|
1
|
Kalmer M, Grasshoff M, Maié T, Pannen K, Toledo MA, Vieri M, Olschok K, Lemanzyk R, Lazarevic J, Junge B, Baumeister J, Galauner A, Chapal Ilani N, Bar D, Colin E, Cheng M, Schifflers J, Kricheldorf K, Schemionek M, Brümmendorf TH, Weiskirchen R, Shlush L, Zenke M, Chatain N, Costa IG, Koschmieder S. Deciphering the complex clonal heterogeneity of polycythemia vera and the response to interferon alfa. Blood Adv 2025; 9:1873-1887. [PMID: 39874500 PMCID: PMC12008703 DOI: 10.1182/bloodadvances.2024012600] [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: 01/08/2024] [Revised: 12/04/2024] [Accepted: 12/31/2024] [Indexed: 01/30/2025] Open
Abstract
ABSTRACT Interferon alfa (IFN-α) is approved for the therapy of patients with polycythemia vera (PV), a subtype of myeloproliferative neoplasm (MPN). Some patients achieve molecular responses (MRs), but clonal factors sensitizing for MRs remain elusive. We integrated colony formation assays with single-cell RNA sequencing (scRNA-seq) and genotyping in PV-derived cells and healthy controls (HCs) to dissect how IFN-α targets diseased clones during erythroid differentiation. IFN-α significantly decreased colony growth in MPNs and HCs with variable transcriptional responses observed in individual colonies. scRNA-seq of colonies demonstrated more mature erythroid colonies in PV than HCs. JAK2V617F-mutant cells exhibited upregulated STAT5A, heme, and G2M checkpoint pathways compared with JAK2WT cells from the same patients. Subgroup analysis revealed that IFN-α significantly decreased immature erythrocytic cells in PV (basophilic erythroblasts P < .05; polychromatic erythroblasts P < .05) but not in HCs. CD71-/CD235a+ cells from HCs (P < .05) but not PV were inhibited by IFN-α, and the number of reticulocytes was less affected in PV. Robust IFN-α responses persisted throughout differentiation, leading to significant apoptosis in PV. Apoptotic cells displayed downregulation of ribosomal genes. This link between apoptosis and ribosomal genes was corroborated through the analysis of mitochondrial variants, demonstrating IFN-α-induced eradication of specific clones, characterized by elevated expression of ribosomal genes. Our findings indicate that PV-derived clones either undergo apoptosis or pass through differentiation, overall reducing the cycling mutant cells over long-term treatment. Furthermore, the significance of ribosomal genes and clonal prerequisites in IFN-α's therapeutic mechanism is underscored, shedding light on the intricate dynamics of IFN-α treatment in PV.
Collapse
Affiliation(s)
- Milena Kalmer
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Martin Grasshoff
- Institute for Computational Genomics, RWTH Aachen University, Aachen, Germany
| | - Tiago Maié
- Institute for Computational Genomics, RWTH Aachen University, Aachen, Germany
| | - Kristina Pannen
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Marcelo A.S. Toledo
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Margherita Vieri
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Kathrin Olschok
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Rebecca Lemanzyk
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Jelena Lazarevic
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Baerbel Junge
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Julian Baumeister
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Angela Galauner
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Noa Chapal Ilani
- Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, Israel
| | - Dror Bar
- Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, Israel
| | - Elia Colin
- Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, Israel
| | - Mingbo Cheng
- Institute for Computational Genomics, RWTH Aachen University, Aachen, Germany
| | - Joelle Schifflers
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Kim Kricheldorf
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Mirle Schemionek
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Tim H. Brümmendorf
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Ralf Weiskirchen
- Institute of Molecular Pathobiochemistry, Experimental Gene Therapy and Clinical Chemistry, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
| | - Liran Shlush
- Department of Molecular Cell Biology, Weizmann Institute of Science, Rehovot, Israel
| | - Martin Zenke
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Nicolas Chatain
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| | - Ivan G. Costa
- Institute for Computational Genomics, RWTH Aachen University, Aachen, Germany
| | - Steffen Koschmieder
- Department of Hematology, Oncology, Hemostaseology, and Stem Cell Transplantation, Faculty of Medicine, RWTH Aachen University, Aachen, Germany
- Center for Integrated Oncology Aachen Bonn Cologne Düsseldorf, Aachen, Germany
| |
Collapse
|
|
2
|
Geng S, Zhang Z, Fan J, Sun H, Yang J, Luo J, Guan G, Yin H, Zeng Q, Niu Q. Transcriptome Profiling Reveals That the African Swine Fever Virus C315R Exploits the IL-6 STAT3 Signaling Axis to Facilitate Virus Replication. Viruses 2025; 17:309. [PMID: 40143240 PMCID: PMC11945413 DOI: 10.3390/v17030309] [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: 12/23/2024] [Revised: 02/17/2025] [Accepted: 02/21/2025] [Indexed: 03/28/2025] Open
Abstract
African swine fever (ASF) is an acute and highly contagious disease that has caused great losses in the past years. It is caused by African swine fever virus (ASFV), which is a large DNA virus encoding about 165 genes. It has been shown that the purified extracellular ASFV is internalized by both constitutive macropinocytosis and clathrin-mediated endocytosis, and the virus utilizes apoptotic bodies for infection and cell cell transmission. The ASFV-encoded RNA polymerase subunit C315R is thought to play an important role in ASFV replication and transcription. However, its involvement in ASFV infection, particularly in host response, remains only partially understood. In this study, the role of C315R in enhancing ASFV replication was investigated through RNA-Seq transcriptomic analysis, which was based on 3D4/21 cells transfected the plasmid expressing HA-tagged C315R or the empty vector. Our findings revealed that C315R significantly upregulates the expression of inflammatory mediators with a particular emphasis on IL-6. The most differentially expressed genes (DEGs) were predominantly associated with the TNF, IL-17, MAPK, and JAK STAT signaling pathways. RNA-seq results were validated through RT-PCR. Subsequently, we observed that ASFV infection increases IL-6 expression and STAT3 phosphorylation, which is regulated by the ASFV C315R protein. Notably, inhibiting STAT3 phosphorylation with specific inhibitors suppressed ASFV replication. In conclusion, our study demonstrates that the ASFV C315R protein actives STAT3 phosphorylation through promoting the transcription of IL-6 to facilitate virus replication. These findings highlight C315R as a positive regulator in the IL-6 STAT3 signaling axis during ASFV infection.
Collapse
Affiliation(s)
- Shuxian Geng
- College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China;
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou University, Lanzhou 730046, China; (Z.Z.); (H.S.); (J.Y.); (J.L.); (G.G.); (H.Y.)
- African Swine Fever Regional Laboratory of China (Lanzhou), Gansu Province Research Center for Basic Disciplines of Pathogen Biology, Lanzhou 730046, China
| | - Zhonghui Zhang
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou University, Lanzhou 730046, China; (Z.Z.); (H.S.); (J.Y.); (J.L.); (G.G.); (H.Y.)
- African Swine Fever Regional Laboratory of China (Lanzhou), Gansu Province Research Center for Basic Disciplines of Pathogen Biology, Lanzhou 730046, China
| | - Jie Fan
- College of Medicine, Northwest Minzu University, Lanzhou 730030, China;
| | - Hualin Sun
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou University, Lanzhou 730046, China; (Z.Z.); (H.S.); (J.Y.); (J.L.); (G.G.); (H.Y.)
- African Swine Fever Regional Laboratory of China (Lanzhou), Gansu Province Research Center for Basic Disciplines of Pathogen Biology, Lanzhou 730046, China
| | - Jifei Yang
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou University, Lanzhou 730046, China; (Z.Z.); (H.S.); (J.Y.); (J.L.); (G.G.); (H.Y.)
- African Swine Fever Regional Laboratory of China (Lanzhou), Gansu Province Research Center for Basic Disciplines of Pathogen Biology, Lanzhou 730046, China
| | - Jianxun Luo
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou University, Lanzhou 730046, China; (Z.Z.); (H.S.); (J.Y.); (J.L.); (G.G.); (H.Y.)
- African Swine Fever Regional Laboratory of China (Lanzhou), Gansu Province Research Center for Basic Disciplines of Pathogen Biology, Lanzhou 730046, China
| | - Guiquan Guan
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou University, Lanzhou 730046, China; (Z.Z.); (H.S.); (J.Y.); (J.L.); (G.G.); (H.Y.)
- African Swine Fever Regional Laboratory of China (Lanzhou), Gansu Province Research Center for Basic Disciplines of Pathogen Biology, Lanzhou 730046, China
| | - Hong Yin
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou University, Lanzhou 730046, China; (Z.Z.); (H.S.); (J.Y.); (J.L.); (G.G.); (H.Y.)
- African Swine Fever Regional Laboratory of China (Lanzhou), Gansu Province Research Center for Basic Disciplines of Pathogen Biology, Lanzhou 730046, China
- Jiangsu Co-Innovation Center for the Prevention and Control of Important Animal Infectious Disease and Zoonosis, Yangzhou University, Yangzhou 225009, China
| | - Qiaoying Zeng
- College of Veterinary Medicine, Gansu Agricultural University, Lanzhou 730070, China;
| | - Qingli Niu
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou University, Lanzhou 730046, China; (Z.Z.); (H.S.); (J.Y.); (J.L.); (G.G.); (H.Y.)
- African Swine Fever Regional Laboratory of China (Lanzhou), Gansu Province Research Center for Basic Disciplines of Pathogen Biology, Lanzhou 730046, China
| |
Collapse
|
|
3
|
Bach FA, Muñoz Sandoval D, Mazurczyk M, Themistocleous Y, Rawlinson TA, Harding AC, Kemp A, Silk SE, Barrett JR, Edwards NJ, Ivens A, Rayner JC, Minassian AM, Napolitani G, Draper SJ, Spence PJ. A systematic analysis of the human immune response to Plasmodium vivax. J Clin Invest 2023; 133:e152463. [PMID: 37616070 PMCID: PMC10575735 DOI: 10.1172/jci152463] [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: 06/17/2021] [Accepted: 08/22/2023] [Indexed: 08/25/2023] Open
Abstract
BACKGROUNDThe biology of Plasmodium vivax is markedly different from that of P. falciparum; how this shapes the immune response to infection remains unclear. To address this shortfall, we inoculated human volunteers with a clonal field isolate of P. vivax and tracked their response through infection and convalescence.METHODSParticipants were injected intravenously with blood-stage parasites and infection dynamics were tracked in real time by quantitative PCR. Whole blood samples were used for high dimensional protein analysis, RNA sequencing, and cytometry by time of flight, and temporal changes in the host response to P. vivax were quantified by linear regression. Comparative analyses with P. falciparum were then undertaken using analogous data sets derived from prior controlled human malaria infection studies.RESULTSP. vivax rapidly induced a type I inflammatory response that coincided with hallmark features of clinical malaria. This acute-phase response shared remarkable overlap with that induced by P. falciparum but was significantly elevated (at RNA and protein levels), leading to an increased incidence of pyrexia. In contrast, T cell activation and terminal differentiation were significantly increased in volunteers infected with P. falciparum. Heterogeneous CD4+ T cells were found to dominate this adaptive response and phenotypic analysis revealed unexpected features normally associated with cytotoxicity and autoinflammatory disease.CONCLUSIONP. vivax triggers increased systemic interferon signaling (cf P. falciparum), which likely explains its reduced pyrogenic threshold. In contrast, P. falciparum drives T cell activation far in excess of P. vivax, which may partially explain why falciparum malaria more frequently causes severe disease.TRIAL REGISTRATIONClinicalTrials.gov NCT03797989.FUNDINGThe European Union's Horizon 2020 Research and Innovation programme, the Wellcome Trust, and the Royal Society.
Collapse
Affiliation(s)
- Florian A. Bach
- Institute of Immunology and Infection Research, University of Edinburgh, Edinburgh, United Kingdom
| | - Diana Muñoz Sandoval
- Institute of Immunology and Infection Research, University of Edinburgh, Edinburgh, United Kingdom
- Insitute of Microbiology, Universidad San Francisco de Quito, Quito, Ecuador
| | | | | | | | - Adam C. Harding
- Institute of Immunology and Infection Research, University of Edinburgh, Edinburgh, United Kingdom
| | - Alison Kemp
- Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom
| | - Sarah E. Silk
- The Jenner Institute, University of Oxford, Oxford, United Kingdom
- Department of Biochemistry, University of Oxford, Oxford, United Kingdom
| | - Jordan R. Barrett
- The Jenner Institute, University of Oxford, Oxford, United Kingdom
- Department of Biochemistry, University of Oxford, Oxford, United Kingdom
| | - Nick J. Edwards
- The Jenner Institute, University of Oxford, Oxford, United Kingdom
| | - Alasdair Ivens
- Institute of Immunology and Infection Research, University of Edinburgh, Edinburgh, United Kingdom
| | - Julian C. Rayner
- Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom
| | - Angela M. Minassian
- The Jenner Institute, University of Oxford, Oxford, United Kingdom
- Department of Biochemistry, University of Oxford, Oxford, United Kingdom
| | - Giorgio Napolitani
- MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, and
| | - Simon J. Draper
- The Jenner Institute, University of Oxford, Oxford, United Kingdom
- Department of Biochemistry, University of Oxford, Oxford, United Kingdom
| | - Philip J. Spence
- Institute of Immunology and Infection Research, University of Edinburgh, Edinburgh, United Kingdom
| |
Collapse
|
|
4
|
Keenum MC, Chatterjee P, Atalis A, Pandey B, Jimenez A, Roy K. Single-cell epitope-transcriptomics reveal lung stromal and immune cell response kinetics to nanoparticle-delivered RIG-I and TLR4 agonists. Biomaterials 2023; 297:122097. [PMID: 37001347 PMCID: PMC10192313 DOI: 10.1016/j.biomaterials.2023.122097] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/09/2022] [Revised: 03/14/2023] [Accepted: 03/20/2023] [Indexed: 03/31/2023]
Abstract
Lung-resident and circulatory lymphoid, myeloid, and stromal cells, expressing various pattern recognition receptors (PRRs), detect pathogen- and danger-associated molecular patterns (PAMPs/DAMPs), and defend against respiratory pathogens and injuries. Here, we report the early responses of murine lungs to nanoparticle-delivered PAMPs, specifically the retinoic acid-inducible gene I (RIG-I) agonist poly-U/UC (PUUC), with or without the TLR4 agonist monophosphoryl lipid A (MPLA). Using cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq), we characterized the responses at 4 and 24 h after intranasal administration. Within 4 h, ribosome-associated transcripts decreased in both stromal and immune cells, followed by widespread interferon-stimulated gene (ISG) expression. Using RNA velocity, we show that lung-neutrophils dynamically regulate the synthesis of cytokines like CXCL-10, IL-1α, and IL-1β. Co-delivery of MPLA and PUUC increased chemokine synthesis and upregulated antimicrobial binding proteins targeting iron, manganese, and zinc in many cell types, including fibroblasts, endothelial cells, and epithelial cells. Overall, our results elucidate the early PAMP-induced cellular responses in the lung and demonstrate that stimulation of the RIG-I pathway, with or without TLR4 agonists, induces a ubiquitous microbial defense state in lung stromal and immune cells. Nanoparticle-delivered combination PAMPs may have applications in intranasal antiviral and antimicrobial therapies and prophylaxis.
Collapse
Affiliation(s)
- M Cole Keenum
- Wallace H. Coulter Department of Biomedical Engineering Georgia Institute of Technology and Emory University Atlanta, GA, USA
| | - Paramita Chatterjee
- Marcus Center for Therapeutic Cell Characterization and Manufacturing Georgia Institute of Technology, Atlanta, GA, USA
| | - Alexandra Atalis
- Wallace H. Coulter Department of Biomedical Engineering Georgia Institute of Technology and Emory University Atlanta, GA, USA
| | - Bhawana Pandey
- Wallace H. Coulter Department of Biomedical Engineering Georgia Institute of Technology and Emory University Atlanta, GA, USA
| | - Angela Jimenez
- Wallace H. Coulter Department of Biomedical Engineering Georgia Institute of Technology and Emory University Atlanta, GA, USA
| | - Krishnendu Roy
- Wallace H. Coulter Department of Biomedical Engineering Georgia Institute of Technology and Emory University Atlanta, GA, USA; Marcus Center for Therapeutic Cell Characterization and Manufacturing Georgia Institute of Technology, Atlanta, GA, USA; The Parker H. Petit Institute for Bioengineering and Biosciences Georgia Institute of Technology, Atlanta, GA, USA.
| |
Collapse
|
|
5
|
Dugger DT, Fung M, Hays SR, Singer JP, Kleinhenz ME, Leard LE, Golden JA, Shah RJ, Lee JS, Deiter F, Greenland NY, Jones KD, Langelier CR, Greenland JR. Chronic lung allograft dysfunction small airways reveal a lymphocytic inflammation gene signature. Am J Transplant 2021; 21:362-371. [PMID: 32885581 PMCID: PMC8009189 DOI: 10.1111/ajt.16293] [Citation(s) in RCA: 23] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/09/2020] [Revised: 07/27/2020] [Accepted: 08/16/2020] [Indexed: 01/25/2023]
Abstract
Chronic lung allograft dysfunction (CLAD) is the major barrier to long-term survival following lung transplantation, and new mechanistic biomarkers are needed. Lymphocytic bronchitis (LB) precedes CLAD and has a defined molecular signature. We hypothesized that this LB molecular signature would be associated with CLAD in small airway brushings independent of infection. We quantified RNA expression from small airway brushings and transbronchial biopsies, using RNAseq and digital RNA counting, respectively, for 22 CLAD cases and 27 matched controls. LB metagene scores were compared across CLAD strata by Wilcoxon rank sum test. We performed unbiased host transcriptome pathway and microbial metagenome analysis in airway brushes and compared machine-learning classifiers between the two tissue types. This LB metagene score was increased in CLAD airway brushes (p = .002) and improved prediction of graft failure (p = .02). Gene expression classifiers based on airway brushes outperformed those using transbronchial biopsies. While infection was associated with decreased microbial alpha-diversity (p ≤ .04), neither infection nor alpha-diversity was associated with LB gene expression. In summary, CLAD was associated with small airway gene expression changes not apparent in transbronchial biopsies in this cohort. Molecular analysis of airway brushings for diagnosing CLAD merits further examination in multicenter cohorts.
Collapse
Affiliation(s)
- Daniel T. Dugger
- Department of Medicine, University of California, San Francisco, CA 94143,Veterans Affairs Health Care System, San Francisco, CA 94121
| | - Monica Fung
- Department of Medicine, University of California, San Francisco, CA 94143
| | - Steven R. Hays
- Department of Medicine, University of California, San Francisco, CA 94143
| | - Jonathan P. Singer
- Department of Medicine, University of California, San Francisco, CA 94143
| | | | - Lorriana E. Leard
- Department of Medicine, University of California, San Francisco, CA 94143
| | - Jeffrey A. Golden
- Department of Medicine, University of California, San Francisco, CA 94143
| | - Rupal J. Shah
- Department of Medicine, University of California, San Francisco, CA 94143
| | - Joyce S. Lee
- Department of Medicine, University of Colorado, Denver, CO 80045
| | - Fred Deiter
- Department of Medicine, University of California, San Francisco, CA 94143
| | - Nancy Y. Greenland
- Veterans Affairs Health Care System, San Francisco, CA 94121,Department of Anatomic Pathology, University of California, San Francisco, CA 94143
| | - Kirk D. Jones
- Department of Anatomic Pathology, University of California, San Francisco, CA 94143
| | | | - John R. Greenland
- Department of Medicine, University of California, San Francisco, CA 94143,Veterans Affairs Health Care System, San Francisco, CA 94121
| |
Collapse
|
|
6
|
Kumar SS, Tandberg JI, Penesyan A, Elbourne LDH, Suarez-Bosche N, Don E, Skadberg E, Fenaroli F, Cole N, Winther-Larsen HC, Paulsen IT. Dual Transcriptomics of Host-Pathogen Interaction of Cystic Fibrosis Isolate Pseudomonas aeruginosa PASS1 With Zebrafish. Front Cell Infect Microbiol 2018; 8:406. [PMID: 30524971 PMCID: PMC6262203 DOI: 10.3389/fcimb.2018.00406] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/12/2018] [Accepted: 10/29/2018] [Indexed: 01/09/2023] Open
Abstract
Pseudomonas aeruginosa is a significant cause of mortality in patients with cystic fibrosis (CF). To explore the interaction of the CF isolate P. aeruginosa PASS1 with the innate immune response, we have used Danio rerio (zebrafish) as an infection model. Confocal laser scanning microscopy (CLSM) enabled visualization of direct interactions between zebrafish macrophages and P. aeruginosa PASS1. Dual RNA-sequencing of host-pathogen was undertaken to profile RNA expression simultaneously in the pathogen and the host during P. aeruginosa infection. Following establishment of infection in zebrafish embryos with PASS1, 3 days post infection (dpi), there were 6739 genes found to be significantly differentially expressed in zebrafish and 176 genes in PASS1. A range of virulence genes were upregulated in PASS1, including genes encoding pyoverdine biosynthesis, flagellin, non-hemolytic phospholipase C, proteases, superoxide dismutase and fimbrial subunits. Additionally, iron and phosphate acquisition genes were upregulated in PASS1 cells in the zebrafish. Transcriptional changes in the host immune response genes highlighted phagocytosis as a key response mechanism to PASS1 infection. Transcriptional regulators of neutrophil and macrophage phagocytosis were upregulated alongside transcriptional regulators governing response to tissue injury, infection, and inflammation. The zebrafish host showed significant downregulation of the ribosomal RNAs and other genes involved in translation, suggesting that protein translation in the host is affected by PASS1 infection.
Collapse
Affiliation(s)
- Sheemal S Kumar
- Department of Molecular Sciences, Faculty of Science and Engineering, Macquarie University, Sydney, NSW, Australia
| | - Julia I Tandberg
- Department of Pharmaceutical Biosciences, Centre of Integrative Microbial Evolution, School of Pharmacy, University of Oslo, Oslo, Norway
| | - Anahit Penesyan
- Department of Molecular Sciences, Faculty of Science and Engineering, Macquarie University, Sydney, NSW, Australia
| | - Liam D H Elbourne
- Department of Molecular Sciences, Faculty of Science and Engineering, Macquarie University, Sydney, NSW, Australia
| | - Nadia Suarez-Bosche
- Microscopy Unit, Faculty of Science and Engineering, Macquarie University, Sydney, NSW, Australia
| | - Emily Don
- Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Macquarie University, Sydney, NSW, Australia
| | - Eline Skadberg
- Department of Pharmaceutical Biosciences, Centre of Integrative Microbial Evolution, School of Pharmacy, University of Oslo, Oslo, Norway
| | - Federico Fenaroli
- Department of Biosciences, The Faculty of Mathematic and Natural Sciences, University of Oslo, Oslo, Norway
| | - Nicholas Cole
- Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, Macquarie University, Sydney, NSW, Australia
| | - Hanne Cecilie Winther-Larsen
- Department of Pharmaceutical Biosciences, Centre of Integrative Microbial Evolution, School of Pharmacy, University of Oslo, Oslo, Norway
| | - Ian T Paulsen
- Department of Molecular Sciences, Faculty of Science and Engineering, Macquarie University, Sydney, NSW, Australia
| |
Collapse
|
|
7
|
Almasry SM, Hassan ZA, Elsaed WM, Elbastawisy YM. Structural evaluation of the peritubular sheath of rat's testes after administration of ribavirin: A possible impact on the testicular function. Int J Immunopathol Pharmacol 2017; 30:282-296. [PMID: 28799438 PMCID: PMC5815259 DOI: 10.1177/0394632017726261] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
Effects of ribavirin on the structure of peritubular sheath (PS) of seminiferous tubules and on testicular functions were studied. We found that ribavirin at a dose of 4 mg/kg/day for 4 weeks produced a significant reduction in testosterone level (6.3 ± 0.2; P < 0.001) and in spermatogenic score count (3.8 ± 0.2; P < 0.001) compared to control values. The thickness of PS (17.8 ± 1.13) and tubular lumen perimeter (1024.7 ± 67) was significantly increased compared to controls (10.7 ± 0.70; P < 0.001 and 808 ± 25; P = 0.004, respectively). The length of germinal epithelium (411.8 ± 39) and tubular external diameters (1661.8 ± 115) was significantly reduced compared to control values (708.4 ± 40; P < 0.001 and 2358.8 ± 169; P < 0.001, respectively). The basement membranes (BMs) were thickened with great deposition of collagen. Myoid cells showed altered structure and extracellular matrix revealed disorganization by excessive collagen I and IV accumulation. Testicular damage was established histologically. Evidence of apoptosis was detected in germ cells. There was a significant increase in integrated density of Casp-3 expression (38,121,743 ± 1,763,420; P < 0.001) in seminiferous tubules compared to control (24,788,409 ± 1,900,140). It is concluded that ribavirin can cause alterations of the testicular function and structure with increased apoptosis in the tissues after 4 weeks of administration. The damaging effect could be persuaded by destruction of the peritubular sheath.
Collapse
Affiliation(s)
- Shaima M Almasry
- 1 Department of Anatomy and Embryology, Taibah University, Medina, Saudi Arabia.,2 Department of Anatomy and Embryology, Mansoura University, Mansoura, Egypt
| | - Zeinab A Hassan
- 1 Department of Anatomy and Embryology, Taibah University, Medina, Saudi Arabia.,3 Department of Histology and Cell Biology, Zagazig University, Zagazig, Egypt
| | - Wael M Elsaed
- 1 Department of Anatomy and Embryology, Taibah University, Medina, Saudi Arabia.,2 Department of Anatomy and Embryology, Mansoura University, Mansoura, Egypt
| | - Yasser M Elbastawisy
- 1 Department of Anatomy and Embryology, Taibah University, Medina, Saudi Arabia.,2 Department of Anatomy and Embryology, Mansoura University, Mansoura, Egypt
| |
Collapse
|
|
8
|
Control of Hepatitis B Virus by Cytokines. Viruses 2017; 9:v9010018. [PMID: 28117695 PMCID: PMC5294987 DOI: 10.3390/v9010018] [Citation(s) in RCA: 92] [Impact Index Per Article: 11.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/17/2016] [Revised: 01/13/2017] [Accepted: 01/13/2017] [Indexed: 02/06/2023] Open
Abstract
Hepatitis B virus (HBV) infection remains a major public health problem worldwide with more than 240 million individuals chronically infected. Current treatments can control HBV replication to a large extent, but cannot eliminate HBV infection. Cytokines have been shown to control HBV replication and contribute to HBV cure in different models. Cytokines play an important role in limiting acute HBV infection in patients and mediate a non-cytolytic clearance of the virus. In this review, we summarize the effects of cytokines and cytokine-induced cellular signaling pathways on different steps of the HBV life cycle, and discuss possible strategies that may contribute to the eradication of HBV through innate immune activation.
Collapse
|
|
9
|
Sullivan KD, Lewis HC, Hill AA, Pandey A, Jackson LP, Cabral JM, Smith KP, Liggett LA, Gomez EB, Galbraith MD, DeGregori J, Espinosa JM. Trisomy 21 consistently activates the interferon response. eLife 2016; 5:e16220. [PMID: 27472900 PMCID: PMC5012864 DOI: 10.7554/elife.16220] [Citation(s) in RCA: 202] [Impact Index Per Article: 22.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2016] [Accepted: 07/28/2016] [Indexed: 12/12/2022] Open
Abstract
Although it is clear that trisomy 21 causes Down syndrome, the molecular events acting downstream of the trisomy remain ill defined. Using complementary genomics analyses, we identified the interferon pathway as the major signaling cascade consistently activated by trisomy 21 in human cells. Transcriptome analysis revealed that trisomy 21 activates the interferon transcriptional response in fibroblast and lymphoblastoid cell lines, as well as circulating monocytes and T cells. Trisomy 21 cells show increased induction of interferon-stimulated genes and decreased expression of ribosomal proteins and translation factors. An shRNA screen determined that the interferon-activated kinases JAK1 and TYK2 suppress proliferation of trisomy 21 fibroblasts, and this defect is rescued by pharmacological JAK inhibition. Therefore, we propose that interferon activation, likely via increased gene dosage of the four interferon receptors encoded on chromosome 21, contributes to many of the clinical impacts of trisomy 21, and that interferon antagonists could have therapeutic benefits.
Collapse
Affiliation(s)
- Kelly D Sullivan
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Pharmacology, University of Colorado School of Medicine, Aurora, United States
- Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder, United States
- Howard Hughes Medical Institute, Chevy Chase, United States
| | - Hannah C Lewis
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Pharmacology, University of Colorado School of Medicine, Aurora, United States
| | - Amanda A Hill
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Pharmacology, University of Colorado School of Medicine, Aurora, United States
| | - Ahwan Pandey
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Pharmacology, University of Colorado School of Medicine, Aurora, United States
- Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder, United States
- Howard Hughes Medical Institute, Chevy Chase, United States
| | - Leisa P Jackson
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder, United States
- Howard Hughes Medical Institute, Chevy Chase, United States
| | - Joseph M Cabral
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder, United States
- Howard Hughes Medical Institute, Chevy Chase, United States
| | - Keith P Smith
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
| | - L Alexander Liggett
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, United States
| | - Eliana B Gomez
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder, United States
- Howard Hughes Medical Institute, Chevy Chase, United States
| | - Matthew D Galbraith
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Pharmacology, University of Colorado School of Medicine, Aurora, United States
- Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder, United States
- Howard Hughes Medical Institute, Chevy Chase, United States
| | - James DeGregori
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Aurora, United States
- Department of Pediatrics, University of Colorado School of Medicine, Aurora, United States
- Integrated Department of Immunology, University of Colorado School of Medicine, Aurora, United States
- Section of Hematology, University of Colorado School of Medicine, Aurora, United States
- Department of Medicine, University of Colorado School of Medicine, Aurora, United States
| | - Joaquín M Espinosa
- Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Aurora, United States
- Department of Pharmacology, University of Colorado School of Medicine, Aurora, United States
- Department of Molecular, Cellular and Developmental Biology, University of Colorado Boulder, Boulder, United States
- Howard Hughes Medical Institute, Chevy Chase, United States
| |
Collapse
|
|
10
|
Sherrill-Mix S, Ocwieja KE, Bushman FD. Gene activity in primary T cells infected with HIV89.6: intron retention and induction of genomic repeats. Retrovirology 2015; 12:79. [PMID: 26377088 PMCID: PMC4574318 DOI: 10.1186/s12977-015-0205-1] [Citation(s) in RCA: 38] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/24/2015] [Accepted: 08/28/2015] [Indexed: 02/07/2023] Open
Abstract
Background HIV infection has been reported to alter cellular gene activity, but published studies have commonly assayed transformed cell lines and lab-adapted HIV strains, yielding inconsistent results. Here we carried out a deep RNA-Seq analysis of primary human T cells infected with the low passage HIV isolate HIV89.6. Results Seventeen percent of cellular genes showed altered activity 48 h after infection. In a meta-analysis including four other studies, our data differed from studies of HIV infection in cell lines but showed more parallels with infections of primary cells. We found a global trend toward retention of introns after infection, suggestive of a novel cellular response to infection. HIV89.6 infection was also associated with activation of several human endogenous retroviruses (HERVs) and retrotransposons, of interest as possible novel antigens that could serve as vaccine targets. The most highly activated group of HERVs was a subset of the ERV-9. Analysis showed that activation was associated with a particular variant of ERV-9 long terminal repeats that contains an indel near the U3-R border. These data also allowed quantification of >70 splice forms of the HIV89.6 RNA and specified the main types of chimeric HIV89.6-host RNAs. Comparison to over 100,000 integration site sequences from the same infected cell populations allowed quantification of authentic versus artifactual chimeric reads, showing that 5′ read-in, splicing out of HIV89.6 from the D4 donor and 3′ read-through were the most common HIV89.6-host cell chimeric RNA forms. Conclusions Analysis of RNA abundance after infection of primary T cells with the low passage HIV89.6 isolate disclosed multiple novel features of HIV-host interactions, notably intron retention and induction of transcription of retrotransposons and endogenous retroviruses. Electronic supplementary material The online version of this article (doi:10.1186/s12977-015-0205-1) contains supplementary material, which is available to authorized users.
Collapse
Affiliation(s)
- Scott Sherrill-Mix
- Department of Microbiology, Perelman School of Medicine at the University of Pennsylvania, 425 Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA, 19104, USA.
| | - Karen E Ocwieja
- Children's Hospital of Philadelphia, 3401 Civic Center Blvd., Philadelphia, PA, 19104, USA.
| | - Frederic D Bushman
- Department of Microbiology, Perelman School of Medicine at the University of Pennsylvania, 425 Johnson Pavilion, 3610 Hamilton Walk, Philadelphia, PA, 19104, USA.
| |
Collapse
|
|
11
|
Almeida M, Garc�a-Montero AC, Orfao A. Cell Purification: A New Challenge for Biobanks. Pathobiology 2015; 81:261-275. [DOI: 10.1159/000358306] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022] Open
|
|
12
|
Wang L, Zhang L, Feng X, Xing L, Zhang W, Jiang K, Song H, Ji G. The Functional Difference of Dendritic Cells in HBeAg Negative Chronic Hepatitis B Patients with Three Different Spleen Deficiency Syndromes and the Therapeutic Evaluation of Chinese Medicine Intervention Based on Syndrome Differentiation. EVIDENCE-BASED COMPLEMENTARY AND ALTERNATIVE MEDICINE : ECAM 2014; 2014:802402. [PMID: 25093028 PMCID: PMC4100377 DOI: 10.1155/2014/802402] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 02/12/2014] [Revised: 05/08/2014] [Accepted: 05/28/2014] [Indexed: 01/17/2023]
Abstract
Objective. To investigate the dendritic cells (DCs) maturity differences of HBeAg negative chronic hepatitis B (CHB) patients with different spleen deficiency (SD) syndromes and explore the role of syndrome differentiation in the therapeutic evaluation of Chinese medicine. Methods. 120 participants were recruited including three treatment groups in different SD syndrome categories as spleen deficiency with liver depression (SDLD), spleen deficiency with damp heat (SDDH), and spleen deficiency with kidney deficiency (SDKD) and one healthy control group; each group had 30 participants. Corresponding drugs were applied. The outcome measures included DC phenotype, liver function, IL-10, IL-12, and HBV-DNA levels. Results. The surface markers of mature DCs and cytokines levels were different in each group; the positive rate of CD80, CD1a, HLA-DR, and CD1a was the lowest in SDKD group. After 3-month intervention, the expression of CD80, CD86, CD1a, HLA-DR, and IL-12 significantly increased, while ALT, AST, and IL-10 significantly decreased (P < 0.05) in treatment groups. HBV-DNA level also significantly reduced in both SDKD and SDLD groups (P < 0.05). Conclusions. HBeAg negative patients had DCs dysmaturity, and there were differences between different SD syndromes. Chinese medicine intervention according to syndrome differentiation could advance the maturity and function of DCs and improve the therapeutic effect.
Collapse
Affiliation(s)
- Lei Wang
- Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
- Department of Hepatology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
| | - Li Zhang
- Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
| | - Xiaoxia Feng
- Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
| | - Lianjun Xing
- Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
- Department of Hepatology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
| | - Wei Zhang
- Department of Hepatology, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
| | - Kaiping Jiang
- Department of Hepatology, Foshan Hospital of Traditional Chinese Medicine, Guangdong 528000, China
| | - Haiyan Song
- Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
| | - Guang Ji
- Institute of Digestive Diseases, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200032, China
- E-Institute of Shanghai Municipal Education Commission, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China
| |
Collapse
|
|
13
|
Lee SC, Tan HT, Chung MCM. Prognostic biomarkers for prediction of recurrence of hepatocellular carcinoma: Current status and future prospects. World J Gastroenterol 2014; 20:3112-3124. [PMID: 24696598 PMCID: PMC3964383 DOI: 10.3748/wjg.v20.i12.3112] [Citation(s) in RCA: 68] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/30/2013] [Revised: 12/03/2013] [Accepted: 01/02/2014] [Indexed: 02/07/2023] Open
Abstract
Hepatocellular carcinoma (HCC) is one of the leading causes of cancer death worldwide, with region specific etiologies. Despite improvements made in the diagnosis of HCC, the prognosis of HCC patients remains poor due to the high recurrence rate of HCC. There is an urgent need for development of prognostic biomarkers to predict the risk of recurrence in HCC patients after “curative” treatment. Such stratification may aid in patient management and development of personalized medicine for HCC treatment. Omics based studies facilitate the study of global changes in biomolecules in a disease in a high throughput manner, and hence are well poised to understand the complex changes which led to HCC recurrence. The quantitative nature of data obtained from omics based studies allow for development of prognostic biomarkers based on changes in gene, protein and metabolite expression. In this review, we surveyed the application of transcriptomics, proteomics and metabolomics in the study of HCC recurrence. We summarised the data in the literature from these three fields of studies that claimed to be prognostic for HCC recurrence. We critiqued on the limitations of each area of research and the challenges faced in translating the research results for clinical application in predicting HCC recurrence.
Collapse
|