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Chen Y, Li Y, Xu Y, Lv Q, Ye Y, Gu J. Revealing the role of natural killer cells in ankylosing spondylitis: identifying diagnostic biomarkers and therapeutic targets. Ann Med 2025; 57:2457523. [PMID: 39853176 PMCID: PMC11770870 DOI: 10.1080/07853890.2025.2457523] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/22/2024] [Revised: 01/06/2025] [Accepted: 01/07/2025] [Indexed: 01/26/2025] Open
Abstract
BACKGROUND Ankylosing spondylitis (AS) is a chronic autoimmune disease that primarily affects the axial joints. Immune cells play a key role in the pathogenesis of AS. This study integrated bioinformatics methods with experimental validation to explore the role of natural killer (NK) cells in AS. METHODS Two microarray datasets, GSE25101 and GSE73754, were selected, and the scRNA-seq data were obtained from GSE194315 and Liu's research. Differentially expressed genes (DEGs) and functional enrichment analysis were performed respectively. Weighted gene co-expression network analysis (WGCNA) was conducted to identify key modules of co-expressed genes and genes involved in NK cell function. The diagnostic value of the identified key genes was evaluated using ROC curves, logistic regression analysis, and a nomogram. Real-time PCR (RT-PCR) was used to quantified the expression of genes. Statistical analysis was conducted using the R software package, and a p-value of less than 0.05 was considered statistically significant. RESULTS Pathways enrichment analysis revealed the involvement of NK cell-mediated immune pathways and regulation of the innate immune response, indicating the crucial role of innate immunity, especially NK cells, in AS pathogenesis. The construction of a co-expression network revealed that the MElightyellow module was most relevant to the NK cell-mediated immune pathway. IL2RB, CD247, PLEKHF1, EOMES, S1PR5, FGFBP2 from the MElightyellow module were identified as key genes involved in NK cell-mediated immune response and served as potential diagnostic biomarkers for AS, with moderate to high diagnostic values based on AUC values. Further analysis using scRNA-seq profiling revealed the higher expression level of IL2RB, CD247, PLEKHF1, S1PR5, FGFBP2 in NK cells compared to that in other cell types. CD247, PLEKHF1, EOMES, S1PR5, and FGFBP2 were reduced expressed in AS patients as compare to control group verified by scRNA-seq data, CD247, EOMES, FGFBP2, IL2RB and S1PR5 were reduced expressed verified by RT-PCR, and PLEKHF1, S1PR5, and FGFBP2 was upregulated after TNF-α blocker therapy. CONCLUSION The study revealed the potential role of NK cells and identified IL2RB, CD247, PLEKHF1, EOMES, S1PR5, and FGFBP2 as key genes associated with NK cells in the pathogenesis of AS.
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Affiliation(s)
- Yuling Chen
- Department of Rheumatology and Immunology, The Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, Guangdong Province, People’s Republic of China
| | - Yan Li
- Department of Scientific Center, The Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, Guangdong Province, People’s Republic of China
| | - Yuan Xu
- Department of Clinical Laboratory, The Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, Guangdong Province, People’s Republic of China
| | - Qing Lv
- Department of Rheumatology and Immunology, The Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, Guangdong Province, People’s Republic of China
| | - Yuanchun Ye
- School of Science, Shenzhen Campus of Sun Yat-sen University, Shenzhen, People’s Republic of China
| | - Jieruo Gu
- Department of Rheumatology and Immunology, The Seventh Affiliated Hospital, Sun Yat-sen University, Shenzhen, Guangdong Province, People’s Republic of China
- Department of Rheumatology and Immunology, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong ProvincePeople’s Republic of China
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2
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Prisinzano M, Seidita I, Bruni P, Petraglia F, Bernacchioni C, Meyer Zu Heringdorf D, Donati C. Characterization of functionally relevant G protein-coupled receptors in endometriotic epithelial cells. Cell Signal 2025; 133:111876. [PMID: 40381972 DOI: 10.1016/j.cellsig.2025.111876] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2025] [Revised: 05/07/2025] [Accepted: 05/15/2025] [Indexed: 05/20/2025]
Abstract
Endometriosis is a chronic inflammatory disease characterized by the invasion of endometrial cells outside the uterine cavity. Current treatments for the disease, whose typical symptoms are pain and infertility, are unsatisfactory, relying on the surgical removal of the lesions and hormonal therapies with high symptom relapse and collateral effects, respectively. The aim of the present study was to exploit the rationale for G protein-coupled receptors (GPCRs) as non-hormonal therapeutic targets for this disease. To this end, human endometriotic epithelial cells 12Z were employed to characterize GPCR-mediated increases in intracellular Ca2+ concentrations ([Ca2+]i) using fluo-4, and cell invasion was measured using Boyden chamber assays. The results showed that the GPCR ligands oxytocin, bradykinin, histamine, lysophosphatidic acid, and sphingosine 1-phosphate (S1P) efficiently increased [Ca2+]i and induced cell invasion in endometriotic cells. In contrast, neuropeptide S, previously identified as a pro-invasive mediator, did not increase [Ca2+]i in 12Z cells. Notably, pretreatment with pertussis toxin significantly reduced S1P-dependent [Ca2+]i increase and cell invasion, highlighting the involvement of Gi-mediated signaling. Employing specific agonists and/or antagonists of S1P receptor isoforms, we demonstrated that S1P1/S1P3/S1P5, but not S1P2/S1P4 mediated the [Ca2+]i increases in this cellular model. Moreover, activation of S1P1/S1P4/S1P5, but not S1P2/S1P3, efficiently stimulated cell invasion. Taken together, we identified several GPCRs that are functionally relevant in human endometriotic epithelial cells and may potentially serve as targets for non-hormonal therapy of endometriosis.
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Affiliation(s)
- Matteo Prisinzano
- Department of Experimental and Clinical Biomedical Sciences "M. Serio", University of Florence, Viale Morgagni 50, 50134 Florence, Italy; Institut für Allgemeine Pharmakologie und Toxikologie, Goethe-Universität Frankfurt, Universitätsklinikum, Theodor-Stern-Kai 7, 60590 Frankfurt am Main, Germany.
| | - Isabelle Seidita
- Department of Experimental and Clinical Biomedical Sciences "M. Serio", University of Florence, Viale Morgagni 50, 50134 Florence, Italy.
| | - Paola Bruni
- Department of Experimental and Clinical Biomedical Sciences "M. Serio", University of Florence, Viale Morgagni 50, 50134 Florence, Italy.
| | - Felice Petraglia
- Department of Experimental and Clinical Biomedical Sciences "M. Serio", University of Florence, Viale Morgagni 50, 50134 Florence, Italy.
| | - Caterina Bernacchioni
- Department of Experimental and Clinical Biomedical Sciences "M. Serio", University of Florence, Viale Morgagni 50, 50134 Florence, Italy.
| | - Dagmar Meyer Zu Heringdorf
- Institut für Allgemeine Pharmakologie und Toxikologie, Goethe-Universität Frankfurt, Universitätsklinikum, Theodor-Stern-Kai 7, 60590 Frankfurt am Main, Germany.
| | - Chiara Donati
- Department of Experimental and Clinical Biomedical Sciences "M. Serio", University of Florence, Viale Morgagni 50, 50134 Florence, Italy.
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3
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Uusi-Mäkelä M, Harjula SKE, Junno M, Sillanpää A, Nätkin R, Niskanen MT, Saralahti AK, Nykter M, Rämet M. The inflammasome adaptor pycard is essential for immunity against Mycobacterium marinum infection in adult zebrafish. Dis Model Mech 2025; 18:dmm052061. [PMID: 39916610 PMCID: PMC11972081 DOI: 10.1242/dmm.052061] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2024] [Accepted: 02/03/2025] [Indexed: 03/25/2025] Open
Abstract
Inflammasomes regulate the host response to intracellular pathogens including mycobacteria. We have previously shown that the course of Mycobacterium marinum infection in adult zebrafish (Danio rerio) mimics the course of tuberculosis in human. To investigate the role of the inflammasome adaptor pycard in zebrafish M. marinum infection, we produced two zebrafish knockout mutant lines for the pycard gene with CRISPR/Cas9 mutagenesis. Although the zebrafish larvae lacking pycard developed normally and had unaltered resistance against M. marinum, the loss of pycard led to impaired survival and increased bacterial burden in the adult zebrafish. Based on histology, immune cell aggregates, granulomas, were larger in pycard-deficient fish than in wild-type controls. Transcriptome analysis with RNA sequencing of a zebrafish haematopoietic tissue, kidney, suggested a role for pycard in neutrophil-mediated defence, haematopoiesis and myelopoiesis during infection. Transcriptome analysis of fluorescently labelled, pycard-deficient kidney neutrophils identified genes that are associated with compromised resistance, supporting the importance of pycard for neutrophil-mediated immunity against M. marinum. Our results indicate that pycard is essential for resistance against mycobacteria in adult zebrafish.
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Affiliation(s)
- Meri Uusi-Mäkelä
- Faculty of Medicine and Health Technology, Tampere University, FI-33014 Tampere, Finland
| | | | - Maiju Junno
- Faculty of Medicine and Health Technology, Tampere University, FI-33014 Tampere, Finland
| | - Alina Sillanpää
- Faculty of Medicine and Health Technology, Tampere University, FI-33014 Tampere, Finland
| | - Reetta Nätkin
- Prostate Cancer Research Center, Faculty of Medicine and Health Technology, Tampere University, FI-33014 Tampere, Finland
- Tays Cancer Center, Tampere University Hospital, FI-33521 Tampere, Finland
| | | | | | - Matti Nykter
- Prostate Cancer Research Center, Faculty of Medicine and Health Technology, Tampere University, FI-33014 Tampere, Finland
- Tays Cancer Center, Tampere University Hospital, FI-33521 Tampere, Finland
| | - Mika Rämet
- Faculty of Medicine and Health Technology, Tampere University, FI-33014 Tampere, Finland
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4
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Nguyen M, Wall BPG, Harrell JC, Dozmorov MG. scHiCcompare: An R Package for Differential Analysis of Single-cell Hi-C Data. J Mol Biol 2025; 437:169155. [PMID: 40246224 PMCID: PMC12145248 DOI: 10.1016/j.jmb.2025.169155] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2024] [Revised: 02/19/2025] [Accepted: 04/09/2025] [Indexed: 04/19/2025]
Abstract
Changes in the three-dimensional (3D) structure of the human genome are associated with various conditions, such as cancer and developmental disorders. Techniques like chromatin conformation capture (Hi-C) have been developed to study these global 3D structures, typically requiring millions of cells and an extremely high sequencing depth (around 1 billion reads per sample) for bulk Hi-C. In contrast, single-cell Hi-C (scHi-C) captures 3D structures at the individual cell level but faces significant data sparsity, characterized by a high proportion of zeros. scHi-C data enable the identification of cell types with distinct 3D structures; consequently, identifying differential chromatin interactions between such groups may offer insights into cell type-specific regulation. While differential analysis methods exist for bulk Hi-C data, they are limited for scHi-C data. To address this, we developed a method for differential scHi-C analysis, extending the HiCcompare R package. Our approach optionally imputes sparse scHi-C data by considering genomic distances and creates pseudo-bulk Hi-C matrices by summing condition-specific data. The data are normalized using locally estimated scatterplot smoothing (LOESS) regression, and differential chromatin interactions are detected via Gaussian Mixture Model (GMM) clustering. Our workflow outperforms existing methods in identifying differential chromatin interactions across various genomic distances, fold changes, resolutions, and sample sizes in both simulated and experimental contexts. This enables the effective detection of cell type-specific differences in chromatin structure and shows expected associations with biological and epigenetic features. Our method is implemented in the scHiCcompare R package, available at https://bioconductor.org/packages/scHiCcompare.
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Affiliation(s)
- My Nguyen
- Department of Biostatistics, Virginia Commonwealth University, Richmond, VA 23298, USA
| | - Brydon P G Wall
- Department of Biostatistics, Virginia Commonwealth University, Richmond, VA 23298, USA
| | - J Chuck Harrell
- Department of Pathology, Virginia Commonwealth University, Richmond, VA 23284, USA; Massey Comprehensive Cancer Center, Virginia Commonwealth University, Richmond, VA 23298, USA
| | - Mikhail G Dozmorov
- Department of Biostatistics, Virginia Commonwealth University, Richmond, VA 23298, USA; Department of Pathology, Virginia Commonwealth University, Richmond, VA 23284, USA.
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5
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Zhang M, Yuan L, Yang X, Zhao X, Xie J, Liu X, Wang F. TRAF1 promotes the progression of Helicobacter pylori-associated gastric cancer through EGFR/STAT/OAS signalling. Life Sci 2025; 373:123656. [PMID: 40287055 DOI: 10.1016/j.lfs.2025.123656] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2025] [Revised: 04/11/2025] [Accepted: 04/21/2025] [Indexed: 04/29/2025]
Abstract
AIMS Helicobacter pylori (H. pylori) is associated with various gastric diseases and is one of the pathogenic factors of gastric cancer (GC). We found that H. pylori induce the expression of TRAF1, but its mechanism of action is still unclear. Therefore, we wanted to determine whether TRAF1 is involved in the mechanism of H. pylori-related GC progression. MATERIALS AND METHODS In this study, we analysed TRAF1 expression and its prognostic significance using clinical specimens, performed functional studies involving TRAF1 overexpression or knockdown in cellular models, identified downstream signalling pathways regulated via RNA-seq, validated these mechanisms through pathway blockade and rescue experiments, and further confirmed the findings in an H. pylori-infected gastritis mouse model. KEY FINDINGS TRAF1 expression was significantly elevated in GC tissues and served as a poor prognostic biomarker. TRAF1 promoted GC cell proliferation, migration and invasion. RNA-seq analysis revealed that TRAF1 activated the EGFR/STAT/OAS signalling axis, upregulated STAT3 expression and increased the transcription of the OAS gene family. Pharmacological inhibition with ruxolitinib and AG490 effectively blocked EGFR/STAT/OAS signalling. In H. pylori-treated cell models, H. pylori infection activated the EGFR/STAT/OAS signalling axis. In vivo, we established an H. pylori-induced gastritis mouse model to validate the activation of this signalling pathway during the gastritis-carcinoma transition. SIGNIFICANCE TRAF1 may promote the proliferation, migration and invasion of H. pylori-associated GC by activating the EGFR/STAT/OAS signalling axis, suggesting that TRAF1 is a promising novel prognostic biomarker and therapeutic target for this malignancy.
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Affiliation(s)
- Minglin Zhang
- Department of Gastroenterology, The Third Xiangya Hospital, Central South University, 138 Tongzipo Road, Changsha, Hunan 410013, China
| | - Lingzhi Yuan
- Department of Digestive Nutrition, Hunan Children's Hospital, Central South University Affiliated Children's Hospital, Changsha, China
| | - Xueer Yang
- Department of Gastroenterology, The Third Xiangya Hospital, Central South University, 138 Tongzipo Road, Changsha, Hunan 410013, China
| | - Xuelin Zhao
- Department of Gastroenterology, The Third Xiangya Hospital, Central South University, 138 Tongzipo Road, Changsha, Hunan 410013, China
| | - Jie Xie
- Department of Gastroenterology, The Third Xiangya Hospital, Central South University, 138 Tongzipo Road, Changsha, Hunan 410013, China
| | - Xiaoming Liu
- Department of Gastroenterology, The Third Xiangya Hospital, Central South University, 138 Tongzipo Road, Changsha, Hunan 410013, China.
| | - Fen Wang
- Department of Gastroenterology, The Third Xiangya Hospital, Central South University, 138 Tongzipo Road, Changsha, Hunan 410013, China.
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6
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Rajebhosale P, Jiang L, Ressa HJ, Johnson KR, Desai NS, Jone A, Role LW, Talmage DA. Diversification of dentate gyrus granule cell subtypes is regulated by Nrg1 nuclear back-signaling. Life Sci Alliance 2025; 8:e202403169. [PMID: 40280713 PMCID: PMC12032840 DOI: 10.26508/lsa.202403169] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2024] [Revised: 04/12/2025] [Accepted: 04/16/2025] [Indexed: 04/29/2025] Open
Abstract
Neuronal heterogeneity is a defining feature of the developing mammalian brain, but the mechanisms regulating the diversification of closely related cell types remain elusive. Here, we investigated granule cell (GC) subtype composition in the dentate gyrus (DG) and the influence of a psychosis-associated V321L mutation in Neuregulin1 (Nrg1). Using morphoelectric characterization, single-nucleus gene expression, and chromatin accessibility profiling, we identified distinctions between typical GCs and a rare subtype known as semilunar granule cells (SGCs). We found that the V321L mutation, which disrupts Nrg1 nuclear back-signaling, results in overabundance of SGC-like cells. Pseudotime analyses suggest a GC-to-SGC transition potential, supported by the accessibility of SGC-enriched genes in non-SGCs. In WT mice, SGC-like gene expression increases during adolescence, coinciding with reduced Nrg1 back-signaling capacity. These results suggest that intact Nrg1 nuclear signaling represses SGC-like fate and that its developmental or pathological loss may permit acquisition of this fate. Our findings reveal a novel role of Nrg1 in maintaining DG cell-type composition and suggest that disrupted subtype regulation may contribute to disease-associated changes in the DG.
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Affiliation(s)
- Prithviraj Rajebhosale
- Genetics of Neuronal Signaling Section, National Institute of Neurological Disorders and Stroke, NIH, Bethesda, MD, USA
| | - Li Jiang
- Genetics of Neuronal Signaling Section, National Institute of Neurological Disorders and Stroke, NIH, Bethesda, MD, USA
| | - Haylee J Ressa
- Undergraduate Program in Fundamental Neuroscience, University of Virginia, Charlottesville, VA, USA
| | - Kory R Johnson
- Bioinformatics Core, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD, USA
| | - Niraj S Desai
- Circuits, Synapses and Molecular Signaling Section, National Institute of Neurological Disorders and Stroke, NIH, Bethesda, MD, USA
| | - Alice Jone
- Program in Neuroscience, State University of New York at Stony Brook, Stony Brook, NY, USA
| | - Lorna W Role
- Circuits, Synapses and Molecular Signaling Section, National Institute of Neurological Disorders and Stroke, NIH, Bethesda, MD, USA
| | - David A Talmage
- Genetics of Neuronal Signaling Section, National Institute of Neurological Disorders and Stroke, NIH, Bethesda, MD, USA
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7
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Huang ZA, Ji GS, Yang S, Yang Y, Wu YC, Tian ZK, Song GS. Design, pharmacology, and toxicology of a novel chemically modified siRNA targeting hepatic angiotensinogen. MOLECULAR THERAPY. NUCLEIC ACIDS 2025; 36:102542. [PMID: 40385635 PMCID: PMC12083919 DOI: 10.1016/j.omtn.2025.102542] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 12/20/2024] [Accepted: 04/14/2025] [Indexed: 05/20/2025]
Abstract
Angiotensinogen (AGT) is the precursor of angiotensin II, a potent vasopressor in the renin-angiotensin-aldosterone system. Small interfering RNAs (siRNAs) targeting hepatic AGT can lower blood pressure in hypertension patients by reducing AGT levels, with effects lasting over 6 months. Existing siRNA molecules are effective, but novel ones with better inhibitory activity and longer duration periods may be developed. In this study, we demonstrated an entire development process for a novel siRNA targeting hepatic AGT. Through the proper combination of bioinformatic on-target/off-target screening on sequences, chemical modification patterns optimization, and liver-targeting delivery ligands conjugation, we have successfully developed several promising siRNAs with equivalent or better inhibitory activity, duration of effect, and safety profile compared with previously reported siRNA. Moreover, our comprehensive analysis has elucidated the correlation between the efficacy and free energy of siRNAs. Currently, there exists no reliable model capable of precisely predicting the activity and off-target risk associated with fully modified siRNAs. Therefore, the implementation of efficient screening procedures is of utmost importance during the development of siRNA candidates. This study presents a meticulous and valuable reference for the development of potent and safe siRNAs on other targets.
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Affiliation(s)
- Ze-Ao Huang
- Beijing Youcare Kechuang Pharmaceutical Technology Co., Ltd., Kechuang 7th Street, BDA, Beijing 100176, China
| | - Guang-Shen Ji
- Beijing Youcare Kechuang Pharmaceutical Technology Co., Ltd., Kechuang 7th Street, BDA, Beijing 100176, China
| | - Shuo Yang
- Beijing Youcare Kechuang Pharmaceutical Technology Co., Ltd., Kechuang 7th Street, BDA, Beijing 100176, China
| | - Yang Yang
- Beijing Youcare Kechuang Pharmaceutical Technology Co., Ltd., Kechuang 7th Street, BDA, Beijing 100176, China
| | - Yu-Cheng Wu
- Beijing Youcare Kechuang Pharmaceutical Technology Co., Ltd., Kechuang 7th Street, BDA, Beijing 100176, China
| | - Zhi-Kang Tian
- Beijing Youcare Kechuang Pharmaceutical Technology Co., Ltd., Kechuang 7th Street, BDA, Beijing 100176, China
| | - Geng-Shen Song
- Beijing Youcare Kechuang Pharmaceutical Technology Co., Ltd., Kechuang 7th Street, BDA, Beijing 100176, China
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8
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Becker SH, Ronayne CE, Bold TD, Jenkins MK. Antigen-specific CD4+ T cells promote monocyte recruitment and differentiation into glycolytic lung macrophages to control Mycobacterium tuberculosis. PLoS Pathog 2025; 21:e1013208. [PMID: 40489538 DOI: 10.1371/journal.ppat.1013208] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2025] [Accepted: 05/12/2025] [Indexed: 06/11/2025] Open
Abstract
Although lung myeloid cells provide an intracellular niche for Mycobacterium tuberculosis (Mtb), CD4+ T cells limit Mtb growth in these cells to protect the host. The CD4+ T cell activities including interferon-γ (IFN-γ) production that account for this protection are poorly understood. Using intravenous antibody labeling and lineage-tracing reporter mice, we show that monocyte-derived macrophages (MDMs), rather than phenotypically similar monocytes or dendritic cells, are preferentially infected with Mtb in murine lungs. MDMs were recruited to the lungs by Mtb-specific CD4+ T cells via IFN-γ, which promoted the extravasation of monocyte precursors from the blood. It was possible that CD4+ T cells recruited infectable MDMs because these cells are uniquely poised to receive cognate MHCII-mediated help to control intracellular bacteria. Mice with MHCII deficiency in monocyte-derived cells had normal Mtb-specific CD4+ T cell activation, expansion and differentiation but the CD4+ T cells were unable to attenuate Mtb growth. Using single cell RNA sequencing, we showed that MDMs receiving cognate MHCII-mediated help from CD4+ T cells upregulated glycolytic genes associated with Mtb control. Overall, the results indicate that CD4+ T cells recruit infectable MDMs to the lungs and then trigger glycolysis-dependent bacterial control in the MDMs by engaging their MHCII-bound Mtb peptides. Moreover, the results suggest that cognate MHCII-mediated help to promote MDM glycolysis is an essential, IFN-γ-independent effector function of Mtb-specific CD4+ T cells.
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Affiliation(s)
- Samuel H Becker
- Center for Immunology, Department of Microbiology and Immunology, University of Minnesota Twin Cities School of Medicine, Minneapolis, Minnesota, United States of America
| | - Christine E Ronayne
- Center for Immunology, Department of Microbiology and Immunology, University of Minnesota Twin Cities School of Medicine, Minneapolis, Minnesota, United States of America
- Division of Infectious Diseases & International Medicine, Department of Medicine, University of Minnesota Twin Cities School of Medicine, Minneapolis, Minnesota, United States of America
| | - Tyler D Bold
- Center for Immunology, Department of Microbiology and Immunology, University of Minnesota Twin Cities School of Medicine, Minneapolis, Minnesota, United States of America
- Division of Infectious Diseases & International Medicine, Department of Medicine, University of Minnesota Twin Cities School of Medicine, Minneapolis, Minnesota, United States of America
| | - Marc K Jenkins
- Center for Immunology, Department of Microbiology and Immunology, University of Minnesota Twin Cities School of Medicine, Minneapolis, Minnesota, United States of America
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9
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Matboli M, Saad M, Ahmed MF, Helmy Hasanin A, Ellithy GM, Abdelwahab MS, Eltantawy EHB, Hamam GG, Hamoud AE, El-Shafei MM, Samir N. Febuxostat alleviate metabolic dysfunction-associated steatohepatitis in rat model via targeting inflammation, cell death, and intestinal barrier dysfunction. Biomed Pharmacother 2025; 187:118086. [PMID: 40306176 DOI: 10.1016/j.biopha.2025.118086] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2025] [Revised: 04/15/2025] [Accepted: 04/23/2025] [Indexed: 05/02/2025] Open
Abstract
Metabolic dysfunction-associated steatotic liver disease (MASLD) constitutes a global health threat with its ability to develop into liver cirrhosis and hepatocellular carcinoma (HCC). Emerging data suggests that oxidative stress and regulated cell death are major driving forces for liver inflammation in MASH. Febuxostat (Feb.), one of the Xanthine oxidase (XO) inhibitors, has shown promise in significantly improving the prognosis of MASH by reducing inflammatory cytokines and cell death. However, the underlying molecular mechanisms remain unclear. In this study, we evaluated the therapeutic effects of febuxostat on MASH through the modulation of cell death, inflammation, and intestinal permeability, focusing on hepatic mRNAs (HGS, SNF8, TSG101) and their epigenetic regulators (rno-miR-6216, rno-miR-1224). MASH was induced in Wistar rats via a High-sucrose high-fat (HSHF) diet over 14 weeks, followed by febuxostat treatment at doses of 1.5, 3, and 6 mg/kg/day for 4 weeks. Febuxostat treatment significantly improved liver function and lipid profiles, reduced hepatic steatosis, intralobular inflammation, and ballooning, and restored normal expression of the hepatic RNA panel by downregulating HGS, SNF8, and TSG101 mRNAs and their epigenetic regulators. Furthermore, febuxostat decreased serum levels of inflammatory (IL6), fibrosis (TGFB1), and cell death (TSG101) markers while reducing apoptosis and regulated cell death via modulation of Caspase-3 and LC3B expression. Improvements in intestinal permeability were evident via reductions in serum haptoglobin (Hpt) and TMAO and restoration of occludin expression. These findings highlight febuxostat as a promising therapeutic candidate for MASH by targeting key molecular mechanisms of liver inflammation and gut-liver axis dysfunction.
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Affiliation(s)
- Marwa Matboli
- Medical biochemistry and molecular biology department, Faculty of Medicine, Ain Shams University, Cairo, Egypt.
| | - Maha Saad
- Basic medical sciences department, Faculty of Medicine, Modern University for Technology and Information, Egypt
| | - Manar Fouad Ahmed
- Medical biochemistry and molecular biology department, Faculty of Medicine, Ain Shams University, Cairo, Egypt
| | - Amany Helmy Hasanin
- Clinical Pharmacology Department, Faculty of Medicine, Ain Shams University, Cairo, Egypt
| | - Ghada M Ellithy
- Clinical Pharmacology Department, Faculty of Medicine, Ain Shams University, Cairo, Egypt
| | | | | | - Ghada Galal Hamam
- Department of Histology, Faculty of Medicine, Ain-Shams University, Cairo, Egypt
| | - Amany E Hamoud
- Anatomy and Embryology Department, Faculty of Medicine, Cairo University, Egypt
| | - Marwa M El-Shafei
- Pathology Department, Faculty of Oral and Dental Medicine, Misr International University, Obour, Qalyubiyya Governorate, Egypt
| | - Nehal Samir
- Medical biochemistry and molecular biology department, Faculty of Medicine, Ain Shams University, Cairo, Egypt
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10
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Cinnamon E, Stein I, Zino E, Rabinovich S, Shovman Y, Schlesinger Y, Salame TM, Reich-Zeliger S, Albrecht T, Roessler S, Schirmacher P, Lotem M, Ben-Neriah Y, Parnas O, Pikarsky E. RORc-expressing immune cells negatively regulate tertiary lymphoid structure formation and support their pro-tumorigenic functions. J Hepatol 2025; 82:1050-1067. [PMID: 39710149 DOI: 10.1016/j.jhep.2024.12.015] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/18/2024] [Revised: 11/28/2024] [Accepted: 12/07/2024] [Indexed: 12/24/2024]
Abstract
BACKGROUND & AIMS RORc-expressing immune cells play important roles in inflammation, autoimmune disease and cancer. They are required for lymphoid organogenesis and have been implicated in tertiary lymphoid structure (TLS) formation. TLSs are formed in many cancer types and have been correlated with better prognosis and response to immunotherapy. In liver cancer, some TLSs are pro-tumorigenic as they harbor tumor progenitor cells and support their growth. The processes involved in TLS development and acquisition of pro- or anti-tumorigenic roles are largely unknown. This study aims to explore the role of RORc-expressing cells in TLS development in the context of inflammation-associated liver cancer. METHODS IKKβ(EE)Hep mice, exhibiting chronic liver inflammation, TLS formation and liver cancer, were crossed with RORc knockout mice to explore RORc's effect on TLS and tumor formation. TLS phenotypes were analyzed using transcriptional, proteomic, and immunohistochemical techniques. CD4, CD8, and B-cell depletions were used to assess their contribution to liver TLS and tumor formation. RESULTS RORc-expressing cells are detected within TLSs of both human patients and mice developing intrahepatic cholangiocarcinoma. In mice, these cells negatively regulate TLS formation, as excess TLSs form in their absence. CD4 cells are essential for liver TLS formation, while B cells are required for TLS formation specifically in the absence of RORc-expressing cells. Importantly, in chronically inflamed livers lacking RORc-expressing cells, TLSs become anti-tumorigenic, reducing tumor load. Anti-tumorigenic TLSs revealed enrichment of exhausted CD8 cells with effector functions, germinal center B cells and plasma cells. B cells are key in limiting tumor development, possibly via tumor-directed antibodies. CONCLUSIONS RORc-expressing cells negatively regulate B-cell responses and facilitate the pro-tumorigenic functions of hepatic TLSs. IMPACT AND IMPLICATIONS RORc-expressing immune cells play critical roles in immune regulation, yet their specific influence on tertiary lymphoid structures (TLSs) in liver pathology and cancer has not been elucidated. Our study reveals that RORc-expressing cells act as negative regulators of TLS formation and shape the immune microenvironment in a manner that promotes tumor development. In the absence of RORc-expressing cells, TLSs not only increase in number but also acquire anti-tumorigenic properties. These findings suggest that RORc-expressing cells serve as key modulators of liver immune dynamics, with potential implications for the use of RORc as a biomarker to differentiate between pro- and anti-tumorigenic immune environments and as a target for manipulating TLS abundance and phenotype in liver cancer.
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Affiliation(s)
- Einat Cinnamon
- The Concern Foundation Laboratories at The Lautenberg Center for Immunology and Cancer Research, Israel-Canada Medical Research Institute, Faculty of Medicine, The Hebrew University, Jerusalem, Israel
| | - Ilan Stein
- The Concern Foundation Laboratories at The Lautenberg Center for Immunology and Cancer Research, Israel-Canada Medical Research Institute, Faculty of Medicine, The Hebrew University, Jerusalem, Israel; Department of Pathology, Hadassah-Hebrew University Medical Center, Jerusalem, Israel
| | - Elvira Zino
- The Concern Foundation Laboratories at The Lautenberg Center for Immunology and Cancer Research, Israel-Canada Medical Research Institute, Faculty of Medicine, The Hebrew University, Jerusalem, Israel
| | - Stav Rabinovich
- The Concern Foundation Laboratories at The Lautenberg Center for Immunology and Cancer Research, Israel-Canada Medical Research Institute, Faculty of Medicine, The Hebrew University, Jerusalem, Israel
| | - Yehuda Shovman
- The Concern Foundation Laboratories at The Lautenberg Center for Immunology and Cancer Research, Israel-Canada Medical Research Institute, Faculty of Medicine, The Hebrew University, Jerusalem, Israel; Department of Neurology, Hadassah-Hebrew University Medical Center, Jerusalem, Israel
| | - Yehuda Schlesinger
- The Concern Foundation Laboratories at The Lautenberg Center for Immunology and Cancer Research, Israel-Canada Medical Research Institute, Faculty of Medicine, The Hebrew University, Jerusalem, Israel
| | - Tomer-Meir Salame
- Flow Cytometry Unit, Life Sciences Core Facilities, Weizmann Institute of Science, Rehovot, Israel
| | | | - Thomas Albrecht
- Institute of Pathology, Heidelberg University Hospital, Heidelberg, Germany
| | - Stephanie Roessler
- Institute of Pathology, Heidelberg University Hospital, Heidelberg, Germany
| | - Peter Schirmacher
- Institute of Pathology, Heidelberg University Hospital, Heidelberg, Germany
| | - Michal Lotem
- Sharett Institute of Oncology, Hadassah-Hebrew University Medical Center, Jerusalem, Israel
| | - Yinon Ben-Neriah
- The Concern Foundation Laboratories at The Lautenberg Center for Immunology and Cancer Research, Israel-Canada Medical Research Institute, Faculty of Medicine, The Hebrew University, Jerusalem, Israel
| | - Oren Parnas
- The Concern Foundation Laboratories at The Lautenberg Center for Immunology and Cancer Research, Israel-Canada Medical Research Institute, Faculty of Medicine, The Hebrew University, Jerusalem, Israel
| | - Eli Pikarsky
- The Concern Foundation Laboratories at The Lautenberg Center for Immunology and Cancer Research, Israel-Canada Medical Research Institute, Faculty of Medicine, The Hebrew University, Jerusalem, Israel; Department of Pathology, Hadassah-Hebrew University Medical Center, Jerusalem, Israel.
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11
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Stapenhorst França F, Gensel JC. Redefining macrophage phenotypes after spinal cord injury: An open data approach. Exp Neurol 2025; 388:115222. [PMID: 40113007 DOI: 10.1016/j.expneurol.2025.115222] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2024] [Revised: 03/13/2025] [Accepted: 03/16/2025] [Indexed: 03/22/2025]
Abstract
Spinal cord injury (SCI) triggers intraspinal inflammation through an influx of blood-derived inflammatory cells such as neutrophils and monocyte-derived macrophages. Macrophages play a complex role in SCI pathophysiology ranging from potentiating secondary injury to facilitating recovery and wound healing. In vitro, macrophages have been classified as having a pro-inflammatory, M1 phenotype, or a regenerative, M2 phenotype. In vivo, however, studies suggest that macrophages exist in a spectrum of phenotypes and can shift from one phenotype to another. Single-cell RNA sequencing (scRNA-seq) allows us to assess immune cell heterogeneity in the spinal cord after injury, and several groups have created publicly available datasets containing valuable data for further exploration. In this study, we compared three different scRNA-seq datasets and analyzed macrophage heterogeneity after SCI based on cell clustering according to gene expression profiles. We analyzed data from 7 days post injury (dpi) in young female mice that received a mid-thoracic SCI contusion. Using the Seurat pipeline, we clustered cells, subsetted macrophages from microglia and other myeloid cells, and identified different macrophage populations. Using SingleR as a cross-dataset cluster comparison tool, we identified similarities in macrophage populations across datasets. To confirm and refine this analysis, we analyzed the top 10 differentially expressed genes for each population in each dataset. Most clusters identified in the SingleR analysis were confirmed to have a unique genetic signature and were consistently present in all datasets analyzed. Taken together, four distinct macrophage populations were consistently identified after SCI at 7 dpi in three datasets from independent research teams. Our identification of biologically conserved macrophage populations after SCI using an unbiased approach highlights the power of data sharing and open data in redefining macrophage heterogeneity.
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Affiliation(s)
- Fernanda Stapenhorst França
- Spinal Cord and Brain Injury Research Center and Department of Physiology, College of Medicine, University of Kentucky, Lexington, KY, United States.
| | - John C Gensel
- Spinal Cord and Brain Injury Research Center and Department of Physiology, College of Medicine, University of Kentucky, Lexington, KY, United States.
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12
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Fullstone T, Rohm H, Kaltofen T, Hierlmayer S, Reichenbach J, Schweikert S, Knodel F, Loeffler AK, Mayr D, Jeschke U, Mahner S, Kessler M, Trillsch F, Rathert P. Identification of FLYWCH1 as a regulator of platinum-resistance in epithelial ovarian cancer. NAR Cancer 2025; 7:zcaf012. [PMID: 40191655 PMCID: PMC11970373 DOI: 10.1093/narcan/zcaf012] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2024] [Revised: 02/20/2025] [Accepted: 03/25/2025] [Indexed: 04/09/2025] Open
Abstract
Platinum-based combination chemotherapy remains the backbone of first-line treatment for patients with advanced epithelial ovarian cancer (EOC). While most patients initially respond well to the treatment, patients with relapse ultimately develop platinum resistance. This study identified FLYWCH-type zinc finger-containing protein 1 (FLYWCH1) as an important regulator in the resistance development process. We showed that the loss of FLYWCH1 promotes platinum resistance in EOC cells, and the low FLYWCH1 expression is correlated with poor prognosis of EOC patients. In platinum-sensitive cells, FLYWCH1 colocalizes with H3K9me3, but this association is significantly reduced when cells acquire resistance. The suppression of FLYWCH1 induces gene expression changes resulting in the deregulation of pathways associated with resistance. In line with its connection to H3K9me3, FLYWCH1 induces gene silencing in a synthetic reporter assay and the suppression of FLYWCH1 alters H3K9me3 at promoter regions and repeat elements. The loss of FLYWCH1 leads to the derepression of LTR and Alu repeats, thereby increasing transcriptional plasticity and driving the resistance development process. Our data highlight the importance of FLYWCH1 in chromatin biology and acquisition of platinum resistance through transcriptional plasticity and propose FLYWCH1 as a potential biomarker for predicting treatment responses in EOC patients.
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MESH Headings
- Female
- Humans
- Drug Resistance, Neoplasm/genetics
- Carcinoma, Ovarian Epithelial/genetics
- Carcinoma, Ovarian Epithelial/drug therapy
- Ovarian Neoplasms/drug therapy
- Ovarian Neoplasms/genetics
- Ovarian Neoplasms/pathology
- Ovarian Neoplasms/metabolism
- Cell Line, Tumor
- Histones/metabolism
- Gene Expression Regulation, Neoplastic/drug effects
- Drosophila Proteins/genetics
- Drosophila Proteins/metabolism
- Neoplasms, Glandular and Epithelial/drug therapy
- Neoplasms, Glandular and Epithelial/genetics
- Neoplasms, Glandular and Epithelial/pathology
- Neoplasms, Glandular and Epithelial/metabolism
- Platinum/pharmacology
- Prognosis
- Promoter Regions, Genetic
- Antineoplastic Agents/pharmacology
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Affiliation(s)
- Tabea L Fullstone
- Department of Molecular Biochemistry, Institute of Biochemistry, University of Stuttgart, 70569 Stuttgart, Germany
| | - Helene Rohm
- Department of Molecular Biochemistry, Institute of Biochemistry, University of Stuttgart, 70569 Stuttgart, Germany
| | - Till Kaltofen
- Department of Obstetrics and Gynaecology, University Hospital, LMU Munich, 81377 Munich, Germany
- Department of Surgery, University Hospital Regensburg, 93053 Regensburg, Germany
| | - Sophia Hierlmayer
- Department of Obstetrics and Gynaecology, University Hospital, LMU Munich, 81377 Munich, Germany
| | - Juliane Reichenbach
- Department of Obstetrics and Gynaecology, University Hospital, LMU Munich, 81377 Munich, Germany
| | - Simon Schweikert
- Department of Molecular Biochemistry, Institute of Biochemistry, University of Stuttgart, 70569 Stuttgart, Germany
| | - Franziska Knodel
- Department of Molecular Biochemistry, Institute of Biochemistry, University of Stuttgart, 70569 Stuttgart, Germany
| | - Ann-Kathrin Loeffler
- Department of Molecular Biochemistry, Institute of Biochemistry, University of Stuttgart, 70569 Stuttgart, Germany
| | - Doris Mayr
- Institute of Pathology, LMU Munich, 81377 Munich, Germany
| | - Udo Jeschke
- Department of Obstetrics and Gynaecology, University Hospital, LMU Munich, 81377 Munich, Germany
- Department of Obstetrics and Gynaecology, University Hospital Augsburg, 86156 Augsburg, Germany
| | - Sven Mahner
- Department of Obstetrics and Gynaecology, University Hospital, LMU Munich, 81377 Munich, Germany
| | - Mirjana Kessler
- Department of Obstetrics and Gynaecology, University Hospital, LMU Munich, 81377 Munich, Germany
| | - Fabian Trillsch
- Department of Obstetrics and Gynaecology, University Hospital, LMU Munich, 81377 Munich, Germany
| | - Philipp Rathert
- Department of Molecular Biochemistry, Institute of Biochemistry, University of Stuttgart, 70569 Stuttgart, Germany
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13
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Ko M, Frankl‐Vilches C, Bakker A, Sohnius‐Wilhelmi N, Alcami P, Gahr M. From silence to song: Testosterone triggers extensive transcriptional changes in the female canary HVC. J Neuroendocrinol 2025; 37:e13476. [PMID: 39647995 PMCID: PMC12145947 DOI: 10.1111/jne.13476] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/21/2024] [Revised: 10/25/2024] [Accepted: 11/16/2024] [Indexed: 12/10/2024]
Abstract
Seasonal song production in canaries is influenced by gonadal hormones, but the molecular mechanisms underlying testosterone-induced song development in adult female canaries, which rarely sing naturally, remain poorly understood. We explored testosterone-induced song development in adult female canaries by comparing gene regulatory networks in the song-controlling brain area HVC at multiple time points (1 h to 14 days) post-treatment with those of placebo-treated controls. Females began vocalizing within 4 days of testosterone treatment, with song complexity and HVC volume increasing progressively over 2 weeks. Rapid transcriptional changes involving 2739 genes preceded song initiation. Over 2 weeks, 9913 genes-approximately 64% of the canary's protein-coding genome-were differentially expressed, with 98% being transiently regulated. These genes are linked to various biological functions, with early changes at the cellular level and later changes affecting the nervous system level after prolonged hormone exposure. Our findings suggest that testosterone-induced song development is accompanied by extensive and dynamic transcriptional changes in the HVC, implicating widespread neuronal involvement. These changes underpin the gradual emergence of singing behavior, providing insights into the neural basis of seasonal behavioral patterns.
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Affiliation(s)
- Meng‐Ching Ko
- Department of Behavioural NeurobiologyMax Planck Institute for Biological IntelligenceSeewiesenGermany
| | - Carolina Frankl‐Vilches
- Department of Behavioural NeurobiologyMax Planck Institute for Biological IntelligenceSeewiesenGermany
| | - Antje Bakker
- Department of Behavioural NeurobiologyMax Planck Institute for Biological IntelligenceSeewiesenGermany
| | - Nina Sohnius‐Wilhelmi
- Department of Behavioural NeurobiologyMax Planck Institute for Biological IntelligenceSeewiesenGermany
| | - Pepe Alcami
- Department of Behavioural NeurobiologyMax Planck Institute for Biological IntelligenceSeewiesenGermany
- Division of Neurobiology, Faculty of BiologyLudwig‐Maximilians‐University MunichPlaneggGermany
| | - Manfred Gahr
- Department of Behavioural NeurobiologyMax Planck Institute for Biological IntelligenceSeewiesenGermany
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14
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Tasneem M, Gupta SD, Ahmed Jony MJ, Minkara M, Dey RK, Ferdoush J. Identification of key biomarker genes in liver hepatocellular carcinoma and kidney renal clear cell carcinoma progression: A shared high-throughput screening and molecular docking method with potentials for targeted therapeutic interventions. J Genet Eng Biotechnol 2025; 23:100497. [PMID: 40390492 PMCID: PMC12049835 DOI: 10.1016/j.jgeb.2025.100497] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2024] [Accepted: 04/14/2025] [Indexed: 05/21/2025]
Abstract
BACKGROUND AND OBJECTIVES Liver Hepatocellular Carcinoma (LIHC) and Kidney Renal Clear Cell Carcinoma (KIRC) are leading causes of cancer death worldwide, but their early detections remain hindered by a lack of genetic markers. Our study aims to find prospective biomarkers that could serve as prognostic indicators for efficient drug candidates for KIRC and LIHC treatment. METHODS To detect differentially expressed genes (DEGs), four datasets were used: GSE66271 and GSE213324 for KIRC, and GSE135631 and GSE202853 for LIHC. Visualization of DEGs was done using heatmaps, volcano plots, and Venn diagrams. Hub genes were identified via PPI analysis and the cytoHubba plugin in Cytoscape. Their expression was evaluated using box plots, stage plots, and survival plots for prognostic assessment via GEPIA2. Molecular docking with PyRx's AutoDock Vina identified optimal binding interactions between compounds and proteins. Pharmacokinetic and toxicity analyses reinforced the drug-likeness and safety of these compounds. RESULTS In this study, 47 DEGs were identified, with the top 10 hub genes being TOP2A, BUB1, PTTG1, CCNB2, NUSAP1, KIF20A, BIRC5, RRM2, NDC80 and CDC45, chosen for their high MCC scores. Data mining revealed a correlation between TOP2A expression and clinical survival outcomes in KIRC and LIHC patients. Docking studies of the TOP2A structure identified a promising compound from Andrographis paniculata with high binding energy and interactions with TOP2A. Pharmacokinetic and toxicity assessments support its potential as a drug candidate. CONCLUSION Our study emphasizes TOP2A's prognostic significance in KIRC and LIHC and recognizes Andrographis paniculata compound as potential therapeutics, offering prospective for enhanced treatment and patient outcomes in these cancers.
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Affiliation(s)
- Maisha Tasneem
- Department of Biotechnology and Genetic Engineering, Noakhali Science and Technology University, Noakhali 3814, Bangladesh
| | - Shipan Das Gupta
- Department of Biotechnology and Genetic Engineering, Noakhali Science and Technology University, Noakhali 3814, Bangladesh
| | - Md Jubair Ahmed Jony
- Department of Biotechnology and Genetic Engineering, Noakhali Science and Technology University, Noakhali 3814, Bangladesh
| | - Maya Minkara
- Department of Biology, Geology, and Environmental Science, University of Tennessee at Chattanooga, 615 McCallie Ave, Chattanooga, TN 37403, USA
| | | | - Jannatul Ferdoush
- Department of Biology, Geology, and Environmental Science, University of Tennessee at Chattanooga, 615 McCallie Ave, Chattanooga, TN 37403, USA.
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15
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Jeong J, Harris J, de Souza LL, Leonelli L. Combining the CowPEAsy Web Application With in Planta Agroinfiltration for Native Promoter Validation in Vigna unguiculata. PLANT, CELL & ENVIRONMENT 2025; 48:4301-4311. [PMID: 39948050 PMCID: PMC12050393 DOI: 10.1111/pce.15431] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/28/2024] [Revised: 01/11/2025] [Accepted: 02/03/2025] [Indexed: 05/06/2025]
Abstract
Cowpea (Vigna unguiculata) is an important protein source in Sub-Saharan Africa. Optimizing resilience and productivity through genetic engineering in cowpea has been slow due in part to a lack of defined species-specific regulatory elements and difficulty testing gene function within the native system. In many plant species, Agrobacterium-mediated transient gene expression is widely used to validate constructs before investing in transgenic lines, but its implementation in legumes has been challenging. In this study, we optimized an in planta agroinfiltration assay in trifoliate cowpea leaves using a betalain reporter. To demonstrate the "intact plant" aspect of this system, we used this assay to characterize drought-inducible promoters by challenging cowpea plants with drought stress. Subsequently, to identify and broaden the pool of native promoters known in cowpea, we developed a user-friendly web application, CowPEAsy, allowing users to interrogate gene expression from our canopy-level, developmental-series RNA-Seq data set. Finally, using CowPEAsy, we identified six promoters that showed constitutive expression across all conditions and verified these promoters with our transient system. This work provides an in vivo platform for preliminary validation of regulatory elements in cowpea and other legumes and enhances current genetic resources by identifying a suite of physiologically relevant promoters of varying strengths.
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Affiliation(s)
- Jooyeon Jeong
- Carl R. Woese Institute for Genomic BiologyUniversity of Illinois at Urbana ChampaignChampaignIllinoisUSA
| | - Jake Harris
- Carl R. Woese Institute for Genomic BiologyUniversity of Illinois at Urbana ChampaignChampaignIllinoisUSA
- Department of Agricultural and Biological EngineeringUniversity of Illinois at Urbana ChampaignChampaignIllinoisUSA
| | - Larissa Larocca de Souza
- Carl R. Woese Institute for Genomic BiologyUniversity of Illinois at Urbana ChampaignChampaignIllinoisUSA
| | - Lauriebeth Leonelli
- Carl R. Woese Institute for Genomic BiologyUniversity of Illinois at Urbana ChampaignChampaignIllinoisUSA
- Department of Agricultural and Biological EngineeringUniversity of Illinois at Urbana ChampaignChampaignIllinoisUSA
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16
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Jiang Y, Immadi MS, Wang D, Zeng S, On Chan Y, Zhou J, Xu D, Joshi T. IRnet: Immunotherapy response prediction using pathway knowledge-informed graph neural network. J Adv Res 2025; 72:319-331. [PMID: 39097091 DOI: 10.1016/j.jare.2024.07.036] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2024] [Revised: 07/10/2024] [Accepted: 07/30/2024] [Indexed: 08/05/2024] Open
Abstract
INTRODUCTION Immune checkpoint inhibitors (ICIs) are potent and precise therapies for various cancer types, significantly improving survival rates in patients who respond positively to them. However, only a minority of patients benefit from ICI treatments. OBJECTIVES Identifying ICI responders before treatment could greatly conserve medical resources, minimize potential drug side effects, and expedite the search for alternative therapies. Our goal is to introduce a novel deep-learning method to predict ICI treatment responses in cancer patients. METHODS The proposed deep-learning framework leverages graph neural network and biological pathway knowledge. We trained and tested our method using ICI-treated patients' data from several clinical trials covering melanoma, gastric cancer, and bladder cancer. RESULTS Our results demonstrate that this predictive model outperforms current state-of-the-art methods and tumor microenvironment-based predictors. Additionally, the model quantifies the importance of pathways, pathway interactions, and genes in its predictions. A web server for IRnet has been developed and deployed, providing broad accessibility to users at https://irnet.missouri.edu. CONCLUSION IRnet is a competitive tool for predicting patient responses to immunotherapy, specifically ICIs. Its interpretability also offers valuable insights into the mechanisms underlying ICI treatments.
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Affiliation(s)
- Yuexu Jiang
- Department of Electrical Engineering and Computer Science, University of Missouri-Columbia, Columbia, MO, USA; Christopher S. Bond Life Sciences Center, University of Missouri-Columbia, Columbia, MO, USA
| | - Manish Sridhar Immadi
- Department of Electrical Engineering and Computer Science, University of Missouri-Columbia, Columbia, MO, USA
| | - Duolin Wang
- Department of Electrical Engineering and Computer Science, University of Missouri-Columbia, Columbia, MO, USA; Christopher S. Bond Life Sciences Center, University of Missouri-Columbia, Columbia, MO, USA
| | - Shuai Zeng
- Department of Electrical Engineering and Computer Science, University of Missouri-Columbia, Columbia, MO, USA; Christopher S. Bond Life Sciences Center, University of Missouri-Columbia, Columbia, MO, USA
| | - Yen On Chan
- Department of Electrical Engineering and Computer Science, University of Missouri-Columbia, Columbia, MO, USA; MU Institute for Data Science and Informatics, University of Missouri-Columbia, Columbia, MO, USA
| | - Jing Zhou
- Department of Surgery, University of Missouri-Columbia, Columbia, MO, USA
| | - Dong Xu
- Department of Electrical Engineering and Computer Science, University of Missouri-Columbia, Columbia, MO, USA; Christopher S. Bond Life Sciences Center, University of Missouri-Columbia, Columbia, MO, USA; MU Institute for Data Science and Informatics, University of Missouri-Columbia, Columbia, MO, USA
| | - Trupti Joshi
- Department of Electrical Engineering and Computer Science, University of Missouri-Columbia, Columbia, MO, USA; Christopher S. Bond Life Sciences Center, University of Missouri-Columbia, Columbia, MO, USA; MU Institute for Data Science and Informatics, University of Missouri-Columbia, Columbia, MO, USA; Department of Biomedical Informatics, Biostatistics and Medical Epidemiology, University of Missouri-Columbia, Columbia, MO, USA.
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17
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Asim MN, Asif T, Hassan F, Dengel A. Protein Sequence Analysis landscape: A Systematic Review of Task Types, Databases, Datasets, Word Embeddings Methods, and Language Models. Database (Oxford) 2025; 2025:baaf027. [PMID: 40448683 DOI: 10.1093/database/baaf027] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2024] [Revised: 02/06/2025] [Accepted: 03/26/2025] [Indexed: 06/02/2025]
Abstract
Protein sequence analysis examines the order of amino acids within protein sequences to unlock diverse types of a wealth of knowledge about biological processes and genetic disorders. It helps in forecasting disease susceptibility by finding unique protein signatures, or biomarkers that are linked to particular disease states. Protein Sequence analysis through wet-lab experiments is expensive, time-consuming and error prone. To facilitate large-scale proteomics sequence analysis, the biological community is striving for utilizing AI competence for transitioning from wet-lab to computer aided applications. However, Proteomics and AI are two distinct fields and development of AI-driven protein sequence analysis applications requires knowledge of both domains. To bridge the gap between both fields, various review articles have been written. However, these articles focus revolves around few individual tasks or specific applications rather than providing a comprehensive overview about wide tasks and applications. Following the need of a comprehensive literature that presents a holistic view of wide array of tasks and applications, contributions of this manuscript are manifold: It bridges the gap between Proteomics and AI fields by presenting a comprehensive array of AI-driven applications for 63 distinct protein sequence analysis tasks. It equips AI researchers by facilitating biological foundations of 63 protein sequence analysis tasks. It enhances development of AI-driven protein sequence analysis applications by providing comprehensive details of 68 protein databases. It presents a rich data landscape, encompassing 627 benchmark datasets of 63 diverse protein sequence analysis tasks. It highlights the utilization of 25 unique word embedding methods and 13 language models in AI-driven protein sequence analysis applications. It accelerates the development of AI-driven applications by facilitating current state-of-the-art performances across 63 protein sequence analysis tasks.
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Affiliation(s)
- Muhammad Nabeel Asim
- German Research Center for Artificial Intelligence, Kaiserslautern 67663, Germany
- Intelligentx GmbH (intelligentx.com), Kaiserslautern, Germany
| | - Tayyaba Asif
- Department of Computer Science, Rheinland Pfälzische Technische Universität, Kaiserslautern 67663, Germany
| | - Faiza Hassan
- Department of Computer Science, Rheinland Pfälzische Technische Universität, Kaiserslautern 67663, Germany
| | - Andreas Dengel
- German Research Center for Artificial Intelligence, Kaiserslautern 67663, Germany
- Department of Computer Science, Rheinland Pfälzische Technische Universität, Kaiserslautern 67663, Germany
- Intelligentx GmbH (intelligentx.com), Kaiserslautern, Germany
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18
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Chumakova A, Vlasov I, Filatova E, Klass A, Lysenko A, Salagaev G, Shadrina M, Slominsky P. Application of RNA-seq for single nucleotide variation identification in a cohort of patients with hypertrophic cardiomyopathy. Sci Rep 2025; 15:18788. [PMID: 40442228 PMCID: PMC12122699 DOI: 10.1038/s41598-025-03226-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/13/2024] [Accepted: 05/19/2025] [Indexed: 06/02/2025] Open
Abstract
A variety of techniques for DNA sequencing, such as specific gene sequencing, whole genome sequencing, or exome sequencing, are currently used to detect single nucleotide variations (SNVs). Although RNA-seq can be used to identify SNVs, studies that employ this approach are uncommon, and those that do often rely on outdated mapping methods or methods that are more suitable for genomic and exomic alignment. In this work, our aim is to apply modern RNA-seq specific alignment method in order to identify SNV in a cohort of HCMP patients, and characterize those SNV to gain insight into possible mechanisms of HCMP pathogenesis. The algorithm of identification of SNV based on transcriptomic sequencing data has been developed and evaluated. The algorithm was evaluated and the optimal quality threshold was determined based on allelic discrimination for the rs397516037 mutation (MYBPC3 c.3697 C > T) among patients. A total of 42,809 SNVs with a quality of 75 or higher were identified in 48 transcriptomes of hypertrophic cardiomyopathy (HCMP) myocardial tissue. Verification of missense and nonsense variants in key HCMP genes using Sanger sequencing confirmed the accuracy of the pipeline results. To identify variants potentially associated with HCMP pathogenesis, a filtration process was conducted based on minor allele frequency, substitution prediction score and ClinVar outcome. 214 missense mutations and 6 nonsense mutations were selected. Together with nonsense mutations, 19 mutations meeting the strictest SIFT and PolypPhen criteria were identified as potential factors influencing HCMP pathogenesis. We have developed and validated a method for identifying SNVs based on transcriptomic data, which can be used to identify putative pathogenic variants. We identified mutations in key HCMP genes MYBPC3 and MYH7 in a cohort of patients. We also found potentially pathologic mutations in genes ANXA6 and FEM1 A and obtained data supporting the role of NEBL in myocardial diseases. This method would be useful in analyzing transcriptomic data available in the Gene Expression Omnibus, but should be used with caution as we have tested it on a specific disease.
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Affiliation(s)
- Anastasia Chumakova
- National Research Centre "Kurchatov Institute", Kurchatov sq. 2, Moscow, 123182, Russia.
| | - Ivan Vlasov
- National Research Centre "Kurchatov Institute", Kurchatov sq. 2, Moscow, 123182, Russia
| | - Elena Filatova
- National Research Centre "Kurchatov Institute", Kurchatov sq. 2, Moscow, 123182, Russia
| | - Anna Klass
- National Research Centre "Kurchatov Institute", Kurchatov sq. 2, Moscow, 123182, Russia
| | - Andrey Lysenko
- Petrovsky National Research Center of Surgery, Abrikosovsky Ln 2, Moscow, 119991, Russia
| | - Gennady Salagaev
- Petrovsky National Research Center of Surgery, Abrikosovsky Ln 2, Moscow, 119991, Russia
| | - Maria Shadrina
- National Research Centre "Kurchatov Institute", Kurchatov sq. 2, Moscow, 123182, Russia
| | - Petr Slominsky
- National Research Centre "Kurchatov Institute", Kurchatov sq. 2, Moscow, 123182, Russia
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19
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Li Y, Jia W, Chen C, Chen C, Chen J, Yang X, Liu P. Identification of biomarkers associated with inflammatory response in Parkinson's disease by bioinformatics and machine learning. PLoS One 2025; 20:e0320257. [PMID: 40435035 PMCID: PMC12118872 DOI: 10.1371/journal.pone.0320257] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2024] [Accepted: 02/14/2025] [Indexed: 06/01/2025] Open
Abstract
Parkinson's disease (PD) is a common and debilitating neurodegenerative disorder. The inflammatory response is essential in the pathogenesis and progression of PD. The goal of this study is to combine bioinformatics and machine learning to screen for biomarker genes related to the inflammatory response in PD. First, differentially expressed genes associated with inflammatory response were screened, PPI networks were constructed and enriched for analysis. LASSO, SVM-RFE and Random Forest algorithms were used to screen biomarker genes. Then, ROC curves were drawn and PD risk predicting models were constructed on the basis of the biomarker genes. Finally, drug sensitivity analysis, mRNA-miRNA network construction and single-cell transcriptome data analysis were performed. The experimental results showed that we screened 31 differentially expressed genes related to inflammatory response. Signaling pathways such as cytokine activity were associated with these genes. Three biomarkers were identified using machine learning algorithms: IL18R1, NMUR1 and RELA. Seventeen co-associated miRNAs were identified by the mRNA-miRNA network as possible regulatory nodes in PD. Finally, these three biomarkers were found to be closely associated with T cells, Endothelial cells, excitatory neurons, inhibitory neurons, and other cells in single-cell transcriptomic analysis. In conclusion, IL18R1, NMUR1 and RELA could be potential therapeutic targets for PD in inflammatory response and new biomarkers for PD diagnosis.
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Affiliation(s)
- Yatan Li
- Department of Pharmacy, the Second Affiliated Hospital of Xinjiang Medical University, Urumqi, China
| | - Wei Jia
- The Affiliated Tumor Hospital of Xinjiang Medical University, Urumqi, China
| | - Chen Chen
- College of Information Science and Engineering, Xinjiang University, Urumqi, China
| | - Cheng Chen
- College of Software, Xinjiang University, Urumqi, China
| | - Jinchao Chen
- College of Information Science and Engineering, Xinjiang University, Urumqi, China
| | - Xinling Yang
- Xinjiang Key Laboratory of Neurology, Urumqi, China
- Xinjiang Medical University, Urumqi, China
| | - Pei Liu
- College of Information Science and Engineering, Xinjiang University, Urumqi, China
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20
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Anuarbekov A, Kléma J. Utilizing RNA-seq data in monotone iterative generalized linear model to elevate prior knowledge quality of the circRNA-miRNA-mRNA regulatory axis. BMC Bioinformatics 2025; 26:139. [PMID: 40426030 PMCID: PMC12117772 DOI: 10.1186/s12859-025-06161-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/25/2025] [Accepted: 05/07/2025] [Indexed: 05/29/2025] Open
Abstract
BACKGROUND Current experimental data on RNA interactions remain limited, particularly for non-coding RNAs, many of which have only recently been discovered and operate within complex regulatory networks. Researchers often rely on in-silico interaction detection algorithms, such as TargetScan, which are based on biochemical sequence alignment. However, these algorithms have limited performance. RNA-seq expression data can provide valuable insights into regulatory networks, especially for understudied interactions such as circRNA-miRNA-mRNA. By integrating RNA-seq data with prior interaction networks obtained experimentally or through in-silico predictions, researchers can discover novel interactions, validate existing ones, and improve interaction prediction accuracy. RESULTS This paper introduces Pi-GMIFS, an extension of the generalized monotone incremental forward stagewise (GMIFS) regression algorithm that incorporates prior knowledge. The algorithm first estimates prior response values through a prior-only regression, interpolates between these prior values and the original data, and then applies the GMIFS method. Our experimental results on circRNA-miRNA-mRNA regulatory interaction networks demonstrate that Pi-GMIFS consistently enhances precision and recall in RNA interaction prediction by leveraging implicit information from bulk RNA-seq expression data, outperforming the initial prior knowledge. CONCLUSION Pi-GMIFS is a robust algorithm for inferring acyclic interaction networks when the variable ordering is known. Its effectiveness was confirmed through extensive experimental validation. We proved that RNA-seq data of a representative size help infer previously unknown interactions available in TarBase v9 and improve the quality of circRNA disease annotation.
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Affiliation(s)
- Alikhan Anuarbekov
- Department of Computer Science, Faculty of Electrical Engineering, Czech Technical University in Prague, Technicka 2, 16627, Prague, Czech Republic
| | - Jiří Kléma
- Department of Computer Science, Faculty of Electrical Engineering, Czech Technical University in Prague, Technicka 2, 16627, Prague, Czech Republic.
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21
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Ulrich L, Schmitz J, Thurow C, Gatz C. CORONATINE INSENSITIVE 1-mediated repression of immunity-related genes in Arabidopsis roots is lifted upon infection with Verticillium longisporum. JOURNAL OF EXPERIMENTAL BOTANY 2025; 76:2356-2372. [PMID: 39945499 DOI: 10.1093/jxb/eraf056] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/03/2024] [Accepted: 02/10/2025] [Indexed: 05/29/2025]
Abstract
Verticillium longisporum is a soil-borne fungal pathogen that causes vascular disease predominantly in Brassicaceae. We have previously reported that the receptor of the plant defense hormone jasmonoyl-isoleucine (JA-Ile), CORONATINE INSENSITIVE 1 (COI1), is required in roots for the fungus to proliferate efficiently in the shoot, suggesting the presence of a mobile root-borne signal that influences the outcome of the disease in shoots. This function of COI1 in promoting susceptibility is independent of JA-Ile. To explore the underlying mechanisms, in this study we compared the root transcriptome of the Arabidopsis coi1 mutant with those of the susceptible JA-Ile-deficient allene oxide synthase (aos) mutant and the susceptible wild-type (WT). The biggest difference between the transcriptomes was due to 316 immunity-related genes that were constitutively higher expressed in coi1 as compared to the susceptible genotypes. Interfering with the expression of a sub-group of these genes partially suppressed the coi1-mediated tolerance phenotype. We therefore hypothesize that secreted defense compounds encoded by genes constitutively expressed in coi1 are transported to the shoot with the transpiration stream where they accumulate and interfere with fungal growth. In addition, we found that 149 of the 316 COI1-repressed genes were induced in the WT and aos upon infection, reaching similar expression levels as in mock-treated coi1. These were not further induced in coi1 upon infection. Thus, the repressive effect of COI1 is either lifted or overridden upon infection with V. longisporum.
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Affiliation(s)
- Louisa Ulrich
- Department of Plant Molecular Biology and Physiology, Albrecht-von-Haller Institute for Plant Sciences, University of Göttingen, Julia-Lermontowa-Weg 3, 37077 Göttingen, Germany
| | - Johanna Schmitz
- Department of Plant Molecular Biology and Physiology, Albrecht-von-Haller Institute for Plant Sciences, University of Göttingen, Julia-Lermontowa-Weg 3, 37077 Göttingen, Germany
| | - Corinna Thurow
- Department of Plant Molecular Biology and Physiology, Albrecht-von-Haller Institute for Plant Sciences, University of Göttingen, Julia-Lermontowa-Weg 3, 37077 Göttingen, Germany
| | - Christiane Gatz
- Department of Plant Molecular Biology and Physiology, Albrecht-von-Haller Institute for Plant Sciences, University of Göttingen, Julia-Lermontowa-Weg 3, 37077 Göttingen, Germany
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22
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Xu Y, Zhang H, Jiao X, Zhang Y, Yin G, Wang C, Du Z, Liang M, Gao X, Gu Z, Jiang Y, Du B, Bi X. Dysregulations of C1QA, C1QB, C1QC and C5AR1 as candidate biomarkers of vascular dementia. NPJ AGING 2025; 11:42. [PMID: 40414977 DOI: 10.1038/s41514-025-00228-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/05/2024] [Accepted: 04/22/2025] [Indexed: 05/27/2025]
Abstract
Vascular dementia (VaD) is the second most common cause of dementia. Few bioinformatic analysis has been done to explore its biomarkers. This study aimed to excavate potential biomarkers for VaD using bioinformatic analysis and validate them at both animal and patient levels. Based on microarray data of GSE122063, bioinformatic analysis revealed 502 DEGs in the frontal and 674 DEGs in the temporal cortex of VaD patients. Afterward, the hub genes between two regions, including C1QA, C1QB, C1QC, and C5AR1, were dugout. Interestingly, compared with sham mice or controls, the above four complements were highly expressed in the cortices of VaD animals and in the peripheral serum of VaD patients. Moreover, receiver operating characteristic curve analysis conformed to good diagnostic powers of these complements, with C1QB having the most prominent capacity (AUC = 0.799, 95%CI 0.722-0.875). That means the complements, especially subunits of C1Q, might be used as specific early VaD diagnostic biomarkers.
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Affiliation(s)
- Yawen Xu
- Department of Neurology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, PR China
- Department of Neurology, Dalian Municipal Central Hospital Affiliated to Dalian University of Technology, Dalian, PR China
| | - Hailing Zhang
- Department of Neurology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, PR China
| | - Xuehao Jiao
- Department of Neurology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, PR China
| | - Yanbo Zhang
- Department of Psychiatry, Faculty of Medicine and Dentistry, University of Alberta, Edmonton, AB, Canada
| | - Ge Yin
- Department of Neurology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, PR China
| | - Cui Wang
- Department of Neurology, Dalian Municipal Central Hospital Affiliated to Dalian University of Technology, Dalian, PR China
| | - Zengkan Du
- Faculty of Basic Medical Sciences, Second Military Medical University, Shanghai, PR China
| | - Meng Liang
- Department of Neurology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, PR China
| | - Xin Gao
- Department of Neurology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, PR China
| | - Zhengsheng Gu
- Department of Neurology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, PR China
| | - Yan Jiang
- School of Pharmacy, Second Military Medical University, Shanghai, PR China
- Department of Oral and Maxillofacial-Head Neck Oncology, Shanghai Ninth People's Hospital, College of Stomatology, Shanghai Jiao Tong University School of Medicine, Shanghai, PR China
| | - Bingying Du
- Department of Neurology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, PR China.
- State Key Laboratory of Medical Neurobiology, Institute for Translational Brain Research, MOE Frontiers Center for Brain Science, MOE Innovative Center for New Drug Development of Immune Inflammatory Disease, Fudan University, Shanghai, PR China.
| | - Xiaoying Bi
- Department of Neurology, Shanghai Changhai Hospital, Second Military Medical University, Shanghai, PR China.
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23
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Huang Y, Yang T, Yang C, Tang B, Su B, Yang X. MiR-485-3p/MELK cascade mediates tumor progression in pancreatic cancer. Sci Rep 2025; 15:17870. [PMID: 40404813 PMCID: PMC12098867 DOI: 10.1038/s41598-025-02586-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/28/2024] [Accepted: 05/14/2025] [Indexed: 05/24/2025] Open
Abstract
Pancreatic cancer remains one of the leading causes of mortality worldwide, largely due to the limitations of current clinical strategies for its treatment. As a result, identifying genetic alterations and potential therapeutic targets could offer new opportunities for improving the diagnosis and treatment of pancreatic cancer. The identification of differentially expressed genes (DEGs) and subsequent analyses, including signaling pathway enrichment, functional classification, and protein-protein interaction (PPI) network construction, were conducted using three public datasets: GSE32676, GSE71989, and GSE16515. Kaplan-Meier survival curves and receiver operating characteristic (ROC) curves were employed to investigate the correlation between hub genes and clinicopathological features in pancreatic cancer patients. Genetic alterations were analyzed using the CBioPortal web tool. Cell proliferation was assessed through CCK-8, colony formation, and EdU assays. Tumor migration, invasion, and angiogenesis were evaluated using transwell and tube formation assays, respectively. Protein and mRNA expression levels were measured via western blot analysis and qPCR assays. The subcutaneous xenografted nude mice models were generated to evaluate the potential effect of miR-485-3p/MELK cascade on tumor growth in vivo. Our analysis revealed that MELK expression is positively correlated with poor prognosis in patients with pancreatic cancer. The overexpression or knockdown of MELK significantly influences cell proliferation, tumor metastasis, and angiogenesis across various pancreatic cancer cell lines. Furthermore, we identified that miR-485-3p regulates MELK expression by directly targeting the MELK 3'UTR binding site in pancreatic cancer cells, which subsequently impacts tumor progression. Additionally, our findings demonstrate that the miR-485-3p/MELK cascade is closely associated with tumor progression in pancreatic cancer cells. Mechanistically, the miR-485-3p/MELK cascade promotes the phosphorylation of Akt to regulate pancreatic cancer cell progression, metastasis, and angiogenesis. Furthermore, overexpression of miR-485-3p inhibits the tumor growth induced by MELK overexpression in subcutaneous xenograft model. MiR-485-3p/MELK cascade may serve as a promising biomarker and therapeutic target for the diagnosis and treatment of pancreatic cancer.
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Affiliation(s)
- Yishan Huang
- Engineering Research Center of Key Technique for Biotherapy of Guangdong Province, Shantou University Medical College, Shantou, 515041, China
| | - Ting Yang
- Engineering Research Center of Key Technique for Biotherapy of Guangdong Province, Shantou University Medical College, Shantou, 515041, China
| | - Chen Yang
- Engineering Research Center of Key Technique for Biotherapy of Guangdong Province, Shantou University Medical College, Shantou, 515041, China
| | - Bo Tang
- Department of Hepatobiliary Surgery, The First Affiliated Hospital of Guangxi Medical University, Nanning, 530021, China
| | - Bo Su
- Department of Cell Biology, School of Basic Medical Science, Shandong University, Jinan, 250012, China.
| | - Xiaojun Yang
- Engineering Research Center of Key Technique for Biotherapy of Guangdong Province, Shantou University Medical College, Shantou, 515041, China.
- The First Dongguan Affiliated Hospital, Guangdong Provincial Key Laboratory of Medical Molecular Diagnostics, School of Basic Medicine, Guangdong Medical University, Dongguan, 523808, China.
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24
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Pepler MAD, Mulholland EL, Montague FR, Elliot MA. Defining the networks that connect RNase III and RNase J-mediated regulation of primary and specialized metabolism in Streptomyces venezuelae. J Bacteriol 2025; 207:e0002425. [PMID: 40227046 PMCID: PMC12096830 DOI: 10.1128/jb.00024-25] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/19/2025] [Accepted: 03/18/2025] [Indexed: 04/15/2025] Open
Abstract
RNA metabolism involves coordinating RNA synthesis with RNA processing and degradation. Ribonucleases play fundamental roles within the cell, contributing to the cleavage, modification, and degradation of RNA molecules, with these actions ensuring appropriate gene regulation and cellular homeostasis. Here, we employed RNA sequencing to explore the impact of RNase III and RNase J on the transcriptome of Streptomyces venezuelae. Differential expression analysis comparing wild-type and RNase mutant strains at distinct developmental stages revealed significant changes in transcript abundance, particularly in pathways related to multicellular development, nutrient acquisition, and specialized metabolism. Both RNase mutants exhibited dysregulation of the BldD regulon, including altered expression of many cyclic-di-GMP-associated enzymes. We also observed precocious chloramphenicol production in these RNase mutants and found that in the RNase III mutant, this was associated with PhoP-mediated regulation. We further found that RNase III directly targeted members of the PhoP regulon, suggesting a link between RNA metabolism and a regulator that bridges primary and specialized metabolism. We connected RNase J function with translation through the observation that RNase J directly targets multiple ribosomal protein transcripts for degradation. These findings establish distinct but complementary roles for RNase III and RNase J in coordinating the gene expression dynamics critical for S. venezuelae development and specialized metabolism. IMPORTANCE RNA processing and metabolism are mediated by ribonucleases and are fundamental processes in all cells. In the morphologically complex and metabolically sophisticated Streptomyces bacteria, RNase III and RNase J influence both development and metabolism through poorly understood mechanisms. Here, we show that both ribonucleases are required for the proper expression of the BldD developmental pathway and contribute to the control of chloramphenicol production, with an interesting connection to phosphate regulation for RNase III. Additionally, we show that both RNases have the potential to impact translation through distinct mechanisms and can function cooperatively in degrading specific transcripts. This study advances our understanding of RNases in Streptomyces biology by providing insight into distinct contributions made by these enzymes and the intriguing interplay between them.
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Affiliation(s)
- Meghan A. D. Pepler
- Department of Biology, McMaster University, Hamilton, Ontario, Canada
- Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario, Canada
| | - Emma L. Mulholland
- Department of Biology, McMaster University, Hamilton, Ontario, Canada
- Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario, Canada
| | - Freddie R. Montague
- Department of Biology, McMaster University, Hamilton, Ontario, Canada
- Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario, Canada
| | - Marie A. Elliot
- Department of Biology, McMaster University, Hamilton, Ontario, Canada
- Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario, Canada
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25
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Wilcken S, Koutsandrea PH, Bakker T, Kulik A, Orthwein T, Franz-Wachtel M, Harbig T, Nieselt KK, Forchhammer K, Brötz-Oesterhelt H, Macek B, Mordhorst S, Kaysser L, Gust B. The TetR-like regulator Sco4385 and Crp-like regulator Sco3571 modulate heterologous production of antibiotics in Streptomyces coelicolor M512. Appl Environ Microbiol 2025; 91:e0231524. [PMID: 40183567 DOI: 10.1128/aem.02315-24] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2024] [Accepted: 03/09/2025] [Indexed: 04/05/2025] Open
Abstract
Heterologous expression in well-studied model strains is a routinely applied method to investigate biosynthetic pathways. Here, we pursue a comparative approach of large-scale DNA-affinity-capturing assays (DACAs) coupled with semi-quantitative mass spectrometry (MS) to identify putative regulatory proteins from Streptomyces coelicolor M512, which bind to the heterologously expressed biosynthetic gene clusters (BGCs) of the liponucleoside antibiotics caprazamycin and liposidomycin. Both gene clusters share an almost identical genetic arrangement, including the location of promoter regions, as detected by RNA sequencing. A total of 2,214 proteins were trapped at the predicted promoter regions, with only three binding to corresponding promoters in both gene clusters. Among these, the overexpression of a yet uncharacterized TetR-family regulator (TFR), Sco4385, increased caprazamycin but not liposidomycin production. Protein-DNA interaction experiments using biolayer interferometry confirmed the binding of Sco4385 to Pcpz10 and PlpmH at different locations within both promoter regions, which might explain its functional variance. Sequence alignment allowed the determination of a consensus sequence present in both promoter regions, to which Sco4385 was experimentally shown to bind. Furthermore, we found that the overexpression of the Crp regulator, Sco3571, leads to a threefold increase in caprazamycin and liposidomycin production yields, possibly due to an increased expression of a precursor pathway.IMPORTANCEStreptomycetes are well-studied model organisms for the biosynthesis of pharmaceutically, industrially, and biotechnologically valuable metabolites. Their naturally broad repertoire of natural products can be further exploited by heterologous expression of biosynthetic gene clusters (BGCs) in non-native host strains. This approach forces the host to adapt to a new regulatory and metabolic environment. In our study, we demonstrate that a host regulator not only interacts with newly incorporated gene clusters but also regulates precursor supply for the produced compounds. We present a comprehensive study of regulatory proteins that interact with two genetically similar gene clusters for the biosynthesis of liponucleoside antibiotics. Thereby, we identified regulators of the heterologous host that influence the production of the corresponding antibiotic. Surprisingly, the regulatory interaction is highly specific for each biosynthetic gene cluster, even though they encode largely structurally similar metabolites.
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Affiliation(s)
- Sarah Wilcken
- Pharmaceutical Biology, Pharmaceutical Institute, Eberhard-Karls-University Tübingen, Tübingen, Germany
- Partner Site Tübingen, German Centre for Infection Research (DZIF), Tübingen, Germany
| | | | - Tomke Bakker
- Pharmaceutical Biology, Pharmaceutical Institute, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Andreas Kulik
- Department of Microbial Bioactive Compounds, Interfaculty Institute of Microbiology and Infection Medicine, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Tim Orthwein
- Department of Microbiology and Organismic Interactions, Interfaculty Institute of Microbiology and Infection Medicine, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Mirita Franz-Wachtel
- Proteome Center Tübingen, Institute of Cell Biology, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Theresa Harbig
- Interfaculty Institute for Bioinformatics and Medical Informatics, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Kay Katja Nieselt
- Interfaculty Institute for Bioinformatics and Medical Informatics, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Karl Forchhammer
- Department of Microbiology and Organismic Interactions, Interfaculty Institute of Microbiology and Infection Medicine, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Heike Brötz-Oesterhelt
- Partner Site Tübingen, German Centre for Infection Research (DZIF), Tübingen, Germany
- Department of Microbial Bioactive Compounds, Interfaculty Institute of Microbiology and Infection Medicine, Eberhard-Karls-University Tübingen, Tübingen, Germany
- Cluster of Excellence Controlling Microbes to Fight Infections, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Boris Macek
- Proteome Center Tübingen, Institute of Cell Biology, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Silja Mordhorst
- Pharmaceutical Biology, Pharmaceutical Institute, Eberhard-Karls-University Tübingen, Tübingen, Germany
| | - Leonard Kaysser
- Institute for Drug Discovery, Department of Pharmaceutical Biology, Leipzig University, Leipzig, Germany
| | - Bertolt Gust
- Pharmaceutical Biology, Pharmaceutical Institute, Eberhard-Karls-University Tübingen, Tübingen, Germany
- Partner Site Tübingen, German Centre for Infection Research (DZIF), Tübingen, Germany
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26
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Grossi E, Marchese FP, González J, Goñi E, Fernández-Justel JM, Amadoz A, Herranz N, Puchades-Carrasco L, Montes M, Huarte M. A lncRNA-mediated metabolic rewiring of cell senescence. Cell Rep 2025; 44:115747. [PMID: 40408249 DOI: 10.1016/j.celrep.2025.115747] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/24/2023] [Revised: 03/21/2025] [Accepted: 05/06/2025] [Indexed: 05/25/2025] Open
Abstract
Despite not proliferating, senescent cells remain metabolically active to maintain the senescence program. However, the mechanisms behind this metabolic reprogramming are not well understood. We identify senescence-induced long noncoding RNA (sin-lncRNA), a previously uncharacterized long noncoding RNA (lncRNA), a key player in this response. While strongly activated in senescence by C/EBPβ, sin-lncRNA loss reinforces the senescence program by altering oxidative phosphorylation and rewiring mitochondrial metabolism. By interacting with dihydrolipoamide S-succinyltransferase (DLST), it facilitates its mitochondrial localization. Depletion of sin-lncRNA causes DLST nuclear translocation, leading to transcriptional changes in oxidative phosphorylation (OXPHOS) genes. While not expressed in highly proliferative cancer cells, it is strongly induced upon cisplatin-induced senescence. Depletion of sin-lncRNA in ovarian cancer cells reduces oxygen consumption and increases extracellular acidification, sensitizing cells to cisplatin treatment. Altogether, these results indicate that sin-lncRNA is specifically induced in senescence to maintain metabolic homeostasis, unveiling an RNA-dependent metabolic rewiring specific to senescent cells.
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Affiliation(s)
- Elena Grossi
- Center for Applied Medical Research, University of Navarra, Pamplona, Spain; Institute of Health Research of Navarra (IdiSNA), Pamplona, Spain
| | - Francesco P Marchese
- Center for Applied Medical Research, University of Navarra, Pamplona, Spain; Institute of Health Research of Navarra (IdiSNA), Pamplona, Spain
| | - Jovanna González
- Center for Applied Medical Research, University of Navarra, Pamplona, Spain; Institute of Health Research of Navarra (IdiSNA), Pamplona, Spain
| | - Enrique Goñi
- Center for Applied Medical Research, University of Navarra, Pamplona, Spain; Institute of Health Research of Navarra (IdiSNA), Pamplona, Spain
| | - José Miguel Fernández-Justel
- Center for Applied Medical Research, University of Navarra, Pamplona, Spain; Institute of Health Research of Navarra (IdiSNA), Pamplona, Spain
| | - Alicia Amadoz
- Center for Applied Medical Research, University of Navarra, Pamplona, Spain; Institute of Health Research of Navarra (IdiSNA), Pamplona, Spain
| | - Nicolás Herranz
- Vall d'Hebron Institute of Oncology (VHIO), Barcelona, Spain; Vall d'Hebron Institute of Research (VHIR), Barcelona, Spain
| | - Leonor Puchades-Carrasco
- Drug Discovery Unit, Instituto de Investigación Sanitaria La Fe (IISLAFE), 46026 Valencia, Spain
| | - Marta Montes
- Center for Applied Medical Research, University of Navarra, Pamplona, Spain; Institute of Health Research of Navarra (IdiSNA), Pamplona, Spain.
| | - Maite Huarte
- Center for Applied Medical Research, University of Navarra, Pamplona, Spain; Institute of Health Research of Navarra (IdiSNA), Pamplona, Spain.
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27
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Bharati J, Kumar S, Buragohain BM, Das D, Devi SJ, Mohan NH, Gupta VK. Identification of differentially expressed genes and pathways in the post-ovulatory ampulla of cyclic pigs through a transcriptomics approach. Mol Biol Rep 2025; 52:481. [PMID: 40397228 DOI: 10.1007/s11033-025-10605-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2025] [Accepted: 05/13/2025] [Indexed: 05/22/2025]
Abstract
BACKGROUND Information on global transcriptomic changes in the porcine ampulla after ovulation is crucial for understanding of oviductal physiology at the molecular level. The objective of the present study was to investigate the differentially expressed genes (DEGs) and signalling pathways regulating the functionality of ampulla in pigs post-ovulation. METHODS AND RESULTS The RNA-sequencing of the post-ovulatory ampulla (POA) and early luteal ampulla (ELA) tissues was conducted using Illumina NextSeq2000. The R package NOISeq was used to obtain significantly differentially expressed genes (DEGs) with the probability of differential expression (1-FDR) value ≥ 0.95 and log2 fold change (log2FC) ≥ 1, which revealed 817 DEGs (657 up- and 160 down-regulated) in the POA vs. ELA group comparison. These DEGs were functionally annotated with various gene ontology terms like sterol biosynthetic process, growth, cell migration, and Reactome pathways like signal transduction, metabolism, and cell cycle, indicating key role of these molecular events in POA. The WNT, TNFR2 non-canonical NF-kB, and hedgehog signalling pathways along with the activation of the immune system process, were enriched in the POA vs. ELA group, which indicates their role in cell-cell interactions and cell fate determination in remodelling the oviductal microenvironment during transition from estrogen to progesterone domination. The highly connected upregulated hub genes ESR1, RAD51, YARS1, TYMS and CDK2 can be regarded as key regulatory factors in synchronizing the changes in POA at the molecular level in the oviduct. CONCLUSION The present study revealed several DEGs, signalling pathways and novel modulatory factors associated with the ampullary physiology during early embryonic development in the POA, which may influence fertility and litter size in pigs.
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Affiliation(s)
- Jaya Bharati
- Animal Physiology, ICAR-National Research Centre on Pig, Guwahati, 781131, Assam, India.
| | - Satish Kumar
- Animal Genetics and Breeding, ICAR-National Research Centre on Pig, Guwahati, 781131, Assam, India.
| | | | - Diptesh Das
- Animal Physiology, ICAR-National Research Centre on Pig, Guwahati, 781131, Assam, India
| | - Salam Jayachitra Devi
- Computer Applications and Information Technology, ICAR-National Research Centre on Pig, Guwahati, 781131, Assam, India
| | - N H Mohan
- Animal Physiology, ICAR-National Research Centre on Pig, Guwahati, 781131, Assam, India
| | - Vivek Kumar Gupta
- Director, ICAR-National Research Centre on Pig, Guwahati, 781131, Assam, India
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28
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Boshnakovska A, Pronto JR, Gall T, Aich A, Prochazka J, Nichtova Z, Sedlacek R, Sobitov I, Ainatzi S, Lenz C, Katschinski DM, Urlaub H, Voigt N, Rehling P, Kremer LS. SMIM20 promotes complex IV biogenesis and Ca 2+ signaling in mice heart. Cell Rep 2025; 44:115723. [PMID: 40402744 DOI: 10.1016/j.celrep.2025.115723] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2024] [Revised: 03/18/2025] [Accepted: 04/29/2025] [Indexed: 05/24/2025] Open
Abstract
Mitochondria are key to cellular energetics, metabolism, and signaling. Their dysfunction is linked to devastating diseases, including mitochondrial disorders, diabetes, neurodegenerative diseases, cardiac disorders, and cancer. Here, we present a knockout mouse model lacking the complex IV assembly factor SMIM20/MITRAC7. SMIM20-/- mice display cardiac pathology with reduced heart weight and cardiac output. Heart mitochondria present with reduced levels of complex IV associated with increased complex I activity, have altered fatty acid oxidation, and display elevated levels of ROS production. Interestingly, mutant mouse ventricular myocytes show unphysiological Ca2+ handling, which can be attributed to the increase in mitochondrial ROS production. Our study presents an example of a tissue-specific phenotype in the context of OXPHOS dysfunction. Moreover, our data suggest a link between complex IV dysfunction and Ca2+ handling at the endoplasmic reticulum through ROS signaling.
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Affiliation(s)
- Angela Boshnakovska
- Department of Cellular Biochemistry, University Medical Center Göttingen, 37073 Göttingen, Germany; German Center for Child and Adolescent Health (DZKJ), 37075 Göttingen, Germany
| | - Julius Ryan Pronto
- Institute of Pharmacology and Toxicology, University Medical Center Göttingen, Georg August University Göttingen, 37075 Göttingen, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Göttingen, 37075 Göttingen, Germany
| | - Tanja Gall
- Department of Cellular Biochemistry, University Medical Center Göttingen, 37073 Göttingen, Germany
| | - Abhishek Aich
- Department of Cellular Biochemistry, University Medical Center Göttingen, 37073 Göttingen, Germany
| | - Jan Prochazka
- Czech Centre for Phenogenomics, Institute of Molecular Genetics of the CAS, 142 20 Prague, Czech Republic
| | - Zuzana Nichtova
- Czech Centre for Phenogenomics, Institute of Molecular Genetics of the CAS, 142 20 Prague, Czech Republic
| | - Radislav Sedlacek
- Czech Centre for Phenogenomics, Institute of Molecular Genetics of the CAS, 142 20 Prague, Czech Republic
| | - Izzatullo Sobitov
- Institute of Pharmacology and Toxicology, University Medical Center Göttingen, Georg August University Göttingen, 37075 Göttingen, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Göttingen, 37075 Göttingen, Germany; Cluster of Excellence "Multiscale Bioimaging: From Molecular Machines to Networks of Excitable Cells" (MBExC), University of Göttingen, 37075 Göttingen, Germany
| | - Sofia Ainatzi
- Bioanalytical Mass Spectrometry Group, Max Planck Institute for Multidisciplinary Sciences, 37077 Göttingen, Germany
| | - Christof Lenz
- Bioanalytical Mass Spectrometry Group, Max Planck Institute for Multidisciplinary Sciences, 37077 Göttingen, Germany; Department of Clinical Chemistry, University Medical Center Göttingen, 37075 Göttingen, Germany
| | - Dörthe M Katschinski
- Department of Cardiovascular Physiology, University Medical Center Göttingen, 37073 Göttingen, Germany
| | - Henning Urlaub
- Bioanalytical Mass Spectrometry Group, Max Planck Institute for Multidisciplinary Sciences, 37077 Göttingen, Germany; Department of Clinical Chemistry, University Medical Center Göttingen, 37075 Göttingen, Germany
| | - Niels Voigt
- Institute of Pharmacology and Toxicology, University Medical Center Göttingen, Georg August University Göttingen, 37075 Göttingen, Germany; German Center for Cardiovascular Research (DZHK), Partner Site Göttingen, 37075 Göttingen, Germany; Cluster of Excellence "Multiscale Bioimaging: From Molecular Machines to Networks of Excitable Cells" (MBExC), University of Göttingen, 37075 Göttingen, Germany
| | - Peter Rehling
- Department of Cellular Biochemistry, University Medical Center Göttingen, 37073 Göttingen, Germany; German Center for Child and Adolescent Health (DZKJ), 37075 Göttingen, Germany; Cluster of Excellence "Multiscale Bioimaging: From Molecular Machines to Networks of Excitable Cells" (MBExC), University of Göttingen, 37075 Göttingen, Germany; Max Planck Institute for Multidisciplinary Science, 37077 Göttingen, Germany; Fraunhofer Institute for Translational Medicine and Pharmacology, Translational Neuroinflammation and Automated Microscopy, 37075 Göttingen, Germany.
| | - Laura S Kremer
- Department of Cellular Biochemistry, University Medical Center Göttingen, 37073 Göttingen, Germany.
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29
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Auer F, Morcos MNF, Sipola M, Akhtar I, Moisio S, Vogt J, Haag R, Lahnalampi M, Tuononen TJ, Hanel A, Viitasalo A, Friedrich UA, Dahl A, Prexler C, Pandyra AA, Stepensky P, Takagi M, Borkhardt A, Heinäniemi M, Hauer J. Trajectories from single-cells to PAX5-driven leukemia reveal PAX5-MYC interplay in vivo. Leukemia 2025:10.1038/s41375-025-02626-2. [PMID: 40394211 DOI: 10.1038/s41375-025-02626-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2024] [Revised: 03/31/2025] [Accepted: 04/22/2025] [Indexed: 05/22/2025]
Abstract
PAX5 acts as a master regulator of B-cell proliferation and differentiation. Its germline and somatic deregulation have both been implicated in the development of B-cell precursor acute lymphoblastic leukemia (BCP-ALL). However, the process how reduced PAX5 transcriptional activity mediates progression to BCP-ALL, is still poorly understood. Here, we characterized the longitudinal effects of PAX5 reduction on healthy, pre-leukemic and BCP-ALL cells at the single-cell level. Cell-surface marker analysis revealed a genotype-driven enrichment of the pre-BII population in healthy Pax5± mice. This population showed downregulated B-cell receptor signaling, while DNA replication/repair and cell-cycle signaling pathways were upregulated. Moreover, we observed a shift in the kappa/lambda light chain ratio toward lambda rearranged B-cells. Transplantation experiments further validated a delay of Pax5± pre-BII cells in maturation and transition to IgM-positivity. Additionally, single-cell RNA-Sequencing and bulk ATAC-Sequencing of different stages of BCP-ALL evolution showed that Pax5± pre-leukemic cells lose their B-cell identity and display Myc activation. Subsequently, BCP-ALLs acquired additional RAG-mediated aberrations and driver mutations in JAK-STAT and RAS-signaling pathways. Together, this study elucidates molecular and functional checkpoints in PAX5-mediated pre-leukemic cell progression exploitable for therapeutic intervention and demonstrates that PAX5 reduction is sufficient to initiate clonal evolution to BCP-ALL through activation of MYC.
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Affiliation(s)
- Franziska Auer
- Technical University of Munich, Germany; School of Medicine and Health; Department of Pediatrics, Munich, Germany
| | - Mina N F Morcos
- Technical University of Munich, Germany; School of Medicine and Health; Department of Pediatrics, Munich, Germany
| | - Mikko Sipola
- Institute of Biomedicine, School of Medicine, University of Eastern Finland, Yliopistonranta 8, FI-70211, Kuopio, Finland
| | - Irfan Akhtar
- Technical University of Munich, Germany; School of Medicine and Health; Department of Pediatrics, Munich, Germany
| | - Sanni Moisio
- Institute of Biomedicine, School of Medicine, University of Eastern Finland, Yliopistonranta 8, FI-70211, Kuopio, Finland
| | - Julia Vogt
- Technical University of Munich, Germany; School of Medicine and Health; Department of Pediatrics, Munich, Germany
| | - Rebecca Haag
- Technical University of Munich, Germany; School of Medicine and Health; Department of Pediatrics, Munich, Germany
| | - Mari Lahnalampi
- Institute of Biomedicine, School of Medicine, University of Eastern Finland, Yliopistonranta 8, FI-70211, Kuopio, Finland
| | - Tiina J Tuononen
- Institute of Biomedicine, School of Medicine, University of Eastern Finland, Yliopistonranta 8, FI-70211, Kuopio, Finland
| | - Andrea Hanel
- Institute of Biomedicine, School of Medicine, University of Eastern Finland, Yliopistonranta 8, FI-70211, Kuopio, Finland
| | - Anna Viitasalo
- Institute of Biomedicine, School of Medicine, University of Eastern Finland, Yliopistonranta 8, FI-70211, Kuopio, Finland
| | - Ulrike A Friedrich
- DRESDEN-concept Genome Center, Technology Platform at the Center for Molecular and Cellular Bioengineering (CMCB), Dresden University of Technology (TUD), Dresden, Germany
- German Center for Diabetes Research (DZD e.V.), 85764, Neuherberg, Germany
- Paul Langerhans Institute Dresden of the Helmholtz Center Munich, University Hospital and Faculty of Medicine Carl Gustav Carus, Dresden University of Technology (TUD), Dresden, Germany
| | - Andreas Dahl
- DRESDEN-concept Genome Center, Technology Platform at the Center for Molecular and Cellular Bioengineering (CMCB), Dresden University of Technology (TUD), Dresden, Germany
| | - Carolin Prexler
- Technical University of Munich, Germany; School of Medicine and Health; Department of Pediatrics, Munich, Germany
| | - Aleksandra A Pandyra
- Institute of Clinical Chemistry and Clinical Pharmacology, University Hospital Bonn, Venusberg-Campus 1, 53127, Bonn, Germany
- German Center for Infection Research (DZIF), Partner Site Bonn-Cologne, Bonn, Germany
- Department of Pediatric Oncology, Hematology and Clinical Immunology, Heinrich-Heine University Duesseldorf, Medical Faculty, Duesseldorf, Germany
| | - Polina Stepensky
- Department of Bone Marrow Transplantation and Cancer Immunotherapy, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel
| | - Masatoshi Takagi
- Department of Pediatrics and Developmental Biology, Institute of Science Tokyo, Tokyo, Japan
| | - Arndt Borkhardt
- Department of Pediatric Oncology, Hematology and Clinical Immunology, Heinrich-Heine University Duesseldorf, Medical Faculty, Duesseldorf, Germany
| | - Merja Heinäniemi
- Institute of Biomedicine, School of Medicine, University of Eastern Finland, Yliopistonranta 8, FI-70211, Kuopio, Finland.
| | - Julia Hauer
- Technical University of Munich, Germany; School of Medicine and Health; Department of Pediatrics, Munich, Germany.
- German Cancer Consortium (DKTK), München, Germany.
- Pediatric Hematology and Oncology, Department of Pediatrics, University Hospital Carl Gustav Carus, Technical University of Dresden, Dresden, Germany.
- German Center for Child and Adolescent Health (DZKJ), partner site Munich, Munich, Germany.
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30
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Deng J, Li P, Li S, Liang F, Hong M, Zhang T, Tan Y, Ren F, Zhang Y, Xu Z, Wang H. SYK identified by bioinformatics analysis promotes the proliferation of multiple myeloma. Expert Rev Hematol 2025:1-15. [PMID: 40387108 DOI: 10.1080/17474086.2025.2505724] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2025] [Revised: 05/05/2025] [Accepted: 05/07/2025] [Indexed: 05/20/2025]
Abstract
BACKGROUND Despite recent advancements, the pathogenesis of multiple myeloma (MM) remains incompletely elucidated, with relapse and therapy resistance persisting as major clinical challenges, underscoring the imperative to identify novel therapeutic targets. RESEARCH DESIGN AND METHODS Differentially expressed genes were initially screened from the GSE6477 and GSE6691 datasets. Subsequent functional annotation and pathway enrichment analyses were conducted utilizing the DAVID bioinformatics platform. A protein-protein interaction network was constructed via the STRING database, followed by module analysis and hub genes identification through CytoHubba plugin. The biological significance of candidate genes was ultimately validated through ex vivo cellular functional assays and in vivo xenograft tumorigenesis experiments in murine models. RESULTS Bioinformatics analysis identified spleen tyrosine kinase (SYK) as the most prognostically significant candidate gene (p = 0.027). The SYK-specific inhibitor BAY61-3606 demonstrated time- (p < 0.05) and dose- (p < 0.01) dependent inhibition of MM cell viability, concomitant induction of G2/M phase cell cycle arrest (p < 0.001), and significant promotion of apoptosis (p < 0.05). In vivo experiments utilizing MM xenograft models demonstrated that BAY61-3606 administration significantly attenuated tumor growth kinetics (p < 0.05). CONCLUSIONS Our findings establish SYK as a therapeutic target in MM, thereby facilitating the development of innovative treatment strategies.
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Affiliation(s)
- Ju Deng
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
| | - Peichun Li
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
| | - Shuo Li
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
| | - Fengting Liang
- Department of Hematology, Bobai Country People's Hospital, Yulin, China
| | - Minglin Hong
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
| | - Ting Zhang
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
| | - Yanhong Tan
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
| | - Fanggang Ren
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
| | - Yaofang Zhang
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
| | - Zhifang Xu
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
| | - Hongwei Wang
- Institute of Hematology, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Key Laboratory of Molecular Diagnosis and Treatment of Hematological Disease of Shanxi Province, The Second Hospital of Shanxi Medical University, Taiyuan, China
- The Second Clinical Medical College, Shanxi Medical University, Taiyuan, China
- Party and Government Affairs Office, Shanxi University of Chinese Medicine, Jinzhong, China
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31
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López-Paleta N, Moreno-Barbosa E, Velázquez-Castro J. A fast validation test of gene regulatory network models via the Fokker-Planck equation. J Biol Phys 2025; 51:16. [PMID: 40388063 PMCID: PMC12089004 DOI: 10.1007/s10867-025-09681-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2025] [Accepted: 04/23/2025] [Indexed: 05/20/2025] Open
Abstract
Since Waddington proposed the concept of the "epigenetic landscape" in 1957, researchers have developed various methodologies to represent it in diverse processes. Studying the epigenetic landscape provides valuable qualitative information regarding cell development and the stability of phenotypic and morphogenetic patterns. Although Waddington's original idea was a visual metaphor, a contemporary perspective relates it to the landscape formed by the basins of attraction of a dynamical system describing the temporal evolution of protein concentrations driven by a gene regulatory network. Transitions among these attractors can be driven by stochastic perturbations, with the cell state more likely to transition to the nearest attractor or to the one that presents the path of least resistance. In this study, we define the epigenetic landscape using the free energy potential obtained from the solution of the Fokker-Planck equation on the regulatory network. Specifically, we obtained a numerical approximate solution of the Fokker-Planck equation describing the Arabidopsis thaliana flower morphogenesis process. We observed good agreement between the coexpression matrix obtained from the Fokker-Planck equation and the experimental coexpression matrix. This paper proposes a method for obtaining this landscape by solving the Fokker-Planck equation (FPE) associated with a dynamical system describing the temporal evolution of protein concentrations involved in the process of interest. As these systems are high-dimensional and analytical solutions are often unfeasible, we propose a gamma mixture model to solve the FPE, transforming this problem into an optimization problem. This methodology can enhance the analysis of gene regulatory networks by directly relating theoretical mathematical models with experimental observations of coexpression matrices, thus providing a discriminating technique for competing models.
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Affiliation(s)
- Natalia López-Paleta
- Facultad de Ciencias Físico Matemáticas, Benemérita Universidad Autónoma de Puebla, Puebla, 72570, Puebla, México
| | - Eduardo Moreno-Barbosa
- Facultad de Ciencias Físico Matemáticas, Benemérita Universidad Autónoma de Puebla, Puebla, 72570, Puebla, México
| | - Jorge Velázquez-Castro
- Facultad de Ciencias Físico Matemáticas, Benemérita Universidad Autónoma de Puebla, Puebla, 72570, Puebla, México.
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32
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Farrell C, Buhidma Y, Mumford P, Heywood WE, Hällqvist J, Flores-Aguilar L, Andrews EJ, Rahimzadah N, Taso OS, Doran E, Swarup V, Head E, Lashley T, Mills K, Toomey CE, Wiseman FK. Apolipoprotein E abundance is elevated in the brains of individuals with Down syndrome-Alzheimer's disease. Acta Neuropathol 2025; 149:49. [PMID: 40387921 PMCID: PMC12089208 DOI: 10.1007/s00401-025-02889-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2025] [Revised: 04/25/2025] [Accepted: 05/01/2025] [Indexed: 05/20/2025]
Abstract
Trisomy of chromosome 21, the cause of Down syndrome (DS), is the most commonly occurring genetic cause of Alzheimer's disease (AD). Here, we compare the frontal cortex proteome of people with Down syndrome-Alzheimer's disease (DSAD) to demographically matched cases of early onset AD and healthy ageing controls. We find dysregulation of the proteome, beyond proteins encoded by chromosome 21, including an increase in the abundance of the key AD-associated protein, APOE, in people with DSAD compared to matched cases of AD. To understand the cell types that may contribute to changes in protein abundance, we undertook a matched single-nuclei RNA-sequencing study, which demonstrated that APOE expression was elevated in subtypes of astrocytes, endothelial cells, and pericytes in DSAD. We further investigate how trisomy 21 may cause increased APOE. Increased abundance of APOE may impact the development of, or response to, AD pathology in the brain of people with DSAD, altering disease mechanisms with clinical implications. Overall, these data highlight that trisomy 21 alters both the transcriptome and proteome of people with DS in the context of AD, and that these differences should be considered when selecting therapeutic strategies for this vulnerable group of individuals who have high risk of early onset dementia.
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Affiliation(s)
- Clíona Farrell
- UK Dementia Research Institute at University College London, London, UK
- Queen Square Institute of Neurology, University College London, London, UK
| | - Yazead Buhidma
- Queen Square Institute of Neurology, University College London, London, UK
| | - Paige Mumford
- UK Dementia Research Institute at University College London, London, UK
- Queen Square Institute of Neurology, University College London, London, UK
| | - Wendy E Heywood
- UCL Great Ormond Street Institute of Child Heath, University College London, London, UK
| | - Jenny Hällqvist
- UCL Great Ormond Street Institute of Child Heath, University College London, London, UK
| | - Lisi Flores-Aguilar
- Department of Pathology and Laboratory Medicine, University of California, Irvine, CA, USA
| | - Elizabeth J Andrews
- Department of Pathology and Laboratory Medicine, University of California, Irvine, CA, USA
| | - Negin Rahimzadah
- Mathematical, Computational, and Systems Biology (MCSB) Program, University of California, Irvine, Irvine, CA, USA
- Institute for Memory Impairments and Neurological Disorders (MIND), University of California, Irvine, Irvine, CA, USA
- Center for Complex Biological Systems (CCBS), University of California Irvine, Irvine, CA, USA
| | - Orjona Stella Taso
- UK Dementia Research Institute at University College London, London, UK
- Queen Square Institute of Neurology, University College London, London, UK
| | - Eric Doran
- Department of Pediatrics, School of Medicine, University of California, Irvine, Orange, CA, USA
| | - Vivek Swarup
- Institute for Memory Impairments and Neurological Disorders (MIND), University of California, Irvine, Irvine, CA, USA
- Center for Complex Biological Systems (CCBS), University of California Irvine, Irvine, CA, USA
- Department of Neurobiology and Behaviour, University of California, Irvine, CA, USA
| | - Elizabeth Head
- Department of Pathology and Laboratory Medicine, University of California, Irvine, CA, USA
| | - Tammaryn Lashley
- Queen Square Institute of Neurology, University College London, London, UK
| | - Kevin Mills
- UCL Great Ormond Street Institute of Child Heath, University College London, London, UK
| | - Christina E Toomey
- Queen Square Institute of Neurology, University College London, London, UK.
- The Francis Crick Institute, London, UK.
| | - Frances K Wiseman
- UK Dementia Research Institute at University College London, London, UK.
- Queen Square Institute of Neurology, University College London, London, UK.
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33
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Mizrachi A, Sadeh M, Ben-Dor S, Dym O, Ku C, Feldmesser E, Zarfin A, Brunson JK, Allen AE, Jinkerson RE, Schatz D, Vardi A. Cathepsin X is a conserved cell death protein involved in algal response to environmental stress. Curr Biol 2025; 35:2240-2255.e6. [PMID: 40233752 DOI: 10.1016/j.cub.2025.03.045] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/22/2025] [Revised: 03/12/2025] [Accepted: 03/21/2025] [Indexed: 04/17/2025]
Abstract
Phytoplankton are responsible for half of the global photosynthesis and form vast blooms in aquatic ecosystems. Bloom demise fuels marine microbial life and is suggested to be mediated by programmed cell death (PCD) induced by diverse environmental stressors. Despite its importance, the molecular basis for algal PCD remains elusive. Here, we reveal novel PCD genes conserved across distant algal lineages using cell-to-cell heterogeneity in the response of the diatom Phaeodactylum tricornutum to oxidative stress. Comparative transcriptomics of sorted sensitive and resilient subpopulations following oxidative stress revealed genes directly linked to their contrasting fates of cell death and survival. Comparing these genes with those found in a large-scale mutant screen in the green alga Chlamydomonas reinhardtii identified functionally relevant conserved PCD gene candidates, including the cysteine protease cathepsin X/Z (CPX). CPX mutants in P. tricornutum CPX1 and C. reinhardtii CYSTEINE ENDOPEPTIDASE 12 (CEP12) exhibited resilience to oxidative stress and infochemicals that induce PCD, supporting a conserved function of these genes in algal PCD. Phylogenetic and predictive structural analyses show that CPX is highly conserved in eukaryotes, and algae exhibit strong structural similarity to human Cathepsin X/Z (CTSZ), a protein linked to various diseases. CPX is expressed by diverse algae across the oceans and correlates with upcoming demise events during toxic Pseudo-nitzschia blooms, providing support for its ecological significance. Elucidating PCD components in algae sheds light on the evolutionary origin of PCD in unicellular organisms and on the cellular strategies employed by the population to cope with stressful conditions.
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Affiliation(s)
- Avia Mizrachi
- Department of Plant and Environmental Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel
| | - Mai Sadeh
- Department of Plant and Environmental Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel
| | - Shifra Ben-Dor
- Bioinformatics Unit, Department of Life Sciences Core Facilities, Weizmann Institute of Science, Rehovot 7610001, Israel
| | - Orly Dym
- Department of Life Sciences Core Facilities, Weizmann Institute of Science, Rehovot 7610001, Israel
| | - Chuan Ku
- Department of Plant and Environmental Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel
| | - Ester Feldmesser
- Bioinformatics Unit, Department of Life Sciences Core Facilities, Weizmann Institute of Science, Rehovot 7610001, Israel
| | - Amichai Zarfin
- Department of Plant and Environmental Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel
| | - John K Brunson
- Scripps Institution of Oceanography, University of California, San Diego, La Jolla, San Diego, CA 92093, USA; Department of Environment and Sustainability, J. Craig Venter Institute, La Jolla, San Diego, CA 92037, USA
| | - Andrew E Allen
- Scripps Institution of Oceanography, University of California, San Diego, La Jolla, San Diego, CA 92093, USA; Department of Environment and Sustainability, J. Craig Venter Institute, La Jolla, San Diego, CA 92037, USA
| | - Robert E Jinkerson
- Department of Chemical and Environmental Engineering, University of California, Riverside, CA 92521, USA
| | - Daniella Schatz
- Department of Plant and Environmental Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel
| | - Assaf Vardi
- Department of Plant and Environmental Sciences, Weizmann Institute of Science, Rehovot 7610001, Israel.
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34
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Heloterä H, Kostanek J, Liukkonen M, Siintamo L, Linna-Kuosmanen S, Watala C, Blasiak J, Kaarniranta K. Serum RNA Profile Reflects Fluid Status and Atrophic Retinal Changes in Neovascular Age-Related Macular Degeneration. Int J Mol Sci 2025; 26:4852. [PMID: 40429992 PMCID: PMC12112293 DOI: 10.3390/ijms26104852] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2025] [Revised: 04/23/2025] [Accepted: 04/30/2025] [Indexed: 05/29/2025] Open
Abstract
The increasing prevalence of age-related macular degeneration (AMD), a disease that can result in the loss of central vision, is an emerging problem worldwide due to aging societies. Growing patient numbers create a challenge for the healthcare system. Understanding the mechanisms of AMD pathogenesis will aid in early, personalized, and efficient intervention, helping to mitigate this issue. Current diagnostic methods rely on optical coherence tomography and angiography imaging, which identify existing damages, but do not provide information on the mechanisms behind them. In the present work, we demonstrate a difference in the serum RNA profile between neovascular AMD (nAMD) patients and controls. Moreover, the RNA profile of nAMD patients corresponded with anatomical changes in the retinal fluid compartments as well as atrophic changes of the retina. We followed two independent ways to control false positive leads, and when these approaches were combined, thioredoxin-related transmembrane protein 4 (TMX4) was observed to be differentially expressed by both approaches. This finding opens a new pathway in AMD studies, which are limited due to restricted access to live human target material and the limited value of animal models of human AMD.
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Affiliation(s)
- Hanna Heloterä
- Department of Ophthalmology, University of Eastern Finland, 70210 Kuopio, Finland; (H.H.); (M.L.)
| | - Joanna Kostanek
- Department of Haemostatic Disorders, Medical University of Lodz, 90-419 Lodz, Poland; (J.K.)
| | - Mikko Liukkonen
- Department of Ophthalmology, University of Eastern Finland, 70210 Kuopio, Finland; (H.H.); (M.L.)
| | - Leea Siintamo
- Department of Ophthalmology, University of Eastern Finland, 70210 Kuopio, Finland; (H.H.); (M.L.)
| | - Suvi Linna-Kuosmanen
- A. I. Virtanen Institute for Molecular Sciences, University of Eastern Finland, 70211 Kuopio, Finland;
| | - Cezary Watala
- Department of Haemostatic Disorders, Medical University of Lodz, 90-419 Lodz, Poland; (J.K.)
| | - Janusz Blasiak
- Faculty of Medicine, Collegium Medicum, Mazovian Academy in Plock, 09-402 Plock, Poland;
| | - Kai Kaarniranta
- Department of Ophthalmology, University of Eastern Finland, 70210 Kuopio, Finland; (H.H.); (M.L.)
- Department of Ophthalmology, Kuopio University Hospital, 70210 Kuopio, Finland
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Qu K, Gu J, Xu Z, Hepeng Y, Ning Q, Wu X. Alcohol consumption and esophageal cancer risk: unveiling DLEU2 as a key immune modulator through Mendelian randomization and transcriptomic analysis. Discov Oncol 2025; 16:804. [PMID: 40383861 PMCID: PMC12086134 DOI: 10.1007/s12672-025-02660-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/15/2024] [Accepted: 05/10/2025] [Indexed: 05/20/2025] Open
Abstract
Esophageal cancer (EC) is a leading cause of cancer-related mortality globally, with alcohol consumption being a significant risk factor. However, the genetic and molecular mechanisms linking alcohol intake to EC remain unclear. This study utilized Mendelian randomization analysis to establish a causal relationship between alcohol consumption and EC (OR: 4.11 [95% CI 1.83-9.23]). Transcriptomic analysis identified 83 differentially expressed genes (log₂ fold change > 1, false discovery rate [FDR] < 0.05), among which DLEU2 was uniquely transcribed into a long non-coding RNA (lncRNA). Pan-cancer analysis revealed its association with the tumor immune microenvironment and cancer progression. Single-cell RNA sequencing localized DLEU2 expression predominantly to T cells, particularly exhausted subpopulations, and pseudo-temporal analysis demonstrated increased DLEU2 expression during late T cell differentiation stages, co-expressing immune suppression markers, with consistent expression patterns observed across multiple patient-derived samples. Additionally, cell communication analysis suggested that DLEU2 modulates TNF signaling through TNFRSF1A/B pathways, contributing to immune evasion and poor prognosis. These findings position DLEU2 as a pivotal regulator of the immune landscape in EC and a potential prognostic biomarker and therapeutic target.
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Affiliation(s)
- Kailin Qu
- Department of Thoracic Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, China
| | - Jingyan Gu
- Department of Neurosurgery, Shanghai General Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, Shanghai, China
| | - Zhi Xu
- Department of General Medicine, Nanfang Hospital, Southern Medical University, Guangzhou, China
| | - Yixiang Hepeng
- Department of Thoracic Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, China
| | - Qi Ning
- Department of General Medicine, Nanfang Hospital, Southern Medical University, Guangzhou, China
| | - Xu Wu
- Department of Thoracic Surgery, Nanfang Hospital, Southern Medical University, Guangzhou, China.
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Wang J, Chu Y, Hu G, Liang S, Cui Y, Zuo J, Luo Z, Chen X, Wang X, Yu Z, Zhang Z, Wang X. Piperlongumine Inhibits Malignant Progression of Esophageal Squamous Cells Through the PI3K/AKT Signaling Pathway. Biochem Genet 2025:10.1007/s10528-025-11139-7. [PMID: 40383869 DOI: 10.1007/s10528-025-11139-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/18/2025] [Accepted: 05/08/2025] [Indexed: 05/20/2025]
Abstract
To examine the impact of Piperlongumine (PL) on the proliferation, migration, invasion, cell cycle progression, and apoptosis in esophageal squamous cell carcinoma (ESCC) cells, as well as to elucidate the underlying molecular mechanisms. The suppressive effects of PL on the viability of ESCC cells were assessed using the CCK-8 assay, bright field imaging, and colony formation assays. Apoptosis induction and cell cycle disruption by PL were evaluated using flow cytometry. The impact of PL on ESCC cell migration and invasion was examined through scratch healing and Transwell assays. Differential gene expression analysis of ESCC tumor and normal tissues from the GSE29886 dataset, integrated with network pharmacology predictions, was conducted to identify core genes and molecular mechanisms involved in PL action. Key protein expression levels in the apoptosis, epithelial-mesenchymal transition (EMT), and PI3K/AKT signaling pathways were quantified by Western blotting. The CCK-8 and colony formation assays demonstrated that PL effectively suppressed cell viability and proliferation in ESCC. Flow cytometry revealed that PL down-regulated CDK1 expression, resulting in G2/M phase arrest, and promoted apoptosis by decreasing Bcl-2 levels and increasing cleaved caspase-3 and PARP. The scratch and Transwell assays indicated that PL inhibited ESCC cell migration and invasion, down-regulated the EMT-associated proteins Vimentin and N-cadherin, and up-regulated E-cadherin. Western blotting confirmed the down-regulation of P-PI3K and P-AKT, indicating the inhibition of the PI3K/AKT pathway by PL. These findings offer a pharmacological foundation for the development of PL as a potential phytotherapeutic agent for the clinical management of ESCC.
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Affiliation(s)
- Jun Wang
- Department of Gastroenterology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Yueming Chu
- School of Pharmacy, North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Guangbing Hu
- Department of Gastroenterology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Shiqi Liang
- Department of Gastroenterology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Yutong Cui
- Department of Gastroenterology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Ji Zuo
- Department of Gastroenterology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Zichen Luo
- Department of Gastroenterology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Xinrui Chen
- Department of Gastroenterology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Xiaobo Wang
- Department of Gastroenterology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Zhenghang Yu
- Department of General Surgery, The Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Zhongyang Zhang
- Department of General Surgery, The Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China
| | - Xianfei Wang
- Department of Gastroenterology, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China.
- Digestive Endoscopy Center, Affiliated Hospital of North Sichuan Medical College, Nanchong, 637000, Sichuan, China.
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Santaeufemia S, Marchetto F, Romano P, Adamska D, Goryca K, Palatini J, Kargul J. Transcriptomics Reveals an Energy-Saving Metabolic Switch in an Extremophilic Red Microalga Cyanidioschyzon merolae Under Nickel Stress. Int J Mol Sci 2025; 26:4813. [PMID: 40429959 PMCID: PMC12112623 DOI: 10.3390/ijms26104813] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/14/2025] [Revised: 05/06/2025] [Accepted: 05/12/2025] [Indexed: 05/29/2025] Open
Abstract
The red microalga Cyanidioschyzon merolae inhabits extreme environments with high temperatures (40-56 °C), high acidity (pH 0.05-4), and high concentrations of heavy metals that are lethal to most forms of life. However, information is scarce on the precise adaptation mechanisms of this extremophile to such hostile conditions. Gaining such knowledge is important for understanding the evolution of microorganisms in the early stages of life on Earth characterized by such extreme environments. Through an analysis of the re-programming of the global transcriptome upon the long-term (up to 15 days) exposure of C. merolae to extremely high concentrations of nickel (1 and 3 mM), the key adaptive metabolic pathways and associated molecular components were identified. Our work shows that the long-term Ni exposure of C. merolae leads to the lagged metabolic switch demonstrated via the transcriptional upregulation of the metabolic pathways critical for cell survival. DNA replication, cell cycle, and protein quality control processes were upregulated, while downregulation occurred with energetically costly processes, including the assembly of the photosynthetic apparatus and lipid biosynthesis. This study paves the way for future multi-omic studies of the molecular mechanisms of abiotic stress adaptation in phototrophs, as well as the future development of rational approaches to the bioremediation of contaminated aquatic environments.
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Affiliation(s)
- Sergio Santaeufemia
- Solar Fuels Laboratory, Center of New Technologies, University of Warsaw, 02-097 Warsaw, Poland; (S.S.); (F.M.)
| | - Francesca Marchetto
- Solar Fuels Laboratory, Center of New Technologies, University of Warsaw, 02-097 Warsaw, Poland; (S.S.); (F.M.)
| | - Patrizia Romano
- Department of Physical Sciences, Earth and Environment, University of Siena, 53100 Siena, Italy;
| | - Dorota Adamska
- Genomics Core Facility, Centre of New Technologies, University of Warsaw, 02-097 Warsaw, Poland; (D.A.); (K.G.); (J.P.)
| | - Krzysztof Goryca
- Genomics Core Facility, Centre of New Technologies, University of Warsaw, 02-097 Warsaw, Poland; (D.A.); (K.G.); (J.P.)
| | - Jeffrey Palatini
- Genomics Core Facility, Centre of New Technologies, University of Warsaw, 02-097 Warsaw, Poland; (D.A.); (K.G.); (J.P.)
| | - Joanna Kargul
- Solar Fuels Laboratory, Center of New Technologies, University of Warsaw, 02-097 Warsaw, Poland; (S.S.); (F.M.)
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Shang K, Huang D, Liu J, Yu Z, Bian W, Chen J, Zhao Y, Liu L, Jiang J, Wang Y, Duan Y, Ge J, Zhang S, Zhou C, Han Y, Hu Y, Zheng W, Sun J, Huang H, Pei S, Qian P, Sun J. CD97-directed CAR-T cells with enhanced persistence eradicate acute myeloid leukemia in diverse xenograft models. Cell Rep Med 2025:102148. [PMID: 40425009 DOI: 10.1016/j.xcrm.2025.102148] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2024] [Revised: 11/11/2024] [Accepted: 04/30/2025] [Indexed: 05/29/2025]
Abstract
Chimeric antigen receptor (CAR)-T therapy on acute myeloid leukemia (AML) is hindered by the absence of a suitable tumor-specific antigen. Here, we propose CD97 as a potential target for CAR-T therapy against AML based on its broader and higher expression on AML cells compared to normal hematopoietic stem and progenitor cells (HSPCs). To resolve the fratricide problem caused by CD97 expression on T cells, we knock out CD97 in CAR-T cells using CRISPR-Cas9. Our CD97KO CAR-T cells eliminate both AML cell lines and primary AML cells effectively while showing tolerable toxicity to HSPCs. Furthermore, we mutate the CD3ζ domain of the CAR and find that the optimized CD97 CAR-T cells exhibit persistent anti-tumor activity both in vitro and in multiple xenograft models. Mechanistically, transcriptional profiles reveal that the optimized CAR-T cells delay differentiation and resist exhaustion. Collectively, our study supports CD97 as a promising target for CAR-T therapy against AML.
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Affiliation(s)
- Kai Shang
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Department of Cell Biology, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China
| | - Deyu Huang
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China; Center for Stem Cell and Regenerative Medicine, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Jun Liu
- Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China
| | - Zebin Yu
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China; Center for Stem Cell and Regenerative Medicine, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Wei Bian
- Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China
| | - Jiangqing Chen
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Department of Cell Biology, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China
| | - Yin Zhao
- Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China
| | - Lina Liu
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China
| | - Jie Jiang
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Department of Cell Biology, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China
| | - Yajie Wang
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Department of Cell Biology, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China
| | - Yanting Duan
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Department of Cell Biology, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China
| | - Jingyu Ge
- Department of Medical Oncology, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310003, China
| | - Shize Zhang
- Center for Genetic Medicine, The Fourth Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Chun Zhou
- School of Public Health, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China
| | - Yingli Han
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China; Center for Stem Cell and Regenerative Medicine, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Yongxian Hu
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China
| | - Weiyan Zheng
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China
| | - Jie Sun
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China
| | - He Huang
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China.
| | - Shanshan Pei
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China.
| | - Pengxu Qian
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China; Center for Stem Cell and Regenerative Medicine, Zhejiang University School of Medicine, Hangzhou 310058, China.
| | - Jie Sun
- Bone Marrow Transplantation Center of the First Affiliated Hospital, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Liangzhu Laboratory, Zhejiang University, 1369 West Wenyi Road, Hangzhou 311121, China; Department of Cell Biology, Zhejiang University School of Medicine, 866 Yuhangtang Road, Hangzhou 310058, China; Institute of Hematology, Zhejiang University, Hangzhou 310058, China; Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou 310058, China.
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Yang Y, Jing W, Zhang L, Zhang Y, Shang Y, Kuang Y. WDR62 affects the progression of ovarian cancer by regulating the cell cycle. Hereditas 2025; 162:78. [PMID: 40369663 PMCID: PMC12076949 DOI: 10.1186/s41065-025-00444-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/05/2025] [Accepted: 05/02/2025] [Indexed: 05/16/2025] Open
Abstract
BACKGROUND Ovarian Cancer (OC) is a gynecological malignant tumor with an extremely high mortality rate, seriously endangering women's health. Due to its insidious clinical manifestations, most patients are diagnosed in the advanced stage of the disease. The currently clinically relied CA125 has limited specificity for the early diagnosis of ovarian cancer. Hence, identifying new promising biomarkers is crucial for the early screening, diagnosis, and treatment of ovarian cancer. Based on differential expression analysis, WGCNA and survival analysis, we identified a centromere-associated gene, WDR62, which is highly expressed in ovarian cancer and highly correlated with ovarian cancer, as well as the poor prognosis of ovarian cancer patients with high expression, suggesting that WDR62 may be a potential biomarker for ovarian cancer. Previous studies have shown that WDR62 is closely associated with the occurrence, development and prognosis of a variety of tumors. However, its role in ovarian cancer has not been studied in depth. METHODS Using combined TCGA and GTEx datasets from the UCSC database, along with WGCNA, and survival analysis, WDR62 was identified as a potential biomarker. GEPIA2 database, GEO database, qRT-PCR, and Western blot proved the expression of WDR62. Enrichment analysis, cell transfection, Western blots and CCK8 demonstrated the regulatory mechanism of WDR62, and the detailed mechanism of WDR62 involvement in the occurrence and development of ovarian cancer was predicted by interaction analysis and correlation analysis. RESULTS WDR62 was highly expressed in ovarian cancer cells compared to normal ovarian epithelial cells, both at the RNA and protein levels. Patients with high WDR62 expression had a poor survival prognosis. Upon WDR62 knockdown, the expression of cell cycle-related proteins CDK1 and C-Myc decreased in ovarian cancer cells, and the cell proliferative capacity was decreased. Based on bioinformatic analysis, it was hypothesized that WDR62 might mediate the JNK signaling pathway by interacting with MAPK8, thus affecting ovarian cancer progression through cell cycle regulation. CONCLUSIONS WDR62 is overexpressed in ovarian cancer and is closely related to the prognosis of ovarian cancer patients. WDR62 promotes ovarian cancer progression by regulating the cell cycle and may influence its development through interaction with MAPK8 to mediate the JNK signaling pathway. These findings suggest that WDR62 could be a potential target for the early screening, diagnosis, and treatment of ovarian cancer.
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Affiliation(s)
- Yuqi Yang
- Department of Gynecology and Obstetrics, The 2nd Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Wanting Jing
- Laboratory of Medical Genetics, Harbin Medical University, Harbin, China
| | - Lingqi Zhang
- Department of Gynecology and Obstetrics, The 2nd Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Yuhang Zhang
- Department of Gynecology and Obstetrics, The 2nd Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Ying Shang
- Department of Gynecology and Obstetrics, The 2nd Affiliated Hospital of Harbin Medical University, Harbin, China
| | - Ye Kuang
- Department of Gynecology and Obstetrics, The 2nd Affiliated Hospital of Harbin Medical University, Harbin, China.
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Moore SPG, Ganesh Krishnan S, Jaswanth Kothari R, Prince NB, Kenny C, Zhang C, Lang D. PAX3 Regulatory Signatures and Gene Targets in Melanoma Cells. Genes (Basel) 2025; 16:577. [PMID: 40428399 PMCID: PMC12111051 DOI: 10.3390/genes16050577] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/24/2025] [Revised: 05/09/2025] [Accepted: 05/13/2025] [Indexed: 05/29/2025] Open
Abstract
BACKGROUND/OBJECTIVES PAX3 is a transcription factor that drives melanoma progression by promoting cell growth, migration, and survival, while inhibiting cellular terminal differentiation. However, known PAX3 target genes are limited and cannot fully explain the wide impact of PAX3 function. The PAX3 protein can regulate DNA through two separate binding domains, the Paired Domain (PD) and Homeodomain (HD), which bind different DNA motifs. It is not clear if these two domains bind and work together to regulate genes and if they promote all or only a subset of downstream cellular events. METHODS PAX3 direct downstream targets were identified using Cleavage Under Targets & Release Using Nuclease (CUT&RUN) assays in SK-MEL-5 melanoma cells. PAX3-binding genomic regions were identified through MACS2 peak calling, and peaks were categorized based on the presence of PD and/or HD binding sites (or neither) through HOMER motif analysis. The peaks were further characterized as Active, Primed, Poised, Repressed, or Closed based on ATAC-seq data and CUT&RUN for histone Post-Translational Modifications H3K4me1, H3K4me3, H3K27me3, and H3K27Ac. RESULTS This analysis revealed that most of the PAX3 binding sites in the SK-MEL-5 cell line were primarily through the PD and connected to Active genes. Surprisingly, PAX3 does not commonly act as a repressor in SK-MEL-5 cells. Pathway analysis identified genes involved with transcription, RNA modification, and cell growth. Peaks located in distal enhancer elements were connected to genes involved in neuronal growth, function, and signaling. CONCLUSIONS Our results reveal novel PAX3 regulatory regions and putative genes in a melanoma cell line, with a predominance of PAX3 PD binding on active sites.
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Affiliation(s)
- Stephen P. G. Moore
- Department of Dermatology, Boston University, Boston, MA 02118, USA; (S.P.G.M.); (S.G.K.); (R.J.K.); (N.B.P.)
| | - Shripushkar Ganesh Krishnan
- Department of Dermatology, Boston University, Boston, MA 02118, USA; (S.P.G.M.); (S.G.K.); (R.J.K.); (N.B.P.)
- Bioinformatics Program, Boston University, Boston, MA 02118, USA
| | - Rutu Jaswanth Kothari
- Department of Dermatology, Boston University, Boston, MA 02118, USA; (S.P.G.M.); (S.G.K.); (R.J.K.); (N.B.P.)
- Bioinformatics Program, Boston University, Boston, MA 02118, USA
| | - Noah B. Prince
- Department of Dermatology, Boston University, Boston, MA 02118, USA; (S.P.G.M.); (S.G.K.); (R.J.K.); (N.B.P.)
| | - Colin Kenny
- Department of Surgery, University of Iowa, Iowa City, IA 52242, USA;
| | - Chao Zhang
- Department of Medicine, Boston University, Boston, MA 02118, USA
| | - Deborah Lang
- Department of Dermatology, Boston University, Boston, MA 02118, USA; (S.P.G.M.); (S.G.K.); (R.J.K.); (N.B.P.)
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Taguchi S, Matsuzawa R, Suda Y, Irie K, Ozaki H. Investigating the effects of liquid handling robot pipetting speed on yeast growth and gene expression using growth assays and RNA-seq. MICROPUBLICATION BIOLOGY 2025; 2025:10.17912/micropub.biology.001566. [PMID: 40438138 PMCID: PMC12117386 DOI: 10.17912/micropub.biology.001566] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/06/2025] [Revised: 05/01/2025] [Accepted: 05/06/2025] [Indexed: 06/01/2025]
Abstract
Assessing the impact of experimental parameters like pipetting speed is essential in life science research but challenging in manual experiments. Recent advancements in laboratory automation enable precise control and systematic evaluation of these parameters. Here, we employed the Opentrons OT-2, an affordable, open-source liquid handling robot, to systematically investigate the effect of pipetting speed on growth and gene expression in the budding yeast Saccharomyces cerevisiae . Growth assays revealed no significant differences across the tested pipetting speeds (ANOVA, p > 0.05). RNA-seq analysis corroborated these findings, demonstrating highly similar gene expression profiles across all 24 samples (minimum Pearson correlation coefficient = 0.9528), with no differentially expressed genes identified by generalized linear model analysis (false discovery rate > 0.01). Our results highlight the utility of robotic platforms in optimizing experimental parameters, improving reproducibility, and enhancing accuracy in biological research, thus providing valuable insights for future applications.
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Affiliation(s)
- Shodai Taguchi
- Ph.D. Program in Humanics, School of Integrative and Global Majors, University of Tsukuba, Tsukuba, Ibaraki, Japan
- Department of Molecular Cell Biology, Institute of Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan
- Bioinformatics Laboratory, Institute of Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan
| | - Ryosuke Matsuzawa
- Bioinformatics Laboratory, Institute of Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan
- Doctoral Program in Medical Sciences, Degree Programs in Comprehensive Human Sciences, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan
| | - Yasuyuki Suda
- Department of Molecular Cell Biology, Institute of Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan
| | - Kenji Irie
- Department of Molecular Cell Biology, Institute of Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan
| | - Haruka Ozaki
- Bioinformatics Laboratory, Institute of Medicine, University of Tsukuba, Tsukuba, Ibaraki, Japan
- Center for Artificial Intelligence Research, University of Tsukuba, Tsukuba, Ibaraki, Japan
- Laboratory for AI Biology, RIKEN Center for Biosystems Dynamics Research, Kobe, Hyōgo, Japan
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42
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Toft K, Mardahl M, Hedberg-Alm Y, Anlén K, Tydén E, Nielsen MK, Honoré ML, Fromm B, Nielsen LN, Nejsum P, Thamsborg SM, Cirera S, Pihl TH. Evaluation of circulating microRNAs in plasma from horses with non-strangulating intestinal infarction and idiopathic peritonitis. Vet J 2025; 313:106378. [PMID: 40374098 DOI: 10.1016/j.tvjl.2025.106378] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2025] [Revised: 05/12/2025] [Accepted: 05/12/2025] [Indexed: 05/17/2025]
Abstract
Non-strangulating intestinal infarctions (NSII) associated with Strongylus vulgaris infection and idiopathic peritonitis (IP) share similar clinical presentation but require different treatment approaches. Horses with NSII need surgical intervention, while idiopathic peritonitis cases can be successfully treated with antimicrobials. A correct diagnosis is thus crucial, but because the two diseases overlap in clinicopathological features, differentiation is difficult in clinical practice. MicroRNAs (miRNAs) are non-coding RNAs that exhibit measurable changes in abundance in tissues and circulation during disease. This study aimed to explore differences in plasma miRNA abundance between patients with NSII and IP. Plasma samples were collected from 43 horses, consisting of 21 with NSII and 22 with IP. A subset (n = 12) was submitted for deep small RNA sequencing to identify miRNAs differing between the groups. Next, a panel of nine miRNAs (two were potential normalizers) were selected for evaluation and confirmation by reverse transcription quantitative real-time PCR (RT-qPCR). Small RNA sequencing detected 628 miRNAs in the blood samples, but no miRNAs were differentially abundant between the disease groups. This finding was confirmed by qPCR. In agreement with previous studies, the top abundant miRNAs in both groups included Eca-Mir-122-5p and Eca-Mir-486-5p, as well as Eca-Mir-223-3p, which has previously been associated with inflammation. Target prediction for the most abundant miRNAs additionally predicted targets in inflammatory pathways. Evaluation of clinicopathological parameters revealed differences between the groups in two measures (white blood cell count and blood neutrophil count), which aligns with findings from previous studies. The results demonstrate that NSII and IP elicit similar miRNA profiles in plasma and are characterized by systemic inflammation.
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Affiliation(s)
- Katrine Toft
- Department of Veterinary Clinical Sciences, University of Copenhagen, Copenhagen, Denmark.
| | | | - Ylva Hedberg-Alm
- Department of Biomedical Science and Veterinary Public Health, Parasitology Unit, Swedish University of Agricultural Sciences, Uppsala, Sweden
| | - Karin Anlén
- Evidensia Equine Hospital Helsingborg, Helsingborg, Sweden
| | - Eva Tydén
- Department of Biomedical Science and Veterinary Public Health, Parasitology Unit, Swedish University of Agricultural Sciences, Uppsala, Sweden
| | - Martin K Nielsen
- M.H. Gluck Equine Research Center, Department of Veterinary Science, University of Kentucky, Lexington, KY, USA
| | - Marie Louise Honoré
- Department of Veterinary Clinical Sciences, University of Copenhagen, Copenhagen, Denmark
| | - Bastian Fromm
- The Arctic University Museum of Norway, UiT The Arctic University of Norway, Tromsø, Norway
| | - Lise N Nielsen
- Department of Veterinary Clinical Sciences, University of Copenhagen, Copenhagen, Denmark
| | - Peter Nejsum
- Department of Clinical Medicine, Aarhus University, Aarhus, Denmark
| | - Stig Milan Thamsborg
- Department of Veterinary and Animal Sciences, University of Copenhagen, Copenhagen, Denmark
| | - Susanna Cirera
- Department of Veterinary and Animal Sciences, University of Copenhagen, Copenhagen, Denmark
| | - Tina Holberg Pihl
- Department of Veterinary Clinical Sciences, University of Copenhagen, Copenhagen, Denmark
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43
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Ebelt ND, Loganathan S, Avsharian LC, Manuel ER. Transcriptomic analysis of non-leukemic cell subsets in azacytidine-responsive AML highlights pathways associated with adhesion, platelet aggregation, and angiogenesis in mice and humans. Mol Med 2025; 31:185. [PMID: 40360989 PMCID: PMC12070539 DOI: 10.1186/s10020-025-01233-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2025] [Accepted: 04/25/2025] [Indexed: 05/15/2025] Open
Abstract
BACKGROUND Hypomethylating agents (HMAs), such as 5-azacytidine (AZA), are valuable treatment options for patients with acute myeloid leukemia (AML). Despite providing significant extensions in survival when used alone or in combination with BCL-2 inhibitors, resistance and eventual relapse is observed. Reported mechanisms of these outcomes are inconsistent when focusing on leukemic populations within bone marrow, indicating a need for studies on the impact of HMAs on non-leukemic cells in the blood and other tissue compartments. METHODS Whole blood and spleens from vehicle- or AZA-treated mice implanted with the syngeneic AML line C1498 were transcriptionally profiled using a comprehensive panel of immune-related gene probes. Publicly available RNAseq data from blood of AZA-responsive, human AML patients were analyzed compared to matched, pre-treatment samples. Genes differentially expressed between vehicle- and AZA-treated (mouse) or pre- and post-AZA treatment (human) samples were analyzed for statistical overrepresentation in gene ontologies using Fisher's one-tailed t-test. Pathological analyses of various tissues in AML relapsed, AZA-responsive mice were compared to the corresponding tissues in vehicle-treated mice. RESULTS We observed hematologic recovery in the peripheral blood of AZA-treated groups, versus vehicle control, that was associated with significant extensions in survival. Transcriptional analysis of AZA-treated samples revealed decreased cell type scores for suppressive subsets and increased pathway scores for T and B cell functions. Comparisons of gene ontology annotations enriched from genes differentially regulated by AZA in human and mouse blood samples revealed overlap in numerous biological pathways including adhesion, thrombosis, and angiogenesis. Consistently, C1498 permeated the liver at end-stage disease in vehicle-treated mice, while AZA treatment limited their spread to just outside the bone after relapse. CONCLUSIONS AZA-induced differences in C1498 spread could be a result of gene expression changes in adhesion, platelet aggregation and/or angiogenesis in non-leukemic compartments; however, further mechanistic studies must be done to confirm a direct link between modulated genes and disease manifestation. Overall, these studies provide rationale for expanding the exploration of biomarkers and therapeutic targets to include normal immune cells in blood, spleen, or other microenvironments of AML patients treated with HMA, rather than limiting studies to the bone marrow and leukemic blasts.
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MESH Headings
- Animals
- Humans
- Leukemia, Myeloid, Acute/drug therapy
- Leukemia, Myeloid, Acute/genetics
- Leukemia, Myeloid, Acute/metabolism
- Leukemia, Myeloid, Acute/pathology
- Mice
- Azacitidine/pharmacology
- Azacitidine/therapeutic use
- Gene Expression Profiling
- Transcriptome
- Cell Adhesion/drug effects
- Cell Adhesion/genetics
- Neovascularization, Pathologic/genetics
- Antimetabolites, Antineoplastic/pharmacology
- Antimetabolites, Antineoplastic/therapeutic use
- Angiogenesis
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Affiliation(s)
- Nancy D Ebelt
- Department of Immuno-Oncology, Beckman Research Institute of the City of Hope, Duarte, CA, USA
| | | | - Lara C Avsharian
- Department of Immuno-Oncology, Beckman Research Institute of the City of Hope, Duarte, CA, USA
- Irell and Manella Graduate School of Biological Sciences, City of Hope, Duarte, CA, USA
| | - Edwin R Manuel
- Department of Immuno-Oncology, Beckman Research Institute of the City of Hope, Duarte, CA, USA.
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44
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Chen Y, Wang M. Revealing roles of PANoptosis-related genes in prognosis and molecular subtypes in lung squamous cell carcinoma by integrated bioinformatic analyses and experiments. Clin Exp Med 2025; 25:154. [PMID: 40353888 PMCID: PMC12069423 DOI: 10.1007/s10238-025-01696-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2025] [Accepted: 04/14/2025] [Indexed: 05/14/2025]
Abstract
The purpose of current study was to reveal the role of PANoptosis-associated genes in lung squamous cell carcinoma (LUSC) and their potential as prognostic biomarkers. We analyzed RNA-seq data from TCGA-LUSC and GEO datasets to identify differentially expressed genes (DEGs) between LUSC and normal samples, followed by VENN analysis to reveal PANoptosis-related DEGs. Functional enrichment analyses were performed by clusterProfiler package. Distinct LUSC subtypes were identified by consensus clustering based on PANoptosis-related DEGs. Univariate Cox and LASSO regression were utilized to identify key prognostic genes, and a prognostic model was developed based on selected genes. Immune infiltration status was evaluated by CIBERSORT and ESTIMATE algorithms. Expression of key prognostic genes was tested in three LUSC cell lines by RT-qPCR and Western blot. Roles of TLR3 in LUSC progression were determined by functional experiments. A total of 76 PANoptosis-related DEGs were identified, with significant enrichment in apoptosis pathways. The clustering analysis revealed four subtypes, in which survival and immune microenvironment were dramatically different. From the 76 genes, four key prognostic genes (CHEK2, PDK4, TLR3, and IL1B) were identified to establish prognostic risk model, which could reflect the survival status and immune cells composition variations for LUSC patients. Besides, these four genes showed significant correlations with infiltrating levels of various immune cells. TLR3 was identified as a more weighted prognostic risk gene in LUSC. Functional assays demonstrated that genes like TLR3 modulated cell proliferation, migration, and inflammatory responses in LUSC cells. This study highlighted the potential of the four key PANoptosis genes as biomarkers or targets in LUSC, and the risk model based on these four genes provided novel insights to develop personalized treatment strategy for patients with LUSC.
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Affiliation(s)
- Ying Chen
- Department of Respiratory Medicine, Hangzhou Xixi Hospital Affiliated to Zhejiang Chinese Medical University (Hangzhou Xixi Hospital), No. 2 Hengbu Road, Hangzhou, 310023, Zhejiang, China
| | - Meihua Wang
- Department of Respiratory Medicine, Hangzhou Xixi Hospital Affiliated to Zhejiang Chinese Medical University (Hangzhou Xixi Hospital), No. 2 Hengbu Road, Hangzhou, 310023, Zhejiang, China.
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45
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de Andrade KP, Michaelsen GL, Dutra LF, Marques RF, Benincasa DER, Portich JP, Loss JF, Gregianin LJ, Brunetto AT, Sinigaglia M, Roesler R, da Cunha Jaeger M, Land M, de Farias CB. Neurotrophins as Potential Biomarkers for Active Disease and Poor Outcome in Pediatric Acute Lymphoblastic Leukemia. Cancers (Basel) 2025; 17:1623. [PMID: 40427122 PMCID: PMC12110685 DOI: 10.3390/cancers17101623] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2025] [Revised: 05/03/2025] [Accepted: 05/05/2025] [Indexed: 05/29/2025] Open
Abstract
BACKGROUND Neurotrophins (NTs) are pivotal growth factors in cellular development and survival. Their precise implications in Acute Lymphoblastic Leukemia (ALL) remain unclear. METHODS Pediatric ALL samples (2011-2021) were analyzed from a Southern Brazil cohort. Neurotrophin levels were quantified via ELISA, with survival analysis using Kaplan-Meier curves. Gene expression data were sourced from public genomic repositories and analyzed with R software version 4.0.5. RESULTS At diagnosis, pro-BDNF, BDNF, and NGF levels were significantly lower than in healthy controls. Reduced pro-BDNF correlated with unfavorable outcomes. NGF and sortilin were highly expressed in healthy samples, while BDNF and p75NTR were predominant in T-ALL and B-ALL, respectively. CONCLUSIONS Neurotrophins show significant alterations in the tumor microenvironment of pediatric ALL. Further studies are needed to elucidate their precise role and prognostic potential.
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Affiliation(s)
- Karine Pereira de Andrade
- Cancer and Neurobiology Laboratory, Experimental Research Center, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
- Postgraduate Program in Pharmacology and Therapeutics, Institute for Basic Health Sciences, Federal University of Rio Grande do Sul, Porto Alegre 90050-170, RS, Brazil
| | | | - Lívia Fratini Dutra
- Cancer and Neurobiology Laboratory, Experimental Research Center, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
| | - Rebeca Ferreira Marques
- Cancer and Neurobiology Laboratory, Experimental Research Center, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
- Pediatric Oncology Service, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
| | | | - Júlia Plentz Portich
- Cancer and Neurobiology Laboratory, Experimental Research Center, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
- Hematology and Hemotherapy Service, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
| | - Jiseh Fagundes Loss
- Pediatric Oncology Service, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
| | - Lauro José Gregianin
- Pediatric Oncology Service, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
- National Institute of Science and Technology in Childhood Cancer Biology and Pediatric Oncology—INCT BioOncoPed, Porto Alegre 90050-170, RS, Brazil
| | - André Tesainer Brunetto
- Children’s Cancer Institute, Porto Alegre 90620-110, RS, Brazil
- National Institute of Science and Technology in Childhood Cancer Biology and Pediatric Oncology—INCT BioOncoPed, Porto Alegre 90050-170, RS, Brazil
| | - Marialva Sinigaglia
- Children’s Cancer Institute, Porto Alegre 90620-110, RS, Brazil
- National Institute of Science and Technology in Childhood Cancer Biology and Pediatric Oncology—INCT BioOncoPed, Porto Alegre 90050-170, RS, Brazil
| | - Rafael Roesler
- Cancer and Neurobiology Laboratory, Experimental Research Center, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
- National Institute of Science and Technology in Childhood Cancer Biology and Pediatric Oncology—INCT BioOncoPed, Porto Alegre 90050-170, RS, Brazil
- Department of Pharmacology, Institute for Basic Health Sciences, Federal University of Rio Grande do Sul, Porto Alegre 90050-170, RS, Brazil
| | - Mariane da Cunha Jaeger
- Cancer and Neurobiology Laboratory, Experimental Research Center, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
- Children’s Cancer Institute, Porto Alegre 90620-110, RS, Brazil
- National Institute of Science and Technology in Childhood Cancer Biology and Pediatric Oncology—INCT BioOncoPed, Porto Alegre 90050-170, RS, Brazil
| | - Marcelo Land
- National Institute of Science and Technology in Childhood Cancer Biology and Pediatric Oncology—INCT BioOncoPed, Porto Alegre 90050-170, RS, Brazil
- Department of Pediatrics, Faculty of Medicine, Federal University of Rio de Janeiro, Rio de Janeiro 21941-912, RJ, Brazil
- Martagão Gesteira Childcare and Pediatrics Institute, Federal University of Rio de Janeiro, Rio de Janeiro 21941-912, RJ, Brazil
| | - Caroline Brunetto de Farias
- Cancer and Neurobiology Laboratory, Experimental Research Center, Porto Alegre’s Clinical Hospital, Federal University of Rio Grande do Sul, Porto Alegre 90035-903, RS, Brazil
- Postgraduate Program in Pharmacology and Therapeutics, Institute for Basic Health Sciences, Federal University of Rio Grande do Sul, Porto Alegre 90050-170, RS, Brazil
- Children’s Cancer Institute, Porto Alegre 90620-110, RS, Brazil
- National Institute of Science and Technology in Childhood Cancer Biology and Pediatric Oncology—INCT BioOncoPed, Porto Alegre 90050-170, RS, Brazil
- Martagão Gesteira Childcare and Pediatrics Institute, Federal University of Rio de Janeiro, Rio de Janeiro 21941-912, RJ, Brazil
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Reierson MM, Acharjee A. Unsupervised machine learning-based stratification and immune deconvolution of liver hepatocellular carcinoma. BMC Cancer 2025; 25:853. [PMID: 40349011 PMCID: PMC12066050 DOI: 10.1186/s12885-025-14242-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/06/2024] [Accepted: 04/29/2025] [Indexed: 05/14/2025] Open
Abstract
BACKGROUND Hepatocellular carcinoma (HCC) is the most prevalent type of liver cancer and a leading cause of cancer-related deaths globally. The tumour microenvironment (TME) influences treatment response and prognosis, yet its heterogeneity remains unclear. METHODS The unsupervised machine learning methods- agglomerative hierarchical clustering, Multi-Omics Factor Analysis with K-means++, and an autoencoder with K-means++ - stratified patients using microarray data from HCC samples. Immune deconvolution algorithms estimated the proportions of infiltrating immune cells across identified clusters. RESULTS Thirteen genes were found to influence HCC subtyping in both primary and validation datasets, with three genes-TOP2A, DCN, and MT1E-showing significant associations with survival and recurrence. DCN, a known tumour suppressor, was significant across datasets and associated with improved survival, potentially by modulating the TME and promoting an anti-tumour immune response. CONCLUSIONS The discovery of the 13 conserved genes is an important step toward understanding HCC heterogeneity and the TME, potentially leading to the identification of more reliable biomarkers and therapeutic targets. We have stratified and validated the liver cancer populations. The findings suggest further research is needed to explore additional factors influencing the TME beyond gene expression, such as tumour microbiome and stromal cell interactions.
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Affiliation(s)
- Mae Montserrat Reierson
- Cancer and Genomic Sciences, School of Medical Sciences, College of Medicine and Health, University of Birmingham, Birmingham, B15 2TT, UK
| | - Animesh Acharjee
- Cancer and Genomic Sciences, School of Medical Sciences, College of Medicine and Health, University of Birmingham, Birmingham, B15 2TT, UK.
- Institute of Translational Medicine, University Hospitals Birmingham NHS Foundation Trust, Birmingham, B15 2TT, UK.
- MRC Health Data Research UK (HDR), Midlands Site, UK.
- Centre for Health Data Research, University of Birmingham, Birmingham, B15 2TT, UK.
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Pickard BS. A mechanism of global gene expression regulation is disrupted by multiple disease states and drug treatments. PLoS One 2025; 20:e0317071. [PMID: 40341320 PMCID: PMC12061403 DOI: 10.1371/journal.pone.0317071] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2024] [Accepted: 02/25/2025] [Indexed: 05/10/2025] Open
Abstract
Conventional expression studies quantify messenger RNA (mRNA) transcript levels gene-by-gene. We recently showed that protein expression is modulated at a global scale by amino acid availability, suggesting that mRNA expression levels might be equivalently affected. Through re-analysis of public transcriptomic datasets, it was confirmed that nucleobase supply interacts with the specific demands of mRNA A + U:C + G sequence composition to shape a global profile of expression, which can be quantified as a gradient of average expression change by average composition change. In mammals, each separate organ and cell-type displays a distinct baseline profile of global expression. These profiles can shift dynamically across the circadian day and the menstrual cycle. They are also significantly distorted by viral infection, multiple complex genetic disorders (including Alzheimer's disease, schizophrenia, and autoimmune disorders), and after treatment with 115 of the 597 chemical entities analysed. These included known toxins and nucleobase analogues, but also many commonly prescribed medications such as antibiotics and proton pump inhibitors, thus revealing a new mechanism of drug action and side-effect. As well as key roles in disease susceptibility, mRNAs with extreme compositions are significantly over-represented in gene ontologies such as transcription and cell division, making these processes particularly sensitive to swings in global expression. This may permit efficient, en bloc transcriptional reprogramming of cell state through simple adjustment of nucleobase proportion and supply. It is also proposed that this mechanism helped mitigate the loss of essential amino acid synthesis in higher organisms. In summary, global expression regulation is invisible to conventional transcriptomic analysis, but its measurement allows a useful distinction between active, promoter-mediated gene expression changes and passive, cell state-dependent transcriptional competence. Linking cell metabolism directly to gene expression offers an entirely new perspective on evolution, disease aetiopathology (including gene x environment - GxE - interactions), and the nature of the pharmacological response.
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Affiliation(s)
- Benjamin S. Pickard
- Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow, United Kingdom
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48
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Lei X, Qiu L, Chen Q, Liao L, Yu P, Wu W, Zhu Z, Li C, Lin G, Zhuang Z, Meng Y, Wang Y, Wang C, Du Y. Exploring the regulatory mechanism of CCNA2 in colorectal cancer: insights from multiomics and experimental analysis. J Biol Chem 2025:110216. [PMID: 40345591 DOI: 10.1016/j.jbc.2025.110216] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2024] [Revised: 03/18/2025] [Accepted: 04/04/2025] [Indexed: 05/11/2025] Open
Abstract
Colorectal cancer (CRC) is the third-most common cancer and the second-leading cause of mortality due to cancer worldwide. The underlying regulatory mechanism of CCNA2 in CRC was explored through multiomics and experimental analyses, thus facilitating diagnosis, therapy and prognosis. Two gene expression datasets (i.e., GSE9348 and GSE110223) were extracted from GEO. Differentially expressed genes (DEGs) were identified via GEO2R, which were used for enrichment analyses through DAVID. PPI network of DEGs was constructed by STRING, and the core genes were identified. CCNA2, a prognostic core gene for CRC, was validated in TCGA and HPA via transcriptomics and proteomics. ROC analysis was performed to evaluate the diagnostic value of CCNA2 in CRC. The therapeutic value of CCNA2 was evaluated in DGIdb through pharmacogenomics. The correlation between CCNA2 and immune infiltration was determined in TIMER by immunomics. TF-mRNA and miRNA-mRNA networks for CCNA2 were constructed in miRnet and miRDB via transcriptomics. The role and mechanism of CCNA2 in CRC were investigated both in vitro and in vivo. The miR-548x-3p/CCNA2 regulatory axis in CRC was investigated in vitro. CCNA2 showed excellent diagnostic, therapeutic, and prognostic value in CRC. CCNA2 was closely associated with tumor-infiltrating immunocytes, TFs, and miRNAs. The upregulation of CCNA2 was observed in CRC, and the knockdown of CCNA2 inhibited the proliferation, migration, and invasion while inducing apoptosis of CRC cells. The knockdown of CCNA2 could inhibit epithelial-mesenchymal transition (EMT) pathway. CCNA2 acted as a target of miR-548x-3p in regulating the biological behavior of CRC cells via the EMT-signaling pathway. CCNA2 is a potential biomarker for the diagnosis, treatment, and prognosis of CRC and is associated with immune infiltration, TF, and miRNA. The miR-548x-3p/CCNA2 axis plays a pivotal role in regulating the tumorigenesis of CRC through the EMT-signaling pathway.
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Affiliation(s)
- Xinyi Lei
- Department of Gastric Surgery, Zhejiang Cancer Hospital, Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China; Department of Gastrointestinal Surgery, the First Affiliated Hospital of Jinan University, Guangzhou, Guangdong 510630, China
| | - Lanying Qiu
- Department of Chest Radiotherapy, Zhejiang Cancer Hospital, Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China
| | - Qiang Chen
- Department of Oncology, Cancer Diagnosis and Therapy Research Center, the First Affiliated Hospital of Jinan University, Guangzhou, Guangdong 510630, China
| | - Lan Liao
- Department of Pathology, the First Affiliated Hospital, University of South China, Hengyang, Hunan 421001, China
| | - Pengfei Yu
- Department of Gastric Surgery, Zhejiang Cancer Hospital, Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China
| | - Wenjie Wu
- Department of Radiation Physics, Zhejiang Cancer Hospital, Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China
| | - Zhengyang Zhu
- Department of Integrated Traditional Chinese and Western Medicine, Zhejiang Cancer Hospital, Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China
| | - Chunying Li
- Zhejiang Cancer Hospital, Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China
| | - Gang Lin
- School of Molecular Medicine, Hangzhou Institute for Advanced Study, University of Chinese Academy of Sciences (UCAS), Hangzhou, Zhejiang 310024, China; Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China
| | - Zirui Zhuang
- School of Molecular Medicine, Hangzhou Institute for Advanced Study, University of Chinese Academy of Sciences (UCAS), Hangzhou, Zhejiang 310024, China; Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China
| | - Yuxin Meng
- College of Pharmaceutical Science, Zhejiang University of Technology, Hangzhou, Zhejiang 310014, China
| | - Yan Wang
- Collaborative Innovation Center of Yangtza River Delta Region Green Pharmaceuticals, Zhejiang University of Technology, Hangzhou, Zhejiang 310014, China
| | - Cunchuan Wang
- Department of Gastrointestinal Surgery, the First Affiliated Hospital of Jinan University, Guangzhou, Guangdong 510630, China.
| | - Yian Du
- Department of Gastric Surgery, Zhejiang Cancer Hospital, Hangzhou Institute of Medicine (HIM), Chinese Academy of Sciences, Hangzhou, Zhejiang 310022, China.
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Tsurukawa FK, Mao Y, Sanchez-Villalobos C, Khanna N, Crasto CJ, Lawrence JJ, Pal R. Cross study transcriptomic investigation of Alzheimer's brain tissue discoveries and limitations. Sci Rep 2025; 15:16041. [PMID: 40341634 PMCID: PMC12062235 DOI: 10.1038/s41598-025-01017-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2024] [Accepted: 05/02/2025] [Indexed: 05/10/2025] Open
Abstract
Developing effective treatments for Alzheimer's disease (AD) likely requires a deep understanding of molecular mechanisms. Integration of transcriptomic datasets and developing innovative computational analyses may yield novel molecular targets with broad applicability. The motivation for this study was conceived from two main observations: (a) most transcriptomic analyses of AD data consider univariate differential expression analysis, and (b) insights are often not transferable across studies. We designed a machine learning-based framework that can elucidate interpretable multivariate relationships from multiple human AD studies to discover robust transcriptomic AD biomarkers transferable across multiple studies. Our analysis of three human hippocampus datasets revealed multiple robust synergistic associations from unrelated pathways along with inconsistencies of gene associations across different studies. Our study underscores the utility of developing AI-assisted next-gen metrics for integration, robustness, and generalization and also highlights the potential benefit of elucidating molecular mechanisms and pathways that are important in targeting a single population.
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Affiliation(s)
- Fernando Koiti Tsurukawa
- Department of Electrical and Computer Engineering, Texas Tech University, Lubbock, TX, 79409, USA
| | - Yixiang Mao
- Department of Electrical and Computer Engineering, Texas Tech University, Lubbock, TX, 79409, USA
| | - Cesar Sanchez-Villalobos
- Department of Electrical and Computer Engineering, Texas Tech University, Lubbock, TX, 79409, USA
| | - Nishtha Khanna
- Center for Biotechnology and Genomics, Texas Tech University, Lubbock, TX, 79409, USA
| | - Chiquito J Crasto
- Center for Biotechnology and Genomics, Texas Tech University, Lubbock, TX, 79409, USA
| | - J Josh Lawrence
- Department of Pharmacology and Neuroscience, Garrison Institute on Aging, Center of Excellence for Translational Neuroscience and Therapeutics, and Center of Excellence in Integrative Health, Texas Tech University Health Sciences Center, Lubbock, TX, 79430, USA.
| | - Ranadip Pal
- Department of Electrical and Computer Engineering, Texas Tech University, Lubbock, TX, 79409, USA.
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50
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Bennett J, Levine AB, Nobre L, Negm L, Chung J, Fang K, Johnson M, Komosa M, Krumholtz S, Nunes NM, Rana M, Ryall S, Sheth J, Siddaway R, Bale TA, Bouffet E, Cusimano MD, Das S, Detsky J, Dirks P, Karajannis MA, Kongkham P, Giantini-Larsen A, Li BK, Lim-Fat MJ, Lin AL, Mason WP, Miller A, Perry JR, Sahgal A, Sait SF, Tsang DS, Zadeh G, Laperriere N, Nguyen L, Gao A, Keith J, Munoz DG, Tabori U, Hawkins C. A population-based analysis of the molecular landscape of glioma in adolescents and young adults reveals insights into gliomagenesis. NATURE CANCER 2025:10.1038/s43018-025-00962-x. [PMID: 40335748 DOI: 10.1038/s43018-025-00962-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/02/2024] [Accepted: 03/28/2025] [Indexed: 05/09/2025]
Abstract
Gliomas are a major cause of cancer-related deaths in adolescents and young adults (AYAs; ages 15-39 years). Different molecular alterations drive gliomas in children and adults, leading to distinct biology and clinical consequences, but the implications of pediatric- versus adult-type alterations in AYAs are unknown. Our population-based analysis of 1,456 clinically and molecularly characterized gliomas in patients aged 0-39 years addresses this gap. Pediatric-type alterations were found in 31% of AYA gliomas and conferred superior outcomes compared to adult-type alterations. AYA low-grade gliomas with specific RAS-MAPK alterations exhibited senescence, tended to arise in different locations and were associated with superior outcomes compared to gliomas in children, suggesting different cellular origins. Hemispheric IDH-mutant, BRAF p.V600E and FGFR-altered gliomas were associated with the risk of malignant transformation, having worse outcomes with increased age. These insights into gliomagenesis may provide a rationale for earlier intervention for certain tumors to disrupt the typical behavior, leading to improved outcomes.
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Affiliation(s)
- Julie Bennett
- Division of Haematology/Oncology, The Hospital for Sick Children, Toronto, Ontario, Canada.
- Division of Medical Oncology and Hematology, Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario, Canada.
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada.
| | - Adrian B Levine
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
- Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
| | - Liana Nobre
- Division of Hematology/Oncology (iHOPE), Department of Pediatrics, Stollery Children's Hospital, University of Alberta, Edmonton, Alberta, Canada
| | - Logine Negm
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Jiil Chung
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Karen Fang
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Monique Johnson
- Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Martin Komosa
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Stacey Krumholtz
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Nuno Miguel Nunes
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Mansuba Rana
- Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Scott Ryall
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
- Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Javal Sheth
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
- Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Robert Siddaway
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
- Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Tejus A Bale
- Department of Pathology and Laboratory Medicine, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Eric Bouffet
- Division of Haematology/Oncology, The Hospital for Sick Children, Toronto, Ontario, Canada
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Michael D Cusimano
- Division of Neurosurgery, Unity Health, Toronto, Ontario, Canada
- Division of Neurosurgery, The Hospital for Sick Children, Toronto, Ontario, Canada
- Department of Neurosurgery, Toronto Western Hospital, University Health Network, Toronto, Ontario, Canada
| | - Sunit Das
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
- Division of Neurosurgery, Unity Health, Toronto, Ontario, Canada
| | - Jay Detsky
- Department of Radiation Oncology, Sunnybrook Health Science Centre, Toronto, Ontario, Canada
| | - Peter Dirks
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
- Division of Neurosurgery, The Hospital for Sick Children, Toronto, Ontario, Canada
| | | | - Paul Kongkham
- Department of Neurosurgery, Toronto Western Hospital, University Health Network, Toronto, Ontario, Canada
| | | | - Bryan Kincheon Li
- Department of Pediatrics, Memorial Sloan Kettering Cancer Center, New York, NY, USA
- Division of Pediatric Hematology/Oncology, Icahn School of Medicine at Mount Sinai, New York, NY, USA
| | - Mary Jane Lim-Fat
- Division of Neurology, Department of Medicine, Sunnybrook Health Sciences Centre, Toronto, Ontario, Canada
| | - Andrew L Lin
- Department of Neurology, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Warren P Mason
- Department of Medicine, Princess Margaret Cancer Centre, University Health Network and University of Toronto, Toronto, Ontario, Canada
| | - Alexandra Miller
- Department of Pediatrics, Memorial Sloan Kettering Cancer Center, New York, NY, USA
- Department of Neurology, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - James R Perry
- Division of Neurology, Department of Medicine, Sunnybrook Health Sciences Centre, Toronto, Ontario, Canada
| | - Arjun Sahgal
- Department of Radiation Oncology, Sunnybrook Health Science Centre, Toronto, Ontario, Canada
| | - Sameer Farouk Sait
- Department of Pediatrics, Memorial Sloan Kettering Cancer Center, New York, NY, USA
| | - Derek S Tsang
- Radiation Medicine Program, Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario, Canada
| | - Gelareh Zadeh
- Department of Neurosurgery, Toronto Western Hospital, University Health Network, Toronto, Ontario, Canada
| | - Normand Laperriere
- Radiation Medicine Program, Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario, Canada
| | - Lananh Nguyen
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
- Department of Laboratory Medicine, Unity Health, Toronto, Ontario, Canada
| | - Andrew Gao
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
- Laboratory Medicine Program, University Health Network, Toronto, Ontario, Canada
| | - Julia Keith
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
- Laboratory Medicine and Molecular Diagnostics, Sunnybrook Health Sciences Centre, Toronto, Ontario, Canada
| | - David G Munoz
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
- Department of Laboratory Medicine, Unity Health, Toronto, Ontario, Canada
| | - Uri Tabori
- Division of Haematology/Oncology, The Hospital for Sick Children, Toronto, Ontario, Canada
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
| | - Cynthia Hawkins
- Arthur and Sonia Labatt Brain Tumour Research Centre, The Hospital for Sick Children, Toronto, Ontario, Canada
- Department of Paediatric Laboratory Medicine, The Hospital for Sick Children, Toronto, Ontario, Canada
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada
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