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1
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Wu X, Cai W, He J, Zhang S, Wang S, Huang L, Zhang H, Sun X, Zhou J, Liu XM. YTHDF2 suppresses the 2C-like state in mouse embryonic stem cells via the DUX-ZSCAN4 molecular circuit. J Biol Chem 2025; 301:108479. [PMID: 40188945 DOI: 10.1016/j.jbc.2025.108479] [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: 01/17/2025] [Revised: 03/21/2025] [Accepted: 03/31/2025] [Indexed: 05/01/2025] Open
Abstract
Mouse embryonic stem cells (ESCs) consist of a rare population of heterogeneous 2-cell-like cells (2CLCs). These cells transiently recapitulate the transcriptional and epigenetic features of the 2-cell embryos, serving as a unique model for studying totipotency acquisition and embryonic development. Accumulating evidence has demonstrated that transcription factors and epigenetic modifications exert crucial functions in the transition of ESCs to 2CLCs. However, the roles of RNA modification in the regulation of the 2C-like state remain elusive. Using a DUX-induced 2CLCs system, we examine N6-methyladenosine (m6A) modification landscape transcriptome-wide and observe dynamic regulation of m6A during DUX-driven 2C-like reprogramming. Notably, many core 2C transcripts like Dux and Zscan4 are highly methylated. We identify the m6A reader protein YTHDF2 as a critical regulator of 2C-like state. Depletion of YTHDF2 facilitates robust expression of 2C-signature genes and ESCs-to-2CLCs transition. Intriguingly, YTHDF2 binds to a subset of m6A-modified 2C transcripts and promotes their decay. We further demonstrate that YTHDF2 suppresses the 2C-like program in a manner that is dependent on both m6A and the DUX-ZSCAN4 molecular circuit. Mechanistically, YTHDF2 interacts with CNOT1, a key component of the RNA deadenylase complex. Consistently, silencing of CNOT1 upregulates the 2C program and promotes ESCs-to-2CLCs transition. Collectively, our findings reveal novel insights into the epitranscriptomic regulation of the 2C-like state in mouse ESCs.
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Affiliation(s)
- Xiang Wu
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China
| | - Wanting Cai
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China
| | - Junjie He
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China
| | - Shiyin Zhang
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China
| | - Shen Wang
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China
| | - Lingci Huang
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China
| | - Haotian Zhang
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China
| | - Xiaoyan Sun
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China
| | - Jun Zhou
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China; Jiangsu Key Laboratory of Drug Design and Optimization, China Pharmaceutical University, Nanjing, China.
| | - Xiao-Min Liu
- School of Life Science and Technology, China Pharmaceutical University, Nanjing, Jiangsu, China; Jiangsu Key Laboratory of Drug Design and Optimization, China Pharmaceutical University, Nanjing, China.
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2
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Moghtaderi H, Mohahammadi S, Sadeghian G, Choudhury M, Al-Harrasi A, Rahman SM. Electrical impedance sensing in stem cell research: Insights, applications, and future directions. PROGRESS IN BIOPHYSICS AND MOLECULAR BIOLOGY 2025; 195:1-14. [PMID: 39557164 DOI: 10.1016/j.pbiomolbio.2024.11.004] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/20/2024] [Revised: 11/14/2024] [Accepted: 11/15/2024] [Indexed: 11/20/2024]
Abstract
The exceptional differentiation abilities of stem cells make them ideal candidates for cell replacement therapies. Considering their great potential, researchers should understand how stem cells interact with other cell types. The production of high-quality differentiated cells is crucial for favorable treatment and makes them an ideal choice for clinical applications. Label-free stem cell monitoring approaches are anticipated to be more effective in this context, as they ensure quality of differentiation while preserving the therapeutic potential. Electric cell-substrate impedance sensing (ECIS) is a nonintrusive technique that enables cell quantification through continuous monitoring of adherent cell behavior using electronic transcellular impedance measurements. This technique also facilitates the study of cell growth, motility, differentiation, drug effects, and cell barrier functions. Therefore, numerous studies have identified ECIS as an effective method for monitoring stem cell quality and differentiation. In this review, we discuss the current understanding of ECIS's achievements in examining cell behaviors and the potential applications of ECIS arrays in preclinical stem cell research. Moreover, we highlight our present knowledge concerning ECIS's contributions in examining cell behaviors and speculate about the future uses of ECIS arrays in preclinical stem cell research. This review also aims to stimulate research on electrochemical biosensors for future applications in regenerative medicine.
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Affiliation(s)
- Hassan Moghtaderi
- Natural and Medical Sciences Research Center, University of Nizwa, Nizwa, 616, Sultanate of Oman
| | - Saeed Mohahammadi
- Natural and Medical Sciences Research Center, University of Nizwa, Nizwa, 616, Sultanate of Oman
| | - Golfam Sadeghian
- Advanced Micro and Nano Device Laboratory, Faculty of New Sciences and Technologies, University of Tehran, Tehran, 1439957131, Iran
| | - Mahua Choudhury
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas A & M University, College Station, TX, 77843, USA
| | - Ahmed Al-Harrasi
- Natural and Medical Sciences Research Center, University of Nizwa, Nizwa, 616, Sultanate of Oman
| | - Shaikh Mizanoor Rahman
- Natural and Medical Sciences Research Center, University of Nizwa, Nizwa, 616, Sultanate of Oman.
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3
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Schüle KM, Probst S. Epigenetic control of cell identities from epiblast to gastrulation. FEBS J 2025. [PMID: 39985220 DOI: 10.1111/febs.70024] [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: 08/22/2024] [Revised: 01/20/2025] [Accepted: 02/04/2025] [Indexed: 02/24/2025]
Abstract
Epigenetic modifications of chromatin are essential for the establishment of cell identities during embryogenesis. Between embryonic days 3.5-7.5 of murine development, major cell lineage decisions are made that discriminate extraembryonic and embryonic tissues, and the embryonic primary germ layers are formed, thereby laying down the basic body plan. In this review, we cover the contribution of dynamic chromatin modifications by DNA methylation, changes of chromatin accessibility, and histone modifications, that in combination with transcription factors control gene expression programs of different cell types. We highlight the differences in regulation of enhancer and promoter marks and discuss their requirement in cell lineage specification. Importantly, in many cases, lineage-specific targeting of epigenetic modifiers is carried out by pioneer or master transcription factors, that in sum mediate the chromatin landscape and thereby control the transcription of cell-type-specific gene programs and thus, cell identities.
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Affiliation(s)
- Katrin M Schüle
- Faculty of Medicine, Institute of Experimental and Clinical Pharmacology and Toxicology, University of Freiburg, Germany
- Signaling Research Centers BIOSS and CIBSS, University of Freiburg, Germany
| | - Simone Probst
- Faculty of Medicine, Institute of Experimental and Clinical Pharmacology and Toxicology, University of Freiburg, Germany
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4
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Ferguson R, Subramanian V. Targeted Deletion in the Basal Body Protein Talpid3 Leads to Loss of Primary Cilia in Embryonic Stem Cells and Defective Lineage-Specific Differentiation. Cells 2024; 13:1957. [PMID: 39682705 PMCID: PMC11639927 DOI: 10.3390/cells13231957] [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: 10/09/2024] [Revised: 11/14/2024] [Accepted: 11/23/2024] [Indexed: 12/18/2024] Open
Abstract
Talpid3 is a basal body protein required for the formation of primary cilia, an organelle involved in signal transduction. Here, we asked if Talpid3 has a role in the regulation of differentiation and/or self-renewal of ES cells and whether cells lacking cilia due to a deletion in Talpid3 can be reprogrammed to induced pluripotent stem (iPS) cells. We show that mouse embryonic limb fibroblasts which lack primary cilia with a targeted deletion in the Talpid3 (Ta3) gene can be efficiently reprogrammed to iPS cells. Furthermore, vector-free Ta3-/- iPS cells retain ES cell features and are able to self-renew. However, both Ta3-/- iPS and ES cells are unable to form visceral endoderm and differentiate poorly into neurons. The observed defects are not a consequence of reprogramming since Ta3-/- ES cells also exhibit this phenotype. Thus, Talpid3 and primary cilia are required for some differentiation events but appear to be dispensable for stem cell self-renewal and reprogramming.
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Affiliation(s)
| | - Vasanta Subramanian
- Department of Life Sciences, University of Bath, Building 4 South, Bath BA2 7AY, UK;
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5
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He X, Tang R, Lou J, Wang R. Pseudo-trajectory inference for identifying essential regulations and molecules in cell fate decisions. J Biol Phys 2024; 51:2. [PMID: 39541052 PMCID: PMC11564433 DOI: 10.1007/s10867-024-09665-3] [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: 02/06/2024] [Accepted: 10/26/2024] [Indexed: 11/16/2024] Open
Abstract
Cell fate decision is crucial in biological development and plays fundamental roles in normal development and functional maintenance of organisms. By identifying key regulatory interactions and molecules involved in these fate decisions, we can shed light on the intricate mechanisms underlying the cell fates. This understanding ultimately reveals the fundamental principles driving biological development and the origins of various diseases. In this study, we present an overarching framework which integrates pseudo-trajectory inference and differential analysis to determine critical regulatory interactions and molecules during cell fate transitions. To demonstrate feasibility and reliability of the approach, we employ the differentiation networks of hepatobiliary system and embryonic stem cells as representative model systems. By applying pseudo-trajectory inference to biological data, we aim to identify critical regulatory interactions and molecules during the cell fate transition processes. Consistent with experimental observations, the approach can allow us to infer dynamical cell fate decision processes and gain insights into the underlying mechanisms which govern cell state decisions.
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Affiliation(s)
- Xinyu He
- Department of Mathematics, Shanghai University, Shanghai, 200444, China
| | - Ruoyu Tang
- Department of Mathematics, Shanghai University, Shanghai, 200444, China
| | - Jie Lou
- Department of Mathematics, Shanghai University, Shanghai, 200444, China.
- Newtouch Center for Mathematics of Shanghai University, Shanghai, 200444, China.
| | - Ruiqi Wang
- Department of Mathematics, Shanghai University, Shanghai, 200444, China.
- Newtouch Center for Mathematics of Shanghai University, Shanghai, 200444, China.
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6
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Arekatla G, Skylaki S, Corredor Suarez D, Jackson H, Schapiro D, Engler S, Auler M, Camargo Ortega G, Hastreiter S, Reimann A, Loeffler D, Bodenmiller B, Schroeder T. Identification of an embryonic differentiation stage marked by Sox1 and FoxA2 co-expression using combined cell tracking and high dimensional protein imaging. Nat Commun 2024; 15:7860. [PMID: 39251590 PMCID: PMC11385471 DOI: 10.1038/s41467-024-52069-z] [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: 05/31/2023] [Accepted: 08/26/2024] [Indexed: 09/11/2024] Open
Abstract
Pluripotent mouse embryonic stem cells (ESCs) can differentiate to all germ layers and serve as an in vitro model of embryonic development. To better understand the differentiation paths traversed by ESCs committing to different lineages, we track individual differentiating ESCs by timelapse imaging followed by multiplexed high-dimensional Imaging Mass Cytometry (IMC) protein quantification. This links continuous live single-cell molecular NANOG and cellular dynamics quantification over 5-6 generations to protein expression of 37 different molecular regulators in the same single cells at the observation endpoints. Using this unique data set including kinship history and live lineage marker detection, we show that NANOG downregulation occurs generations prior to, but is not sufficient for neuroectoderm marker Sox1 upregulation. We identify a developmental cell type co-expressing both the canonical Sox1 neuroectoderm and FoxA2 endoderm markers in vitro and confirm the presence of such a population in the post-implantation embryo. RNASeq reveals cells co-expressing SOX1 and FOXA2 to have a unique cell state characterized by expression of both endoderm as well as neuroectoderm genes suggesting lineage potential towards both germ layers.
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Affiliation(s)
- Geethika Arekatla
- Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland
- Department of Neurosciences, Leuven Brain Institute (LBI), KU Leuven - University of Leuven, Leuven, Belgium
- Laboratory of Neurobiology, VIB Center for Brain & Disease Research, Leuven, Belgium
| | - Stavroula Skylaki
- Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland
| | | | - Hartland Jackson
- Department of Quantitative Biomedicine, University of Zurich, Zurich, Switzerland
- Institute of Molecular Health Sciences, ETH Zürich, Zürich, Switzerland
- Lunenfeld Tanenbaum Research Institute, Mount Sinai Health Systems; Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada
| | - Denis Schapiro
- Department of Quantitative Biomedicine, University of Zurich, Zurich, Switzerland
- Institute of Molecular Health Sciences, ETH Zürich, Zürich, Switzerland
- Institute for Computational Biomedicine, Heidelberg University, Faculty of Medicine, Heidelberg University Hospital, Heidelberg, Germany
- Institute of Pathology, Heidelberg University Hospital, Heidelberg, Germany
- Translational Spatial Profiling Center (TSPC), Heidelberg, Germany
| | - Stefanie Engler
- Department of Quantitative Biomedicine, University of Zurich, Zurich, Switzerland
| | - Markus Auler
- Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland
| | | | - Simon Hastreiter
- Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland
| | - Andreas Reimann
- Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland
| | - Dirk Loeffler
- Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland
- Department of Hematology, St. Jude Children's Research Hospital, Memphis, TN, USA
- Department of Pathology and Laboratory Medicine, The University of Tennessee, Memphis, TN, USA
| | - Bernd Bodenmiller
- Department of Quantitative Biomedicine, University of Zurich, Zurich, Switzerland
- Institute of Molecular Health Sciences, ETH Zürich, Zürich, Switzerland
| | - Timm Schroeder
- Department of Biosystems Science and Engineering, ETH Zurich, Basel, Switzerland.
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7
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Llewellyn J, Charrier A, Cuciniello R, Helfer E, Dono R. Substrate stiffness alters layer architecture and biophysics of human induced pluripotent stem cells to modulate their differentiation potential. iScience 2024; 27:110557. [PMID: 39175774 PMCID: PMC11340605 DOI: 10.1016/j.isci.2024.110557] [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: 02/01/2024] [Revised: 06/10/2024] [Accepted: 07/17/2024] [Indexed: 08/24/2024] Open
Abstract
Lineage-specific differentiation of human induced pluripotent stem cells (hiPSCs) relies on complex interactions between biochemical and physical cues. Here we investigated the ability of hiPSCs to undergo lineage commitment in response to inductive signals and assessed how this competence is modulated by substrate stiffness. We showed that Activin A-induced hiPSC differentiation into mesendoderm and its derivative, definitive endoderm, is enhanced on gel-based substrates softer than glass. This correlated with changes in tight junction formation and extensive cytoskeletal remodeling. Further, live imaging and biophysical studies suggested changes in cell motility and interfacial contacts underlie hiPSC layer reshaping on soft substrates. Finally, we repurposed an ultra-soft silicone gel, which may provide a suitable substrate for culturing hiPSCs at physiological stiffnesses. Our results provide mechanistic insight into how epithelial mechanics dictate the hiPSC response to chemical signals and provide a tool for their efficient differentiation in emerging stem cell therapies.
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Affiliation(s)
- Jack Llewellyn
- Aix Marseille University, CNRS, IBDM, Turing Centre for Living Systems, NeuroMarseille, Marseille, France
- Aix Marseille University, CNRS, CINAM, Turing Centre for Living Systems, 13009 Marseille, France
| | - Anne Charrier
- Aix Marseille University, CNRS, CINAM, Turing Centre for Living Systems, 13009 Marseille, France
| | - Rossana Cuciniello
- Aix Marseille University, CNRS, IBDM, Turing Centre for Living Systems, NeuroMarseille, Marseille, France
| | - Emmanuèle Helfer
- Aix Marseille University, CNRS, CINAM, Turing Centre for Living Systems, 13009 Marseille, France
| | - Rosanna Dono
- Aix Marseille University, CNRS, IBDM, Turing Centre for Living Systems, NeuroMarseille, Marseille, France
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8
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Darsi SP, Baishya S, Nagati V, Bharani KK, Cheekatla SS, Darsi SK, Kamireddy AR, Barra RR, Devarasetti AK, Surampudi S, Singireddy JR, Kandula SK, Pasupulati AK. Safety assessment of rat embryonic fraction for in vivo regenerative therapy. Biol Open 2024; 13:bio060266. [PMID: 38984587 PMCID: PMC11360137 DOI: 10.1242/bio.060266] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/07/2023] [Accepted: 07/03/2024] [Indexed: 07/11/2024] Open
Abstract
Regenerative therapy is considered a novel option for treating various diseases, whereas a developing embryo is a prime source of molecules that help repair diseased tissue and organs. Organoid culture studies also confirmed the inherent biological functions of several embryonic factors. However, the in vivo safety and efficacy of embryonic protein fraction (EPF) were not validated. In this study, we investigated the effectiveness of EPF on healthy adult rats. We obtained embryos from Sprague-Dawley (SD) female rats of E14, E16, and E19 embryonic days and collected protein lysate. This lysate was administered intravenously into adult SD rats on sequential days. We collected blood and performed hematological and biochemical parameters of rats that received EPF. C-reactive protein levels, interleukin-6, blood glucose levels, serum creatinine, blood urea, total leucocyte counts, and % of neutrophils and lymphocytes were comparable between rats receiving EPF and saline. Histological examination of rats' tissues administered with EPF is devoid of abnormalities. Our study revealed that intravenous administration of EPF to healthy adult rats showed that EPF is non-immunogenic, non-inflammatory, non-tumorigenic, and safe for in vivo applications. Our analysis suggests that EPF or its components could be recommended for validating its therapeutic abilities in organ regenerative therapy.
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Affiliation(s)
- Sivarama Prasad Darsi
- Department of Biotechnology, School of Life Sciences, Gitam University, Visakhapatnam, AP, India530045
| | - Somorita Baishya
- Department of Biochemistry, University of Hyderabad, Hyderabad, TG, India500046
| | - Veerababu Nagati
- Department of Biochemistry, University of Hyderabad, Hyderabad, TG, India500046
| | - Kala Kumar Bharani
- Department of Veterinary Pharmacology and Toxicology, P.V. Narasimha Rao University of Veterinary Sciences, Rajendra Nagar, TG, India500030
| | | | - Sujesh Kumar Darsi
- Department of General Medicine, ESI Corporation, Gunadala, Vijayawada, AP, India520004
| | - Adi Reddy Kamireddy
- Department of Internal Medicine, Banner Health Center, Maricopa, AZ, USA85138
| | - Ram Reddy Barra
- Department of Physiology, Apollo Institute of Medical Sciences and Research, Hyderabad, TG 500090, India
| | - Ashok Kumar Devarasetti
- Department of Veterinary Biochemistry, P.V. Narasimha Rao University of Veterinary Sciences, Mamnoor, Warangal, TG, India506166
| | - Sreedhar Surampudi
- Department of Biochemistry, Aware College of Medical Lab Technology, Bairamalguda, Hyderabad 500035, India
| | - Jayaram Reddy Singireddy
- Department of Urology, Hyderabad Kidney & Laparoscopic Centre, Malakpet, Hyderabad, TG 500036, India
| | - Siva Kumar Kandula
- Department of Biotechnology, School of Life Sciences, Gitam University, Visakhapatnam, AP, India530045
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9
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Bello AB, Canlas KKV, Kim D, Park H, Lee SH. Stepwise dual-release microparticles of BMP-4 and SCF in induced pluripotent stem cell spheroids enhance differentiation into hematopoietic stem cells. J Control Release 2024; 371:386-405. [PMID: 38844177 DOI: 10.1016/j.jconrel.2024.06.011] [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: 12/26/2023] [Revised: 06/03/2024] [Accepted: 06/03/2024] [Indexed: 06/11/2024]
Abstract
Recently, the formation of three-dimensional (3D) cell aggregates known as embryoid bodies (EBs) grown in media supplemented with HSC-specific morphogens has been utilized for the directed differentiation of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), into clinically relevant hematopoietic stem cells (HSCs). However, delivering growth factors and nutrients have become ineffective in inducing synchronous differentiation of cells due to their 3D conformation. Moreover, irregularly sized EBs often lead to the formation of necrotic cores in larger EBs, impairing differentiation. Here, we developed two gelatin microparticles (GelMPs) with different release patterns and two HSC-related growth factors conjugated to them. Slow and fast releasing GelMPs were conjugated with bone morphogenic factor-4 (BMP-4) and stem cell factor (SCF), respectively. The sequential presentation of BMP-4 and SCF in GelMPs resulted in efficient and effective hematopoietic differentiation, shown by the enhanced gene and protein expression of several mesoderm and HSC-related markers, and the increased concentration of released HSC-related cytokines. In the present study, we were able to generate CD34+, CD133+, and FLT3+ cells with similar cellular and molecular morphology as the naïve HSCs that can produce colony units of different blood cells, in vitro.
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Affiliation(s)
- Alvin Bacero Bello
- Department of Biomedical Engineering, Dongguk University, Seoul 04620, Republic of Korea; School of Integrative Engineering, Chung-Ang University, Seoul 06911, Republic of Korea
| | | | - Deogil Kim
- Department of Biomedical Engineering, Dongguk University, Seoul 04620, Republic of Korea
| | - Hansoo Park
- School of Integrative Engineering, Chung-Ang University, Seoul 06911, Republic of Korea.
| | - Soo-Hong Lee
- Department of Biomedical Engineering, Dongguk University, Seoul 04620, Republic of Korea.
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10
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Li W, Guo J, Hobson EC, Xue X, Li Q, Fu J, Deng CX, Guo Z. Metabolic-Glycoengineering-Enabled Molecularly Specific Acoustic Tweezing Cytometry for Targeted Mechanical Stimulation of Cell Surface Sialoglycans. Angew Chem Int Ed Engl 2024; 63:e202401921. [PMID: 38498603 PMCID: PMC11073901 DOI: 10.1002/anie.202401921] [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: 01/27/2024] [Revised: 03/14/2024] [Accepted: 03/18/2024] [Indexed: 03/20/2024]
Abstract
In this study, we developed a novel type of dibenzocyclooctyne (DBCO)-functionalized microbubbles (MBs) and validated their attachment to azide-labelled sialoglycans on human pluripotent stem cells (hPSCs) generated by metabolic glycoengineering (MGE). This enabled the application of mechanical forces to sialoglycans on hPSCs through molecularly specific acoustic tweezing cytometry (mATC), that is, displacing sialoglycan-anchored MBs using ultrasound (US). It was shown that subjected to the acoustic radiation forces of US pulses, sialoglycan-anchored MBs exhibited significantly larger displacements and faster, more complete recovery after each pulse than integrin-anchored MBs, indicating that sialoglycans are more stretchable and elastic than integrins on hPSCs in response to mechanical force. Furthermore, stimulating sialoglycans on hPSCs using mATC reduced stage-specific embryonic antigen-3 (SSEA-3) and GD3 expression but not OCT4 and SOX2 nuclear localization. Conversely, stimulating integrins decreased OCT4 nuclear localization but not SSEA-3 and GD3 expression, suggesting that mechanically stimulating sialoglycans and integrins initiated distinctive mechanoresponses during the early stages of hPSC differentiation. Taken together, these results demonstrated that MGE-enabled mATC uncovered not only different mechanical properties of sialoglycans on hPSCs and integrins but also their different mechanoregulatory impacts on hPSC differentiation, validating MGE-based mATC as a new, powerful tool for investigating the roles of glycans and other cell surface biomolecules in mechanotransduction.
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Affiliation(s)
- Weiping Li
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI 48109, USA
| | - Jiatong Guo
- Department of Chemistry, University of Florida, Gainesville, FL 32611, USA
| | - Eric C. Hobson
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI 48109, USA
| | - Xufeng Xue
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, MI 48109, USA
| | - Qingjiang Li
- Department of Chemistry, University of Florida, Gainesville, FL 32611, USA
- Department of Chemistry, University of Massachusetts Boston, Boston, MA 02125, USA
| | - Jianping Fu
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI 48109, USA
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, MI 48109, USA
- Department of Cell and Developmental Biology, University of Michigan, Ann Arbor, MI 48109, USA
| | - Cheri X. Deng
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI 48109, USA
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, MI 48109, USA
| | - Zhongwu Guo
- Department of Chemistry, University of Florida, Gainesville, FL 32611, USA
- UF Health Cancer Center, University of Florida, Gainesville, FL 32611, USA
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11
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Maruyama K, Miyazaki S, Kobayashi R, Hikita H, Tsubone T, Ohnuma K. The migration pattern of cells during the mesoderm and endoderm differentiation from human pluripotent stem cells. In Vitro Cell Dev Biol Anim 2024; 60:535-543. [PMID: 38656570 DOI: 10.1007/s11626-024-00904-4] [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: 12/26/2023] [Accepted: 03/16/2024] [Indexed: 04/26/2024]
Abstract
Gastrulation is the first major differentiation process in animal embryos. However, the dynamics of human gastrulation remain mostly unknown owing to the ethical limitations. We studied the dynamics of the mesoderm and endoderm cell differentiation from human pluripotent stem cells for insight into the cellular dynamics of human gastrulation. Human pluripotent stem cells have properties similar to those of the epiblast, which gives rise to the three germ layers. The mesoderm and endoderm were induced with more than 75% purity from human induced pluripotent stem cells. Single-cell dynamics of pluripotent stem cell-derived mesoderm and endoderm cells were traced using time-lapse imaging. Both mesoderm and endoderm cells migrate randomly, accompanied by short-term directional persistence. No substantial differences were detected between mesoderm and endoderm migration. Computer simulations created using the measured parameters revealed that random movement and external force, such as the spread out of cells from the primitive streak area, mimicked the homogeneous discoidal germ layer formation. These results were consistent with the development of amniotes, which suggests the effectiveness of human pluripotent stem cells as a good model for studying human embryogenesis.
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Affiliation(s)
- Kenshiro Maruyama
- Department of Science of Technology Innovation, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata, 940-2188, Japan
| | - Shota Miyazaki
- Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata, 940-2188, Japan
| | - Ryo Kobayashi
- Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata, 940-2188, Japan
| | - Haru Hikita
- Department of Electrical, Electronics and Information Engineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata, 940-2188, Japan
| | - Tadashi Tsubone
- Department of Electrical, Electronics and Information Engineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata, 940-2188, Japan
| | - Kiyoshi Ohnuma
- Department of Science of Technology Innovation, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata, 940-2188, Japan.
- Department of Bioengineering, Nagaoka University of Technology, 1603-1 Kamitomioka, Nagaoka, Niigata, 940-2188, Japan.
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12
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Cao Y, Wu C, Ma L. Lysine demethylase 5B (KDM5B): A key regulator of cancer drug resistance. J Biochem Mol Toxicol 2024; 38:e23587. [PMID: 38014925 DOI: 10.1002/jbt.23587] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2023] [Revised: 10/17/2023] [Accepted: 11/10/2023] [Indexed: 11/29/2023]
Abstract
Chemoresistance, a roadblock in the chemotherapy process, has been impeding its effective treatment. KDM5B, a member of the histone demethylase family, has been crucial in the emergence and growth of malignancies. More significantly, KDM5B has recently been linked closely to cancer's resistance to chemotherapy. In this review, we explain the biological properties of KDM5B, its function in the emergence and evolution of cancer treatment resistance, and our hopes for future drug resistance-busting combinations involving KDM5B and related targets or medications.
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Affiliation(s)
- Yaquan Cao
- College of Basic Medicine and Forensic Medicine, Henan University of Science and Technology, Luoyang, China
| | - Chunli Wu
- School of Pharmaceutical Sciences, Zhengzhou University, Zhengzhou, China
| | - Liying Ma
- State Key Laboratory of Esophageal Cancer Prevention and Treatment, Key Laboratory of Advanced Pharmaceutical Technology, Ministry of Education of China, School of Pharmaceutical Science and Institute of Pharmaceutical Science, Zhengzhou University, Zhengzhou, Henan, China
- Key Laboratory of Cardio-Cerebrovascular Drug, China Meheco Topfond Pharmaceutical Company, Zhumadian, China
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13
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Dubau M, Tripetchr T, Mahmoud L, Kral V, Kleuser B. Advancing skin model development: A focus on a self-assembled, induced pluripotent stem cell-derived, xeno-free approach. J Tissue Eng 2024; 15:20417314241291848. [PMID: 39502328 PMCID: PMC11536386 DOI: 10.1177/20417314241291848] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2024] [Accepted: 10/01/2024] [Indexed: 11/08/2024] Open
Abstract
The demand for skin models as alternatives to animal testing has grown due to ethical concerns and the need for accurate substance evaluation. These alternatives, known as New Approach Methodologies (NAMs), are increasingly used for regulatory decisions. Current skin models from primary human cells often rely on bovine collagen, raising ethical issues. This study explores self-assembled skin models (SASM) as a new method, utilizing hair follicle-derived keratinocytes reprogrammed into induced pluripotent stem cells (iPSC) and differentiated into fibroblasts and keratinocytes. The model relies on the ability of fibroblasts to secrete collagen to produce a xeno-free dermal layer and on the differentiation of keratinocytes to create a functional epidermal layer. These layers exhibited confirmed metabolic activity and the capability to withstand test substances. The successful development of SASM underscores the significance of accurate alternatives in dermatological research, providing an ethical and reliable option for substance evaluation and regulatory testing.
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Affiliation(s)
| | | | - Lava Mahmoud
- Department of Pharmacology and Toxicology, Institute for Pharmacy, Freie Universität Berlin, Berlin, Germany
| | - Vivian Kral
- Department of Pharmacology and Toxicology, Institute for Pharmacy, Freie Universität Berlin, Berlin, Germany
| | - Burkhard Kleuser
- Department of Pharmacology and Toxicology, Institute for Pharmacy, Freie Universität Berlin, Berlin, Germany
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14
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He Q, Sze SK, Ng KS, Koh CG. Paxillin interactome identified by SILAC and label-free approaches coupled to TurboID sheds light on the compositions of focal adhesions in mouse embryonic stem cells. Biochem Biophys Res Commun 2023; 680:73-85. [PMID: 37725837 DOI: 10.1016/j.bbrc.2023.09.017] [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: 07/09/2023] [Revised: 08/25/2023] [Accepted: 09/08/2023] [Indexed: 09/21/2023]
Abstract
Self-renewal and differentiation of mouse embryonic stem cells (mESCs) are greatly affected by the extracellular matrix (ECM) environment; the composition and stiffness of which are sensed by the cells via integrin-associated focal adhesions (FAs) which link the cells to the ECM. Although FAs have been studied extensively in differentiated cells, their composition and function in mESCs are not as well elucidated. To gain more detailed knowledge of the molecular compositions of FAs in mESCs, we adopted the proximity-dependent biotinylation (BioID) proteomics approach. Paxillin, a known FA protein (FAP), is fused to the promiscuous biotin ligase TurboID as bait. We employed both SILAC- and label-free (LF)-based quantitative proteomics to strengthen as well as complement individual approach. The mass spectrometry data derived from SILAC and LF identified 38 and 443 proteins, respectively, with 35 overlapping candidates. Fifteen of these shared proteins are known FAPs based on literature-curated adhesome and 7 others are among the reported "meta-adhesome", suggesting the components of FAs are largely conserved between mESCs and differentiated cells. Furthermore, the LF data set contained an additional 18 literature-curated FAPs. Notably, the overlapped proteomics data failed to detect LIM-domain proteins such as zyxin family proteins, which suggests that FAs in mESCs are less mature than differentiated cells. Using the LF approach, we are able to identify PDLIM7, a LIM-domain protein, as a FAP in mESCs. This study illustrates the effectiveness of TurboID in mESCs. Importantly, we found that application of both SILAC and LF methods in combination allowed us to analyze the TurboID proteomics data in an unbiased, stringent and yet comprehensive manner.
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Affiliation(s)
- Qianqian He
- School of Biological Sciences, Nanyang Technological University, Singapore, 637551, Singapore
| | - Siu Kwan Sze
- School of Biological Sciences, Nanyang Technological University, Singapore, 637551, Singapore
| | - Kai Soon Ng
- School of Biological Sciences, Nanyang Technological University, Singapore, 637551, Singapore
| | - Cheng-Gee Koh
- School of Biological Sciences, Nanyang Technological University, Singapore, 637551, Singapore.
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15
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Xu Z, Liu S, Xue X, Li W, Fu J, Deng CX. Rapid responses of human pluripotent stem cells to cyclic mechanical strains applied to integrin by acoustic tweezing cytometry. Sci Rep 2023; 13:18030. [PMID: 37865697 PMCID: PMC10590420 DOI: 10.1038/s41598-023-45397-5] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2023] [Accepted: 10/19/2023] [Indexed: 10/23/2023] Open
Abstract
Acoustic tweezing cytometry (ATC) is an ultrasound-based biophysical technique that has shown the capability to promote differentiation of human pluripotent stem cells (hPSCs). This study systematically examined how hPSCs respond to cyclic mechanical strains applied by ATC via displacement of integrin-bound microbubbles (averaged diameter of 4.3 µm) using ultrasound pulses (acoustic pressure 0.034 MPa, center frequency 1.24 MHz and pulse repetition frequency 1 Hz). Our data show downregulation of pluripotency marker Octamer-binding transcription factor 4 (OCT4) by at least 10% and increased nuclear localization of Yes-associated protein (YAP) by almost 100% in hPSCs immediately after ATC application for as short as 1 min and 5 min respectively. Analysis of the movements of integrin-anchored microbubbles under ATC stimulations reveals different stages of viscoelastic characteristic behavior and increasing deformation of the integrin-cytoskeleton (CSK) linkage. The peak displacement of integrin-bound microbubbles increased from 1.45 ± 0.16 to 4.74 ± 0.67 μm as the duty cycle of ultrasound pulses increased from 5% to 50% or the duration of each ultrasound pulse increased from 0.05 to 0.5 s. Real-time tracking of integrin-bound microbubbles during ATC application detects high correlation of microbubble displacements with OCT4 downregulation in hPSCs. Together, our data showing fast downregulation of OCT4 in hPSCs in respond to ATC stimulations highlight the unique mechanosensitivity of hPSCs to integrin-targeted cyclic force/strain dependent on the pulse duration or duty cycle of ultrasound pulses, providing insights into the mechanism of ATC-induced accelerated differentiation of hPSCs.
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Affiliation(s)
- Zhaoyi Xu
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, MI, 48109, USA
| | - Shiying Liu
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, 48109, USA
| | - Xufeng Xue
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, MI, 48109, USA
| | - Weiping Li
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, 48109, USA
| | - Jianping Fu
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, MI, 48109, USA.
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, 48109, USA.
- Department of Cell and Developmental Biology, University of Michigan Medical School, Ann Arbor, MI, 48109, USA.
| | - Cheri X Deng
- Department of Mechanical Engineering, University of Michigan, Ann Arbor, MI, 48109, USA.
- Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, 48109, USA.
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16
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Ku T, Tan X, Liu Y, Wang R, Fan L, Ren Z, Ning X, Li G, Sang N. Triazole fungicides exert neural differentiation alteration through H3K27me3 modifications: In vitro and in silico study. JOURNAL OF HAZARDOUS MATERIALS 2023; 459:132225. [PMID: 37557044 DOI: 10.1016/j.jhazmat.2023.132225] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 06/10/2023] [Revised: 07/29/2023] [Accepted: 08/03/2023] [Indexed: 08/11/2023]
Abstract
Considering that humans are unavoidably exposed to triazole fungicides through the esophagus, respiratory tract, and skin contact, revealing the developmental toxicity of triazole fungicides is vital for health risk assessment. This study aimed to screen and discriminate neural developmental disorder chemicals in commonly used triazole fungicides, and explore the underlying harmful impacts on neurogenesis associated with histone modification abnormality in mouse embryonic stem cells (mESCs). The triploblastic and neural differentiation models were constructed based on mESCs to expose six typical triazole fungicides (myclobutanil, tebuconazole, hexaconazole, propiconazole, difenoconazole, and flusilazole). The result demonstrated that although no cytotoxicity was observed, different triazole fungicides exhibited varying degrees of alterations in neural differentiation, including increased ectodermal differentiation, promoted neurogenesis, increased intracellular calcium ion levels, and disturbance of neurotransmitters. Molecular docking, cluster analysis, and multiple linear regressions demonstrated that the binding affinities between triazole fungicides and the Kdm6b-ligand binding domain were the dominant determinants of the neurodevelopmental response. This partially resulted in the reduced enrichment of H3K27me3 at the promoter region of the serotonin receptor 2 C gene, finally leading to disturbed neural differentiation. The data suggested potential adverse outcomes of triazole fungicides on embryonic neurogenesis even under sublethal doses through interfering histone modification, providing substantial evidence on the safety control of fungicides.
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Affiliation(s)
- Tingting Ku
- College of Environment and Resource, Research Center of Environment and Health, Shanxi University, Taiyuan, Shanxi 030006, China
| | - Xin Tan
- College of Environment and Resource, Research Center of Environment and Health, Shanxi University, Taiyuan, Shanxi 030006, China
| | - Yutong Liu
- College of Environment and Resource, Research Center of Environment and Health, Shanxi University, Taiyuan, Shanxi 030006, China
| | - Rui Wang
- College of Environment and Resource, Research Center of Environment and Health, Shanxi University, Taiyuan, Shanxi 030006, China
| | - Lifan Fan
- College of Environment and Resource, Research Center of Environment and Health, Shanxi University, Taiyuan, Shanxi 030006, China
| | - Zhihua Ren
- College of Environment and Resource, Research Center of Environment and Health, Shanxi University, Taiyuan, Shanxi 030006, China
| | - Xia Ning
- College of Environment and Resource, Research Center of Environment and Health, Shanxi University, Taiyuan, Shanxi 030006, China
| | - Guangke Li
- College of Environment and Resource, Research Center of Environment and Health, Shanxi University, Taiyuan, Shanxi 030006, China
| | - Nan Sang
- College of Environment and Resource, Research Center of Environment and Health, Shanxi University, Taiyuan, Shanxi 030006, China.
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17
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Ghorbani S, Christine Füchtbauer A, Møllebjerg A, Møller Martensen P, Hvidbjerg Laursen S, Christian Evar Kraft D, Kjems J, Meyer RL, Rahimi K, Foss M, Füchtbauer EM, Sutherland DS. Protein ligand and nanotopography separately drive the phenotype of mouse embryonic stem cells. Biomaterials 2023; 301:122244. [PMID: 37459700 DOI: 10.1016/j.biomaterials.2023.122244] [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: 01/02/2023] [Revised: 07/09/2023] [Accepted: 07/12/2023] [Indexed: 09/06/2023]
Abstract
Biochemical and biomechanical signals regulate stem cell function in the niche environments in vivo. Current in vitro culture of mouse embryonic stem cells (mESC) uses laminin (LN-511) to provide mimetic biochemical signaling (LN-521 for human systems) to maintain stemness. Alternative approaches propose topographical cues to provide biomechanical cues, however combined biochemical and topographic cues may better mimic the in vivo environment, but are largely unexplored for in vitro stem cell expansion. In this study, we directly compare in vitro signals from LN-511 and/or topographic cues to maintain stemness, using systematically-varied submicron pillar patterns or flat surfaces with or without preadsorbed LN-511. The adhesion of cells, colony formation, expression of the pluripotency marker,octamer-binding transcription factor 4 (Oct4), and transcriptome profiling were characterized. We observed that either biochemical or topographic signals could maintain stemness of mESCs in feeder-free conditions, indicated by high-level Oct4 and gene profiling by RNAseq. The combination of LN-511 with nanotopography reduced colony growth, while maintaining stemness markers, shifted the cellular phenotype indicating that the integration of biochemical and topographic signals is antagonistic. Overall, significantly faster (up to 2.5 times) colony growth was observed at nanotopographies without LN-511, suggesting for improved ESC expansion.
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Affiliation(s)
- Sadegh Ghorbani
- Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, Aarhus C, 8000, Denmark; The Centre for Cellular Signal Patterns (CELLPAT), Gustav Wieds Vej 14, Aarhus C, 8000, Denmark; Department of Materials Science and Engineering, Stanford University, Stanford, CA, USA.
| | | | - Andreas Møllebjerg
- Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, Aarhus C, 8000, Denmark
| | | | - Sara Hvidbjerg Laursen
- Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, Aarhus C, 8000, Denmark
| | - David Christian Evar Kraft
- Department of Dentistry and Oral Health, Faculty of Health, University of Aarhus, Aarhus C, 8000, Denmark
| | - Jørgen Kjems
- Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, Aarhus C, 8000, Denmark; The Centre for Cellular Signal Patterns (CELLPAT), Gustav Wieds Vej 14, Aarhus C, 8000, Denmark; Department of Molecular Biology, University of Aarhus, Aarhus C, 8000, Denmark
| | - Rikke Louise Meyer
- Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, Aarhus C, 8000, Denmark
| | - Karim Rahimi
- Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, Aarhus C, 8000, Denmark; Department of Molecular Biology, University of Aarhus, Aarhus C, 8000, Denmark
| | - Morten Foss
- Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, Aarhus C, 8000, Denmark
| | | | - Duncan S Sutherland
- Interdisciplinary Nanoscience Center, Aarhus University, Gustav Wieds Vej 14, Aarhus C, 8000, Denmark; The Centre for Cellular Signal Patterns (CELLPAT), Gustav Wieds Vej 14, Aarhus C, 8000, Denmark.
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18
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Yi Y, Lan X, Li Y, Yan C, Lv J, Zhang T, Jiang W. Fatty acid synthesis and oxidation regulate human endoderm differentiation by mediating SMAD3 nuclear localization via acetylation. Dev Cell 2023; 58:1670-1687.e4. [PMID: 37516106 DOI: 10.1016/j.devcel.2023.07.005] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2022] [Revised: 05/02/2023] [Accepted: 07/07/2023] [Indexed: 07/31/2023]
Abstract
Metabolic remodeling is one of the earliest events that occur during cell differentiation. Here, we define fatty acid metabolism as a key player in definitive endoderm differentiation from human embryonic stem cells. Fatty acid β-oxidation is enhanced while lipogenesis is decreased, and this is due to the phosphorylation of lipogenic enzyme acetyl-CoA carboxylase by AMPK. More importantly, inhibition of fatty acid synthesis by either its inhibitors or AMPK agonist significantly promotes human endoderm differentiation, while blockade of fatty acid oxidation impairs differentiation. Mechanistically, reduced de novo fatty acid synthesis and enhanced fatty acid β-oxidation both contribute to the accumulation of intracellular acetyl-CoA, which guarantees the acetylation of SMAD3 and further causes nuclear localization to promote endoderm differentiation. Thus, our current study identifies a fatty acid synthesis/oxidation shift during early differentiation and presents an instructive role for fatty acid metabolism in regulating human endoderm differentiation.
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Affiliation(s)
- Ying Yi
- Department of Biological Repositories, Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan 430071, China
| | - Xianchun Lan
- Department of Biological Repositories, Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan 430071, China
| | - Yinglei Li
- Department of Biological Repositories, Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan 430071, China
| | - Chenchao Yan
- Department of Biological Repositories, Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan 430071, China
| | - Jing Lv
- Department of Biological Repositories, Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan 430071, China; College of Life Science, Cangzhou Normal University, Cangzhou 061000, China
| | - Tianzhe Zhang
- Department of Biological Repositories, Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan 430071, China
| | - Wei Jiang
- Department of Biological Repositories, Frontier Science Center for Immunology and Metabolism, Medical Research Institute, Zhongnan Hospital of Wuhan University, Wuhan University, Wuhan 430071, China; Hubei Provincial Key Laboratory of Developmentally Originated Disease, Wuhan 430071, China.
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19
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Ahmed TA, Eldaly B, Eldosuky S, Elkhenany H, El-Derby AM, Elshazly MF, El-Badri N. The interplay of cells, polymers, and vascularization in three-dimensional lung models and their applications in COVID-19 research and therapy. Stem Cell Res Ther 2023; 14:114. [PMID: 37118810 PMCID: PMC10144893 DOI: 10.1186/s13287-023-03341-4] [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: 10/15/2022] [Accepted: 04/14/2023] [Indexed: 04/30/2023] Open
Abstract
Millions of people have been affected ever since the emergence of the corona virus disease of 2019 (COVID-19) outbreak, leading to an urgent need for antiviral drug and vaccine development. Current experimentation on traditional two-dimensional culture (2D) fails to accurately mimic the in vivo microenvironment for the disease, while in vivo animal model testing does not faithfully replicate human COVID-19 infection. Human-based three-dimensional (3D) cell culture models such as spheroids, organoids, and organ-on-a-chip present a promising solution to these challenges. In this report, we review the recent 3D in vitro lung models used in COVID-19 infection and drug screening studies and highlight the most common types of natural and synthetic polymers used to generate 3D lung models.
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Affiliation(s)
- Toka A Ahmed
- Center of Excellence for Stem Cells and Regenerative Medicine (CESC), Zewail City of Science and Technology, October Gardens, 6th of October City, Giza, 12582, Egypt
- Egypt Center for Research and Regenerative Medicine (ECRRM), Cairo, Egypt
| | - Bassant Eldaly
- Center of Excellence for Stem Cells and Regenerative Medicine (CESC), Zewail City of Science and Technology, October Gardens, 6th of October City, Giza, 12582, Egypt
| | - Shadwa Eldosuky
- Center of Excellence for Stem Cells and Regenerative Medicine (CESC), Zewail City of Science and Technology, October Gardens, 6th of October City, Giza, 12582, Egypt
| | - Hoda Elkhenany
- Department of Surgery, Faculty of Veterinary Medicine, Alexandria University, Alexandria, 22785, Egypt
| | - Azza M El-Derby
- Center of Excellence for Stem Cells and Regenerative Medicine (CESC), Zewail City of Science and Technology, October Gardens, 6th of October City, Giza, 12582, Egypt
| | - Muhamed F Elshazly
- Center of Excellence for Stem Cells and Regenerative Medicine (CESC), Zewail City of Science and Technology, October Gardens, 6th of October City, Giza, 12582, Egypt
| | - Nagwa El-Badri
- Center of Excellence for Stem Cells and Regenerative Medicine (CESC), Zewail City of Science and Technology, October Gardens, 6th of October City, Giza, 12582, Egypt.
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20
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Ruan Y, Wang J, Yu M, Wang F, Wang J, Xu Y, Liu L, Cheng Y, Yang R, Zhang C, Yang Y, Wang J, Wu W, Huang Y, Tian Y, Chen G, Zhang J, Jian R. A multi-omics integrative analysis based on CRISPR screens re-defines the pluripotency regulatory network in ESCs. Commun Biol 2023; 6:410. [PMID: 37059858 PMCID: PMC10104827 DOI: 10.1038/s42003-023-04700-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/04/2022] [Accepted: 03/13/2023] [Indexed: 04/16/2023] Open
Abstract
A comprehensive and precise definition of the pluripotency gene regulatory network (PGRN) is crucial for clarifying the regulatory mechanisms in embryonic stem cells (ESCs). Here, after a CRISPR/Cas9-based functional genomics screen and integrative analysis with other functional genomes, transcriptomes, proteomes and epigenome data, an expanded pluripotency-associated gene set is obtained, and a new PGRN with nine sub-classes is constructed. By integrating the DNA binding, epigenetic modification, chromatin conformation, and RNA expression profiles, the PGRN is resolved to six functionally independent transcriptional modules (CORE, MYC, PAF, PRC, PCGF and TBX). Spatiotemporal transcriptomics reveal activated CORE/MYC/PAF module activity and repressed PRC/PCGF/TBX module activity in both mouse ESCs (mESCs) and pluripotent cells of early embryos. Moreover, this module activity pattern is found to be shared by human ESCs (hESCs) and cancers. Thus, our results provide novel insights into elucidating the molecular basis of ESC pluripotency.
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Affiliation(s)
- Yan Ruan
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
| | - Jiaqi Wang
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
- Department of Pathophysiology, College of High Altitude Military Medicine, Army Medical University, Chongqing, 400038, China
| | - Meng Yu
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
- Department of Joint Surgery, The First Affiliated Hospital, Army Medical University, Chongqing, 400038, China
| | - Fengsheng Wang
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
- State Key Laboratory of NBC Protection for Civilian, Beijing, 102205, China
| | - Jiangjun Wang
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
- Department of Cell Biology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
| | - Yixiao Xu
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
| | - Lianlian Liu
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
| | - Yuda Cheng
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
| | - Ran Yang
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
- Department of Pathophysiology, College of High Altitude Military Medicine, Army Medical University, Chongqing, 400038, China
| | - Chen Zhang
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
| | - Yi Yang
- Experimental Center of Basic Medicine, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
| | - JiaLi Wang
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
| | - Wei Wu
- Thoracic Surgery Department, Southwest Hospital, The First Hospital Affiliated to Army Medical University, Chongqing, 400038, China
| | - Yi Huang
- Biomedical Analysis Center, Army Medical University, Chongqing, 400038, China
| | - Yanping Tian
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China
| | - Guangxing Chen
- Department of Joint Surgery, The First Affiliated Hospital, Army Medical University, Chongqing, 400038, China.
| | - Junlei Zhang
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China.
| | - Rui Jian
- Laboratory of Stem Cell & Developmental Biology, Department of Histology and Embryology, College of Basic Medical Sciences, Army Medical University, Chongqing, 400038, China.
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21
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Xie G, Lee JE, Senft AD, Park YK, Jang Y, Chakraborty S, Thompson JJ, McKernan K, Liu C, Macfarlan TS, Rocha PP, Peng W, Ge K. MLL3/MLL4 methyltransferase activities control early embryonic development and embryonic stem cell differentiation in a lineage-selective manner. Nat Genet 2023; 55:693-705. [PMID: 37012455 DOI: 10.1038/s41588-023-01356-4] [Citation(s) in RCA: 33] [Impact Index Per Article: 16.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2020] [Accepted: 02/27/2023] [Indexed: 04/05/2023]
Abstract
H3K4me1 methyltransferases MLL3 (KMT2C) and MLL4 (KMT2D) are critical for enhancer activation, cell differentiation and development. However, roles of MLL3/4 enzymatic activities and MLL3/4-mediated enhancer H3K4me1 in these processes remain unclear. Here we report that constitutive elimination of both MLL3 and MLL4 enzymatic activities prevents initiation of gastrulation and leads to early embryonic lethality in mice. However, selective elimination of MLL3/4 enzymatic activities in embryonic, but not extraembryonic, lineages leaves gastrulation largely intact. Consistent with this, embryonic stem cells (ESCs) lacking MLL3/4 enzymatic activities can differentiate toward the three embryonic germ layers but show aberrant differentiation to extraembryonic endoderm (ExEn) and trophectoderm. The failure in ExEn differentiation can be attributed to markedly reduced enhancer-binding of the lineage-determining transcription factor GATA6. Furthermore, we show that MLL3/4-catalyzed H3K4me1 is largely dispensable for enhancer activation during ESC differentiation. Together, our findings suggest a lineage-selective, but enhancer activation-independent, role of MLL3/4 methyltransferase activities in early embryonic development and ESC differentiation.
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Affiliation(s)
- Guojia Xie
- National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
| | - Ji-Eun Lee
- National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
| | - Anna D Senft
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, USA
| | - Young-Kwon Park
- National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
| | - Younghoon Jang
- National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
| | - Shreeta Chakraborty
- Unit on Genome Structure and Regulation, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, USA
| | - Joyce J Thompson
- Unit on Genome Structure and Regulation, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, USA
| | - Kaitlin McKernan
- National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA
| | - Chengyu Liu
- Transgenic Core, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD, USA
| | - Todd S Macfarlan
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, USA
| | - Pedro P Rocha
- Unit on Genome Structure and Regulation, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD, USA
- National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Weiqun Peng
- Departments of Physics and Anatomy and Cell Biology, The George Washington University, Washington, DC, USA
| | - Kai Ge
- National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA.
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22
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Kim MH, Kuroda M, Ke D, Thanuthanakhun N, Kino-Oka M. An in vitro culture platform for studying the effect of collective cell migration on spatial self-organization within induced pluripotent stem cell colonies. J Biol Eng 2023; 17:25. [PMID: 36998087 PMCID: PMC10064534 DOI: 10.1186/s13036-023-00341-z] [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: 11/11/2022] [Accepted: 03/13/2023] [Indexed: 04/01/2023] Open
Abstract
BACKGROUND Human induced pluripotent stem cells (hiPSCs) provide an in vitro system to identify the impact of cell behavior on the earliest stages of cell fate specification during human development. Here, we developed an hiPSC-based model to study the effect of collective cell migration in meso-endodermal lineage segregation and cell fate decisions through the control of space confinement using a detachable ring culture system. RESULTS The actomyosin organization of cells at the edge of undifferentiated colonies formed in a ring barrier differed from that of the cells in the center of the colony. In addition, even in the absence of exogenous supplements, ectoderm, mesoderm, endoderm, and extraembryonic cells differentiated following the induction of collective cell migration at the colony edge by removing the ring-barrier. However, when collective cell migration was inhibited by blocking E-cadherin function, this fate decision within an hiPSC colony was altered to an ectodermal fate. Furthermore, the induction of collective cell migration at the colony edge using an endodermal induction media enhanced endodermal differentiation efficiency in association with cadherin switching, which is involved in the epithelial-mesenchymal transition. CONCLUSIONS Our findings suggest that collective cell migration can be an effective way to drive the segregation of mesoderm and endoderm lineages, and cell fate decisions of hiPSCs.
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Affiliation(s)
- Mee-Hae Kim
- Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan.
| | - Masaki Kuroda
- Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan
| | - Ding Ke
- Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan
| | - Naruchit Thanuthanakhun
- Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan
| | - Masahiro Kino-Oka
- Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan
- Research Base for Cell Manufacturability, Osaka University, 2-1 Yamadaoka, Suita, Osaka, 565-0871, Japan
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23
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Investigation of SAMD1 ablation in mice. Sci Rep 2023; 13:3000. [PMID: 36810619 PMCID: PMC9944271 DOI: 10.1038/s41598-023-29779-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2022] [Accepted: 02/10/2023] [Indexed: 02/23/2023] Open
Abstract
SAM domain-containing protein 1 (SAMD1) has been implicated in atherosclerosis, as well as in chromatin and transcriptional regulation, suggesting a versatile and complex biological function. However, its role at an organismal level is currently unknown. Here, we generated SAMD1-/- and SAMD1+/- mice to explore the role of SAMD1 during mouse embryogenesis. Homozygous loss of SAMD1 was embryonic lethal, with no living animals seen after embryonic day 18.5. At embryonic day 14.5, organs were degrading and/or incompletely developed, and no functional blood vessels were observed, suggesting failed blood vessel maturation. Sparse red blood cells were scattered and pooled, primarily near the embryo surface. Some embryos had malformed heads and brains at embryonic day 15.5. In vitro, SAMD1 absence impaired neuronal differentiation processes. Heterozygous SAMD1 knockout mice underwent normal embryogenesis and were born alive. Postnatal genotyping showed a reduced ability of these mice to thrive, possibly due to altered steroidogenesis. In summary, the characterization of SAMD1 knockout mice suggests a critical role of SAMD1 during developmental processes in multiple organs and tissues.
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24
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Huang L, Li F, Ye L, Yu F, Wang C. Epigenetic regulation of embryonic ectoderm development in stem cell differentiation and transformation during ontogenesis. Cell Prolif 2023; 56:e13413. [PMID: 36727213 PMCID: PMC10068960 DOI: 10.1111/cpr.13413] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/01/2022] [Revised: 01/09/2023] [Accepted: 01/18/2023] [Indexed: 02/03/2023] Open
Abstract
Dynamic chromatin accessibility regulates stem cell fate determination and tissue homeostasis via controlling gene expression. As a histone-modifying enzyme that predominantly mediates methylation of lysine 27 in histone H3 (H3K27me1/2/3), Polycomb repressive complex 2 (PRC2) plays the canonical role in targeting developmental regulators during stem cell differentiation and transformation. Embryonic ectoderm development (EED), the core scaffold subunit of PRC2 and as an H3K27me3-recognizing protein, has been broadly implicated with PRC2 stabilization and allosterically stimulated PRC2. Accumulating evidences from experimental data indicate that EED-associating epigenetic modifications are indispensable for stem cell maintenance and differentiation into specific cell lineages. In this review, we discuss the most updated advances to summarize the structural architecture of EED and its contributions and underlying mechanisms to mediating lineage differentiation of different stem cells during epigenetic modification to expand our understanding of PRC2.
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Affiliation(s)
- Liuyan Huang
- State Key Laboratory of Oral Diseases & National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China.,Department of Endodontics, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Feifei Li
- State Key Laboratory of Oral Diseases & National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Ling Ye
- State Key Laboratory of Oral Diseases & National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China.,Department of Endodontics, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Fanyuan Yu
- State Key Laboratory of Oral Diseases & National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China.,Department of Endodontics, West China Hospital of Stomatology, Sichuan University, Chengdu, China
| | - Chenglin Wang
- State Key Laboratory of Oral Diseases & National Clinical Research Center for Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China.,Department of Endodontics, West China Hospital of Stomatology, Sichuan University, Chengdu, China
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25
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Augustyniak J, Kozlowska H, Buzanska L. Genes Involved in DNA Repair and Mitophagy Protect Embryoid Bodies from the Toxic Effect of Methylmercury Chloride under Physioxia Conditions. Cells 2023; 12:cells12030390. [PMID: 36766732 PMCID: PMC9913246 DOI: 10.3390/cells12030390] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2022] [Revised: 01/17/2023] [Accepted: 01/19/2023] [Indexed: 01/24/2023] Open
Abstract
The formation of embryoid bodies (EBs) from human pluripotent stem cells resembles the early stages of human embryo development, mimicking the organization of three germ layers. In our study, EBs were tested for their vulnerability to chronic exposure to low doses of MeHgCl (1 nM) under atmospheric (21%O2) and physioxia (5%O2) conditions. Significant differences were observed in the relative expression of genes associated with DNA repair and mitophagy between the tested oxygen conditions in nontreated EBs. When compared to physioxia conditions, the significant differences recorded in EBs cultured at 21% O2 included: (1) lower expression of genes associated with DNA repair (ATM, OGG1, PARP1, POLG1) and mitophagy (PARK2); (2) higher level of mtDNA copy number; and (3) higher expression of the neuroectodermal gene (NES). Chronic exposure to a low dose of MeHgCl (1 nM) disrupted the development of EBs under both oxygen conditions. However, only EBs exposed to MeHgCl at 21% O2 revealed downregulation of mtDNA copy number, increased oxidative DNA damage and DNA fragmentation, as well as disturbances in SOX17 (endoderm) and TBXT (mesoderm) genes expression. Our data revealed that physioxia conditions protected EBs genome integrity and their further differentiation.
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Affiliation(s)
- Justyna Augustyniak
- Department of Neurochemistry, Mossakowski Medical Research Institute, Polish Academy of Sciences, 02-106 Warsaw, Poland
- Correspondence: (J.A.); (L.B.); Tel.: +48-668500988 (L.B.)
| | - Hanna Kozlowska
- Laboratory of Advanced Microscopy Technique, Mossakowski Medical Research Institute, Polish Academy of Sciences, 02-106 Warsaw, Poland
| | - Leonora Buzanska
- Department of Stem Cell Bioengineering, Mossakowski Medical Research Institute, Polish Academy of Sciences, 02-106 Warsaw, Poland
- Correspondence: (J.A.); (L.B.); Tel.: +48-668500988 (L.B.)
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26
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Jia Y, Guo Z, Zhu J, Qin G, Sun W, Yin Y, Wang H, Guo R. Snap29 Is Dispensable for Self-Renewal Maintenance but Required for Proper Differentiation of Mouse Embryonic Stem Cells. Int J Mol Sci 2023; 24:ijms24010750. [PMID: 36614195 PMCID: PMC9821219 DOI: 10.3390/ijms24010750] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2022] [Revised: 12/28/2022] [Accepted: 12/29/2022] [Indexed: 01/04/2023] Open
Abstract
Pluripotent embryonic stem cells (ESCs) can self-renew indefinitely and are able to differentiate into all three embryonic germ layers. Synaptosomal-associated protein 29 (Snap29) is implicated in numerous intracellular membrane trafficking pathways, including autophagy, which is involved in the maintenance of ESC pluripotency. However, the function of Snap29 in the self-renewal and differentiation of ESCs remains elusive. Here, we show that Snap29 depletion via CRISPR/Cas does not impair the self-renewal and expression of pluripotency-associated factors in mouse ESCs. However, Snap29 deficiency enhances the differentiation of ESCs into cardiomyocytes, as indicated by heart-like beating cells. Furthermore, transcriptome analysis reveals that Snap29 depletion significantly decreased the expression of numerous genes required for germ layer differentiation. Interestingly, Snap29 deficiency does not cause autophagy blockage in ESCs, which might be rescued by the SNAP family member Snap47. Our data show that Snap29 is dispensable for self-renewal maintenance, but required for the proper differentiation of mouse ESCs.
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Affiliation(s)
- Yumei Jia
- College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China
| | - Zhaoyuan Guo
- College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China
| | - Jiahao Zhu
- College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China
| | - Guanyu Qin
- College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China
| | - Wenwen Sun
- College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China
| | - Yu Yin
- Yunnan Key Laboratory of Primate Biomedical Research, Institute of Primate Translational Medicine, Kunming University of Science and Technology, Kunming 650500, China
| | - Haiying Wang
- State Key Laboratory of Medicinal Chemical Biology, Department of Cell Biology and Genetics, College of Life Sciences, Nankai University, Tianjin 300071, China
| | - Renpeng Guo
- College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China
- Correspondence:
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27
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Wang CM, Wu BR, Xiang P, Xiao J, Hu XC. Management of male erectile dysfunction: From the past to the future. Front Endocrinol (Lausanne) 2023; 14:1148834. [PMID: 36923224 PMCID: PMC10008940 DOI: 10.3389/fendo.2023.1148834] [Citation(s) in RCA: 18] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/20/2023] [Accepted: 02/13/2023] [Indexed: 03/02/2023] Open
Abstract
Erectile dysfunction is a common disease of the male reproductive system, which seriously affects the life quality of patients and their partners. At present, erectile dysfunction is considered as a social-psychological-physiological disease with complex etiology and various treatment methods. Oral PDE5I is the first-line treatment for erectile dysfunction with the advantages of high safety, good effect and non-invasiveness. But intracavernosal injection, hormonal replacement therapy, vacuum erection device, penile prosthesis implantation can also be alternative treatments for patients have organic erectile dysfunction or tolerance to PDE5I. With the rapid development of technologies, some new methods, such as low-intensity extracorporeal shock wave and stem cell injection therapy can even repair the organic damage of the corpora cavernosa. These are important directions for the treatment of male erectile dysfunction in the future. In this mini-review, we will introduce these therapies in detail.
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Affiliation(s)
| | | | | | - Jun Xiao
- *Correspondence: Jun Xiao, ; Xue-Chun Hu,
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28
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Quesenberry PJ, Wen S, Goldberg LR, Dooner MS. The universal stem cell. Leukemia 2022; 36:2784-2792. [PMID: 36307485 PMCID: PMC9712109 DOI: 10.1038/s41375-022-01715-w] [Citation(s) in RCA: 10] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2022] [Revised: 08/26/2022] [Accepted: 09/22/2022] [Indexed: 11/08/2022]
Abstract
Current dogma is that there exists a hematopoietic pluripotent stem cell, resident in the marrow, which is quiescent, but with tremendous proliferative and differentiative potential. Furthermore, the hematopoietic system is essentially hierarchical with progressive differentiation from the pluripotent stem cells to different classes of hematopoietic cells. However, results summarized here indicate that the marrow pluripotent hematopoietic stem cell is actively cycling and thus continually changing phenotype. As it progresses through cell cycle differentiation potential changes as illustrated by sequential changes in surface expression of B220 and GR-1 epitopes. Further data indicated that the potential of purified hematopoietic stem cells extends to multiple other non-hematopoietic cells. It appears that marrow stem cells will give rise to epithelial pulmonary cells at certain points in cell cycle. Thus, it appears that the marrow "hematopoietic" stem cell is also a stem cell for other non-hematopoietic tissues. These observations give rise to the concept of a universal stem cell. The marrow stem cell is not limited to hematopoiesis and its differentiation potential continually changes as it transits cell cycle. Thus, there is a universal stem cell in the marrow which alters its differentiation potential as it progresses through cell cycle. This potential is expressed when it resides in tissues compatible with its differentiation potential, at a particular point in cell cycle transit, or when it interacts with vesicles from that tissue.
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Affiliation(s)
- Peter J Quesenberry
- Division of Hematology/Oncology, Brown University, Rhode Island Hospital, Providence, RI, 02903, USA.
| | - Sicheng Wen
- Division of Hematology/Oncology, Brown University, Rhode Island Hospital, Providence, RI, 02903, USA
| | - Laura R Goldberg
- Division of Hematology/Oncology, Brown University, Rhode Island Hospital, Providence, RI, 02903, USA
- Division of Hematology, Brigham and Women's Hospital, Boston, MA, 02115, USA
| | - Mark S Dooner
- Division of Hematology/Oncology, Brown University, Rhode Island Hospital, Providence, RI, 02903, USA
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29
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Koh YE, Choi EH, Kim JW, Kim KP. The Kleisin Subunits of Cohesin are Involved in the Fate Determination of Embryonic Stem Cells. Mol Cells 2022; 45:820-832. [PMID: 36172976 PMCID: PMC9676991 DOI: 10.14348/molcells.2022.2042] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/09/2021] [Revised: 06/20/2022] [Accepted: 07/24/2022] [Indexed: 11/27/2022] Open
Abstract
As a potential candidate to generate an everlasting cell source to treat various diseases, embryonic stem cells are regarded as a promising therapeutic tool in the regenerative medicine field. Cohesin, a multi-functional complex that controls various cellular activities, plays roles not only in organizing chromosome dynamics but also in controlling transcriptional activities related to self-renewal and differentiation of stem cells. Here, we report a novel role of the α-kleisin subunits of cohesin (RAD21 and REC8) in the maintenance of the balance between these two stem-cell processes. By knocking down REC8, RAD21, or the non-kleisin cohesin subunit SMC3 in mouse embryonic stem cells, we show that reduction in cohesin level impairs their self-renewal. Interestingly, the transcriptomic analysis revealed that knocking down each cohesin subunit enables the differentiation of embryonic stem cells into specific lineages. Specifically, embryonic stem cells in which cohesin subunit RAD21 were knocked down differentiated into cells expressing neural alongside germline lineage markers. Thus, we conclude that cohesin appears to control the fate determination of embryonic stem cells.
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Affiliation(s)
- Young Eun Koh
- Department of Life Sciences, Chung-Ang University, Seoul 06974, Korea
- Genexine Inc., Bio Innovation Park, Seoul 07789, Korea
| | - Eui-Hwan Choi
- Department of Life Sciences, Chung-Ang University, Seoul 06974, Korea
- New Drug Development Center, Daegu-Gyeongbuk Medical Innovation Foundation, Daegu 41061, Korea
| | - Jung-Woong Kim
- Department of Life Sciences, Chung-Ang University, Seoul 06974, Korea
| | - Keun Pil Kim
- Department of Life Sciences, Chung-Ang University, Seoul 06974, Korea
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30
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Abstract
'Age reprogramming' refers to the process by which the molecular and cellular pathways of a cell that are subject to age-related decline are rejuvenated without passage through an embryonic stage. This process differs from the rejuvenation observed in differentiated derivatives of induced pluripotent stem cells, which involves passage through an embryonic stage and loss of cellular identity. Accordingly, the study of age reprogramming can provide an understanding of how ageing can be reversed while retaining cellular identity and the specialised function(s) of a cell, which will be of benefit to regenerative medicine. Here, we highlight recent work that has provided a more nuanced understanding of age reprogramming and point to some open questions in the field that might be explored in the future.
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Affiliation(s)
- Prim B. Singh
- Department of Medicine, Nazarbayev University School of Medicine, 5/1 Kerei Zhanibek Khandar Street, Astana 010000, Republic of Kazakhstan
| | - Assem Zhakupova
- Department of Medicine, Nazarbayev University School of Medicine, 5/1 Kerei Zhanibek Khandar Street, Astana 010000, Republic of Kazakhstan
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31
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Rajabi H, Mortazavi D, Konyalilar N, Aksoy GT, Erkan S, Korkunc SK, Kayalar O, Bayram H, Rahbarghazi R. Forthcoming complications in recovered COVID-19 patients with COPD and asthma; possible therapeutic opportunities. Cell Commun Signal 2022; 20:173. [PMID: 36320055 PMCID: PMC9623941 DOI: 10.1186/s12964-022-00982-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2022] [Accepted: 10/01/2022] [Indexed: 11/21/2022] Open
Abstract
Infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been growing swiftly worldwide. Patients with background chronic pulmonary inflammations such as asthma or chronic obstructive pulmonary diseases (COPD) are likely to be infected with this virus. Of note, there is an argument that COVID-19 can remain with serious complications like fibrosis or other pathological changes in the pulmonary tissue of patients with chronic diseases. Along with conventional medications, regenerative medicine, and cell-based therapy could be alternative approaches to compensate for organ loss or restore injured sites using different stem cell types. Owing to unique differentiation capacity and paracrine activity, these cells can accelerate the healing procedure. In this review article, we have tried to scrutinize different reports related to the harmful effects of SARS-CoV-2 on patients with asthma and COPD, as well as the possible therapeutic effects of stem cells in the alleviation of post-COVID-19 complications. Video abstract.
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Affiliation(s)
- Hadi Rajabi
- Koç University Research Centre for Translational Medicine (KUTTAM), Koç University School of Medicine, Istanbul, Turkey
| | - Deniz Mortazavi
- Koç University Research Centre for Translational Medicine (KUTTAM), Koç University School of Medicine, Istanbul, Turkey
| | - Nur Konyalilar
- Koç University Research Centre for Translational Medicine (KUTTAM), Koç University School of Medicine, Istanbul, Turkey
| | - Gizem Tuse Aksoy
- Koç University Research Centre for Translational Medicine (KUTTAM), Koç University School of Medicine, Istanbul, Turkey
| | - Sinem Erkan
- Koç University Research Centre for Translational Medicine (KUTTAM), Koç University School of Medicine, Istanbul, Turkey
| | - Seval Kubra Korkunc
- Koç University Research Centre for Translational Medicine (KUTTAM), Koç University School of Medicine, Istanbul, Turkey
| | - Ozgecan Kayalar
- Koç University Research Centre for Translational Medicine (KUTTAM), Koç University School of Medicine, Istanbul, Turkey
| | - Hasan Bayram
- Koç University Research Centre for Translational Medicine (KUTTAM), Koç University School of Medicine, Istanbul, Turkey.
- Department of Pulmonary Medicine, School of Medicine, Koç University, Istanbul, Turkey.
| | - Reza Rahbarghazi
- Stem Cell Research Centre, Tabriz University of Medical Sciences, Tabriz, Iran.
- Department of Applied Cell Sciences, Faculty of Advanced Medical Sciences, Tabriz University of Medical Sciences, Tabriz, Iran.
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32
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Liu D, Zinski A, Mishra A, Noh H, Park GH, Qin Y, Olorife O, Park JM, Abani CP, Park JS, Fung J, Sawaqed F, Coyle JT, Stahl E, Bendl J, Fullard JF, Roussos P, Zhang X, Stanton PK, Yin C, Huang W, Kim HY, Won H, Cho JH, Chung S. Impact of schizophrenia GWAS loci converge onto distinct pathways in cortical interneurons vs glutamatergic neurons during development. Mol Psychiatry 2022; 27:4218-4233. [PMID: 35701597 DOI: 10.1038/s41380-022-01654-z] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/26/2022] [Revised: 05/24/2022] [Accepted: 05/31/2022] [Indexed: 02/07/2023]
Abstract
Remarkable advances have been made in schizophrenia (SCZ) GWAS, but gleaning biological insight from these loci is challenging. Genetic influences on gene expression (e.g., eQTLs) are cell type-specific, but most studies that attempt to clarify GWAS loci's influence on gene expression have employed tissues with mixed cell compositions that can obscure cell-specific effects. Furthermore, enriched SCZ heritability in the fetal brain underscores the need to study the impact of SCZ risk loci in specific developing neurons. MGE-derived cortical interneurons (cINs) are consistently affected in SCZ brains and show enriched SCZ heritability in human fetal brains. We identified SCZ GWAS risk genes that are dysregulated in iPSC-derived homogeneous populations of developing SCZ cINs. These SCZ GWAS loci differential expression (DE) genes converge on the PKC pathway. Their disruption results in PKC hyperactivity in developing cINs, leading to arborization deficits. We show that the fine-mapped GWAS locus in the ATP2A2 gene of the PKC pathway harbors enhancer marks by ATACseq and ChIPseq, and regulates ATP2A2 expression. We also generated developing glutamatergic neurons (GNs), another population with enriched SCZ heritability, and confirmed their functionality after transplantation into the mouse brain. Then, we identified SCZ GWAS risk genes that are dysregulated in developing SCZ GNs. GN-specific SCZ GWAS loci DE genes converge on the ion transporter pathway, distinct from those for cINs. Disruption of the pathway gene CACNA1D resulted in deficits of Ca2+ currents in developing GNs, suggesting compromised neuronal function by GWAS loci pathway deficits during development. This study allows us to identify cell type-specific and developmental stage-specific mechanisms of SCZ risk gene function, and may aid in identifying mechanism-based novel therapeutic targets.
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Affiliation(s)
- Dongxin Liu
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA.
- Department of Developmental Cell Biology, Key Laboratory of Cell Biology, Ministry of Public Health, and Key Laboratory of Medical Cell Biology, Ministry of Education, China Medical University, Shenyang, China.
| | - Amy Zinski
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Akanksha Mishra
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Haneul Noh
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
- Department of Psychiatry, McLean Hospital/Harvard Medical School, Belmont, MA, 02478, USA
| | - Gun-Hoo Park
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Yiren Qin
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Oshoname Olorife
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - James M Park
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Chiderah P Abani
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Joy S Park
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Janice Fung
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Farah Sawaqed
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Joseph T Coyle
- Department of Psychiatry, McLean Hospital/Harvard Medical School, Belmont, MA, 02478, USA
| | - Eli Stahl
- Pamela Sklar Division of Psychiatric Genomics, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Department of Psychiatry, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
| | - Jaroslav Bendl
- Pamela Sklar Division of Psychiatric Genomics, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Department of Psychiatry, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Friedman Brain Institute, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Icahn Institute for Data Science and Genomic Technology, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
| | - John F Fullard
- Pamela Sklar Division of Psychiatric Genomics, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Department of Psychiatry, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Friedman Brain Institute, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Icahn Institute for Data Science and Genomic Technology, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
| | - Panos Roussos
- Pamela Sklar Division of Psychiatric Genomics, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Department of Psychiatry, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Friedman Brain Institute, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Icahn Institute for Data Science and Genomic Technology, Icahn School of Medicine at Mount Sinai, 1470 Madison Avenue, New York, NY, 10029, USA
- Mental Illness Research Education and Clinical Center (VISN 2 South), James J. Peters VA Medical Center, Bronx, NY, 10468, USA
| | - Xiaolei Zhang
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Patric K Stanton
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA
| | - Changhong Yin
- Department of Pathology, New York Medical College, Valhalla, NY, 10595, USA
| | - Weihua Huang
- Department of Pathology, New York Medical College, Valhalla, NY, 10595, USA
| | - Hae-Young Kim
- Department of Public Health, New York Medical College, Valhalla, NY, USA
| | - Hyejung Won
- Department of Genetics, University of North Carolina, Chapel Hill, NC, 27599, USA
| | - Jun-Hyeong Cho
- Department of Molecular, Cell and Systems Biology, University of California, Riverside, CA, 92521, USA
| | - Sangmi Chung
- Department of Cell biology and Anatomy, New York Medical College, Valhalla, NY, 10595, USA.
- Department of Psychiatry, McLean Hospital/Harvard Medical School, Belmont, MA, 02478, USA.
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Liu Y, Chen C, Wang X, Sun Y, Zhang J, Chen J, Shi Y. An Epigenetic Role of Mitochondria in Cancer. Cells 2022; 11:cells11162518. [PMID: 36010594 PMCID: PMC9406960 DOI: 10.3390/cells11162518] [Citation(s) in RCA: 65] [Impact Index Per Article: 21.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2022] [Revised: 08/03/2022] [Accepted: 08/09/2022] [Indexed: 12/14/2022] Open
Abstract
Mitochondria are not only the main energy supplier but are also the cell metabolic center regulating multiple key metaborates that play pivotal roles in epigenetics regulation. These metabolites include acetyl-CoA, α-ketoglutarate (α-KG), S-adenosyl methionine (SAM), NAD+, and O-linked beta-N-acetylglucosamine (O-GlcNAc), which are the main substrates for DNA methylation and histone post-translation modifications, essential for gene transcriptional regulation and cell fate determination. Tumorigenesis is attributed to many factors, including gene mutations and tumor microenvironment. Mitochondria and epigenetics play essential roles in tumor initiation, evolution, metastasis, and recurrence. Targeting mitochondrial metabolism and epigenetics are promising therapeutic strategies for tumor treatment. In this review, we summarize the roles of mitochondria in key metabolites required for epigenetics modification and in cell fate regulation and discuss the current strategy in cancer therapies via targeting epigenetic modifiers and related enzymes in metabolic regulation. This review is an important contribution to the understanding of the current metabolic-epigenetic-tumorigenesis concept.
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Affiliation(s)
- Yu’e Liu
- Tongji University Cancer Center, Shanghai Tenth People’s Hospital of Tongji University, School of Medicine, Tongji University, Shanghai 200092, China
| | - Chao Chen
- Department of Neurosurgery, Changhai Hospital, Second Military Medical University, 168 Changhai Road, Shanghai 200433, China
| | - Xinye Wang
- Tongji University Cancer Center, Shanghai Tenth People’s Hospital of Tongji University, School of Medicine, Tongji University, Shanghai 200092, China
| | - Yihong Sun
- Tongji University Cancer Center, Shanghai Tenth People’s Hospital of Tongji University, School of Medicine, Tongji University, Shanghai 200092, China
| | - Jin Zhang
- Department of Pharmacology and Toxicology, University of Mississippi Medical Center, Jackson, MS 39216, USA
| | - Juxiang Chen
- Department of Neurosurgery, Changhai Hospital, Second Military Medical University, 168 Changhai Road, Shanghai 200433, China
- Correspondence: (J.C.); (Y.S.)
| | - Yufeng Shi
- Tongji University Cancer Center, Shanghai Tenth People’s Hospital of Tongji University, School of Medicine, Tongji University, Shanghai 200092, China
- Clinical Center for Brain and Spinal Cord Research, Tongji University, Shanghai 200092, China
- Correspondence: (J.C.); (Y.S.)
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Johnson HJ, Chakraborty S, Muckom RJ, Balsara NP, Schaffer DV. A scalable and tunable thermoreversible polymer for 3D human pluripotent stem cell biomanufacturing. iScience 2022; 25:104971. [PMID: 36147944 PMCID: PMC9485071 DOI: 10.1016/j.isci.2022.104971] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2021] [Revised: 03/07/2022] [Accepted: 08/15/2022] [Indexed: 11/25/2022] Open
Abstract
Human pluripotent stem cells (hPSCs) are an exciting and promising source to enable cell replacement therapies for a variety of unmet medical needs. Though hPSCs can be successfully derived into numerous physiologically relevant cell types, effective translation to the clinic is limited by challenges in scalable production of high-quality cells, cellular immaturity following the differentiation process, and the use of animal-derived components in culture. To address these limitations, we have developed a fully defined, reproducible, and tunable thermoreversible polymer for high-quality, scalable 3D cell production. Our reproducible synthesis method enables precise control of gelation temperature (24°C–32°C), hydrogel stiffness (100–4000 Pa), and the prevention of any unintended covalent crosslinking. After material optimization, we demonstrated hPSC expansion, pluripotency maintenance, and differentiation into numerous lineages within the hydrogel. Overall, this 3D thermoreversible hydrogel platform has broad applications in scalable, high-quality cell production to overcome the biomanufacturing burden of stem cell therapy.
Synthesis of a scalable, tunable, and reproducible thermoreversible hydrogel Optimization of hydrogel properties including stiffness, LCST, and viscosity Expansion and pluripotency maintenance of hESCs in the hydrogel platform Differentiation of neurons, cardiomyocytes, and hepatocytes in the hydrogel platform
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Chen G, Yin S, Zeng H, Li H, Wan X. Regulation of Embryonic Stem Cell Self-Renewal. LIFE (BASEL, SWITZERLAND) 2022; 12:life12081151. [PMID: 36013330 PMCID: PMC9410528 DOI: 10.3390/life12081151] [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: 06/09/2022] [Revised: 07/12/2022] [Accepted: 07/21/2022] [Indexed: 11/16/2022]
Abstract
Embryonic stem cells (ESCs) are a type of cells capable of self-renewal and multi-directional differentiation. The self-renewal of ESCs is regulated by factors including signaling pathway proteins, transcription factors, epigenetic regulators, cytokines, and small molecular compounds. Similarly, non-coding RNAs, small RNAs, and microRNAs (miRNAs) also play an important role in the process. Functionally, the core transcription factors interact with helper transcription factors to activate the expression of genes that contribute to maintaining pluripotency, while suppressing the expression of differentiation-related genes. Additionally, cytokines such as leukemia suppressor factor (LIF) stimulate downstream signaling pathways and promote self-renewal of ESCs. Particularly, LIF binds to its receptor (LIFR/gp130) to trigger the downstream Jak-Stat3 signaling pathway. BMP4 activates the downstream pathway and acts in combination with Jak-Stat3 to promote pluripotency of ESCs in the absence of serum. In addition, activation of the Wnt-FDZ signaling pathway has been observed to facilitate the self-renewal of ESCs. Small molecule modulator proteins of the pathway mentioned above are widely used in in vitro culture of stem cells. Multiple epigenetic regulators are involved in the maintenance of ESCs self-renewal, making the epigenetic status of ESCs a crucial factor in this process. Similarly, non-coding RNAs and cellular energetics have been described to promote the maintenance of the ESC's self-renewal. These factors regulate the self-renewal and differentiation of ESCs by forming signaling networks. This review focused on the role of major transcription factors, signaling pathways, small molecular compounds, epigenetic regulators, non-coding RNAs, and cellular energetics in ESC's self-renewal.
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Affiliation(s)
- Guofang Chen
- Shanghai Key Laboratory of Maternal Fetal Medicine, Clinical and Translational Research Center of Shanghai First Maternity and Infant Hospital, Tongji University School of Medicine, Shanghai 201204, China;
- Correspondence: (G.C.); (H.L.); (X.W.); Tel./Fax: +86-021-20261000 (ext. 1379) (G.C.)
| | - Shasha Yin
- Shanghai Key Laboratory of Maternal Fetal Medicine, Clinical and Translational Research Center of Shanghai First Maternity and Infant Hospital, Tongji University School of Medicine, Shanghai 201204, China;
| | - Hongliang Zeng
- Institute of Chinese Materia Medica, Hunan Academy of Chinese Medicine, Changsha 410013, China;
| | - Haisen Li
- School of Medicine, Wayne State University, Detroit, MI 48201, USA
- Correspondence: (G.C.); (H.L.); (X.W.); Tel./Fax: +86-021-20261000 (ext. 1379) (G.C.)
| | - Xiaoping Wan
- Shanghai Key Laboratory of Maternal Fetal Medicine, Clinical and Translational Research Center of Shanghai First Maternity and Infant Hospital, Tongji University School of Medicine, Shanghai 201204, China;
- Correspondence: (G.C.); (H.L.); (X.W.); Tel./Fax: +86-021-20261000 (ext. 1379) (G.C.)
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Jafarzadeh E, Soodi M, Tiraihi T, Zarei M, Qasemian-Lemraski M. Study of lead-induced neurotoxicity in cholinergic cells differentiated from bone marrow-derived mesenchymal stem cells. Toxicol Ind Health 2022; 38:655-664. [PMID: 35838060 DOI: 10.1177/07482337221115514] [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] [Indexed: 11/15/2022]
Abstract
The developing brain is susceptible to the neurotoxic effects of lead. Exposure to lead has main effects on the cholinergic system and causes reduction of cholinergic neuron function during brain development. Disruption of the cholinergic system by chemicals, which play important roles during brain development, causes of neurodevelopmental toxicity. Differentiation of stem cells to neural cells is recently considered a promising tool for neurodevelopmental toxicity studies. This study evaluated the toxicity of lead acetate exposure during the differentiation of bone marrow-derived mesenchyme stem cells (bone marrow stem cells, BMSCs) to cholinergic neurons. Following institutional animal care review board approval, BMSCs were obtained from adult rats. The differentiating protocol included two stages that were pre-induction with β-mercaptoethanol (BME) for 24 h and differentiation to cholinergic neurons with nerve growth factor (NGF) over 5 days. The cells were exposed to different lead acetate concentrations (0.1-100 μm) during three stages, including undifferentiated, pre-induction, and neuronal differentiation stages; cell viability was measured by MTT assay. Lead exposure (0.01-100 μg/ml) had no cytotoxic effect on BMSCs but could significantly reduce cell viability at 50 and 100 μm concentrations during pre-induction and neuronal differentiation stages. MAP2 and choline acetyltransferase (ChAT) protein expression were investigated by immunocytochemistry. Although cells treated with 100 μm lead concentration expressed MAP2 protein in the differentiation stages, they had no neuronal cell morphology. The ChAT expression was negative in cells treated with lead. The present study showed that differentiated neuronal BMSCs are sensitive to lead toxicity during differentiation, and it is suggested that these cells be used to study neurodevelopmental toxicity.
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Affiliation(s)
- Emad Jafarzadeh
- Department of Toxicology, Faculty of Medical Sciences, 48503Tarbiat Modares University, Tehran, Iran
| | - Maliheh Soodi
- Department of Toxicology, Faculty of Medical Sciences, 48503Tarbiat Modares University, Tehran, Iran
| | - Taki Tiraihi
- Department of Anatomical Sciences, Faculty of Medical Sciences, 41616Tarbiat Modares University, Tehran, Iran
| | - Mohammadhadi Zarei
- Medical Plants Research Center, 154205Basic Health Sciences Institute, Shahrekord University of Medical Sciences, Shahrekord, Iran
| | - Mehdi Qasemian-Lemraski
- Department of Toxicology, Faculty of Medical Sciences, 48503Tarbiat Modares University, Tehran, Iran
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Wang J, Xiao B, Kimura E, Mongan M, Xia Y. The combined effects of Map3k1 mutation and dioxin on differentiation of keratinocytes derived from mouse embryonic stem cells. Sci Rep 2022; 12:11482. [PMID: 35798792 PMCID: PMC9263165 DOI: 10.1038/s41598-022-15760-z] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2022] [Accepted: 06/29/2022] [Indexed: 11/25/2022] Open
Abstract
Epithelial development starts with stem cell commitment to ectoderm followed by differentiation to the basal keratinocytes. The basal keratinocytes, first committed in embryogenesis, constitute the basal layer of the epidermis. They have robust proliferation and differentiation potential and are responsible for epidermal expansion, maintenance and regeneration. We generated basal epithelial cells in vitro through differentiation of mouse embryonic stem cells (mESCs). Early on in differentiation, the expression of stem cell markers, Oct4 and Nanog, decreased sharply along with increased ectoderm marker keratin (Krt) 18. Later on, Krt 18 expression was subdued when cells displayed basal keratinocyte characteristics, including regular polygonal shape, adherent and tight junctions and Krt 14 expression. These cells additionally expressed abundant Sca-1, Krt15 and p63, suggesting epidermal progenitor characteristics. Using Map3k1 mutant mESCs and environmental dioxin, we examined the gene and environment effects on differentiation. Neither Map3k1 mutation nor dioxin altered mESC differentiation to ectoderm and basal keratinocytes, but they, individually and in combination, potentiated Krt 1 expression and basal to spinous differentiation. Similar gene-environment effects were observed in vivo where dioxin exposure increased Krt 1 more substantially in the epithelium of Map3k1+/- than wild type embryos. Thus, the in vitro model of epithelial differentiation can be used to investigate the effects of genetic and environmental factors on epidermal development.
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Affiliation(s)
- Jingjing Wang
- Department of Environmental and Public Health Sciences, College of Medicine, University of Cincinnati, Cincinnati, OH, 45267-0056, USA
| | - Bo Xiao
- Department of Environmental and Public Health Sciences, College of Medicine, University of Cincinnati, Cincinnati, OH, 45267-0056, USA
| | - Eiki Kimura
- Department of Environmental and Public Health Sciences, College of Medicine, University of Cincinnati, Cincinnati, OH, 45267-0056, USA
| | - Maureen Mongan
- Department of Environmental and Public Health Sciences, College of Medicine, University of Cincinnati, Cincinnati, OH, 45267-0056, USA
| | - Ying Xia
- Department of Environmental and Public Health Sciences, College of Medicine, University of Cincinnati, Cincinnati, OH, 45267-0056, USA.
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Guo YL, Gurung C, Fendereski M, Huang F. Dicer and PKR as Novel Regulators of Embryonic Stem Cell Fate and Antiviral Innate Immunity. JOURNAL OF IMMUNOLOGY (BALTIMORE, MD. : 1950) 2022; 208:2259-2266. [PMID: 35577384 PMCID: PMC9179006 DOI: 10.4049/jimmunol.2200042] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/12/2022] [Accepted: 03/21/2022] [Indexed: 05/17/2023]
Abstract
Embryonic stem cells (ESCs) represent a unique cell population in the blastocyst stage embryo. They have been intensively studied as a promising cell source for regenerative medicine. Recent studies have revealed that both human and mouse ESCs are deficient in expressing IFNs and have attenuated inflammatory responses. Apparently, the ability to express IFNs and respond to certain inflammatory cytokines is not "innate" to ESCs but rather is developmentally acquired by somatic cells during differentiation. Accumulating evidence supports a hypothesis that the attenuated innate immune response may serve as a protective mechanism allowing ESCs to avoid immunological cytotoxicity. This review describes our current understanding of the molecular basis that shapes the immune properties of ESCs. We highlight the recent findings on Dicer and dsRNA-activated protein kinase R as novel regulators of ESC fate and antiviral immunity and discuss how ESCs use alternative mechanisms to accommodate their stem cell properties.
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Affiliation(s)
- Yan-Lin Guo
- Cell and Molecular Biology Program, University of Southern Mississippi, Hattiesburg, MS; and
| | - Chandan Gurung
- Cell and Molecular Biology Program, University of Southern Mississippi, Hattiesburg, MS; and
| | - Mona Fendereski
- Cell and Molecular Biology Program, University of Southern Mississippi, Hattiesburg, MS; and
| | - Faqing Huang
- Chemistry and Biochemistry Program, University of Southern Mississippi, Hattiesburg, MS
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Urade R, Chiu YH, Chiu CC, Wu CY. Small GTPases and Their Regulators: A Leading Road toward Blood Vessel Development in Zebrafish. Int J Mol Sci 2022; 23:4991. [PMID: 35563380 PMCID: PMC9099977 DOI: 10.3390/ijms23094991] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2022] [Revised: 04/27/2022] [Accepted: 04/27/2022] [Indexed: 12/26/2022] Open
Abstract
Members of the Ras superfamily have been found to perform several functions leading to the development of eukaryotes. These small GTPases are divided into five major subfamilies, and their regulators can "turn on" and "turn off" signals. Recent studies have shown that this superfamily of proteins has various roles in the process of vascular development, such as vasculogenesis and angiogenesis. Here, we discuss the role of these subfamilies in the development of the vascular system in zebrafish.
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Affiliation(s)
- Ritesh Urade
- Department of Biological Sciences, National Sun Yat-sen University, Kaohsiung 804, Taiwan; (R.U.); (Y.-H.C.)
| | - Yan-Hui Chiu
- Department of Biological Sciences, National Sun Yat-sen University, Kaohsiung 804, Taiwan; (R.U.); (Y.-H.C.)
| | - Chien-Chih Chiu
- Department of Biological Sciences, National Sun Yat-sen University, Kaohsiung 804, Taiwan; (R.U.); (Y.-H.C.)
- Department of Biotechnology, Kaohsiung Medical University, Kaohsiung 807, Taiwan
| | - Chang-Yi Wu
- Department of Biological Sciences, National Sun Yat-sen University, Kaohsiung 804, Taiwan; (R.U.); (Y.-H.C.)
- Department of Biotechnology, Kaohsiung Medical University, Kaohsiung 807, Taiwan
- Doctoral Degree Program in Marine Biotechnology, National Sun Yat-sen University, Kaohsiung 804, Taiwan
- Institute of Medical Science and Technology, National Sun Yat-sen University, Kaohsiung 804, Taiwan
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40
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Hidalgo Aguilar A, Smith L, Owens D, Quelch R, Przyborski S. Recreating Tissue Structures Representative of Teratomas In Vitro Using a Combination of 3D Cell Culture Technology and Human Embryonic Stem Cells. Bioengineering (Basel) 2022; 9:bioengineering9050185. [PMID: 35621463 PMCID: PMC9138123 DOI: 10.3390/bioengineering9050185] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2022] [Revised: 04/11/2022] [Accepted: 04/19/2022] [Indexed: 11/22/2022] Open
Abstract
In vitro studies using human embryonic stem cells (hESCs) are a valuable method to study aspects of embryogenesis, avoiding ethical issues when using embryonic materials and species dissimilarities. The xenograft teratoma assay is often traditionally used to establish pluripotency in putative PSC populations, but also has additional applications, including the study of tissue differentiation. The stem cell field has long sought an alternative due to various well-established issues with the in vivo technique, including significant protocol variability and animal usage. We have established a two-step culture method which combines PSC-derived embryoid bodies (EBs) with porous scaffolds to enhance their viability, prolonging the time these structures can be maintained, and therefore, permitting more complex, mature differentiation. Here, we have utilised human embryonic stem cell-derived EBs, demonstrating the formation of tissue rudiments of increasing complexity over time and the ability to manipulate their differentiation through the application of exogenous morphogens to achieve specific lineages. Crucially, these EB-derived tissues are highly reminiscent of xenograft teratoma samples derived from the same cell line. We believe this in vitro approach represents a reproducible, animal-free alternative to the teratoma assay, which can be used to study human tissue development.
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Affiliation(s)
| | - Lucy Smith
- Department of Biosciences, Durham University, Durham DH1 3LE, UK; (A.H.A.); (L.S.); (D.O.); (R.Q.)
| | - Dominic Owens
- Department of Biosciences, Durham University, Durham DH1 3LE, UK; (A.H.A.); (L.S.); (D.O.); (R.Q.)
| | - Rebecca Quelch
- Department of Biosciences, Durham University, Durham DH1 3LE, UK; (A.H.A.); (L.S.); (D.O.); (R.Q.)
| | - Stefan Przyborski
- Department of Biosciences, Durham University, Durham DH1 3LE, UK; (A.H.A.); (L.S.); (D.O.); (R.Q.)
- Reprocell Europe, NETPark, Sedgefield TS21 3FD, UK
- Correspondence:
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41
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Yan J, Huangfu D. Epigenome rewiring in human pluripotent stem cells. Trends Cell Biol 2022; 32:259-271. [PMID: 34955367 PMCID: PMC8840982 DOI: 10.1016/j.tcb.2021.12.001] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/10/2021] [Revised: 11/29/2021] [Accepted: 12/02/2021] [Indexed: 01/10/2023]
Abstract
The epigenome plays a crucial role in modulating the activity of regulatory elements, thereby orchestrating diverse transcriptional programs during embryonic development. Human (h)PSC stepwise differentiation provides an excellent platform for capturing dynamic epigenomic events during lineage transition in human development. Here we discuss how recent technological advances, from epigenomic mapping to targeted perturbation, are providing a more comprehensive appreciation of remodeling of the chromatin landscape during human development with implications for aberrant rewiring in disease. We predict that the continuous innovation of hPSC differentiation methods, epigenome mapping, and CRISPR (clustered regularly interspaced short palindromic repeats) perturbation technologies will allow researchers to build toward not only a comprehensive understanding of the epigenomic mechanisms governing development, but also a highly flexible way to model diseases with opportunities for translation.
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Affiliation(s)
- Jielin Yan
- Sloan Kettering Institute, 1275 York Avenue, New York, NY 10065, USA; Louis V. Gerstner Jr. Graduate School of Biomedical Sciences, Memorial Sloan Kettering Cancer Center, 1275 York Avenue, New York, NY 10065, USA
| | - Danwei Huangfu
- Sloan Kettering Institute, 1275 York Avenue, New York, NY 10065, USA.
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A simple and efficient cryopreservation method for mouse small intestinal and colon organoids for regenerative medicine. Biochem Biophys Res Commun 2022; 595:14-21. [DOI: 10.1016/j.bbrc.2021.12.021] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2021] [Revised: 12/07/2021] [Accepted: 12/07/2021] [Indexed: 11/18/2022]
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43
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Chen PH, Tjong WY, Yang HC, Liu HY, Stern A, Chiu DTY. Glucose-6-Phosphate Dehydrogenase, Redox Homeostasis and Embryogenesis. Int J Mol Sci 2022; 23:ijms23042017. [PMID: 35216131 PMCID: PMC8878822 DOI: 10.3390/ijms23042017] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/30/2021] [Revised: 02/04/2022] [Accepted: 02/08/2022] [Indexed: 12/04/2022] Open
Abstract
Normal embryogenesis requires complex regulation and precision, which depends on multiple mechanistic details. Defective embryogenesis can occur by various mechanisms. Maintaining redox homeostasis is of importance during embryogenesis. NADPH, as produced from the action of glucose-6-phosphate dehydrogenase (G6PD), has an important role in redox homeostasis, serving as a cofactor for glutathione reductase in the recycling of glutathione from oxidized glutathione and for NADPH oxidases and nitric oxide synthases in the generation of reactive oxygen (ROS) and nitrogen species (RNS). Oxidative stress differentially influences cell fate and embryogenesis. While low levels of stress (eustress) by ROS and RNS promote cell growth and differentiation, supra-physiological concentrations of ROS and RNS can lead to cell demise and embryonic lethality. G6PD-deficient cells and organisms have been used as models in embryogenesis for determining the role of redox signaling in regulating cell proliferation, differentiation and migration. Embryogenesis is also modulated by anti-oxidant enzymes, transcription factors, microRNAs, growth factors and signaling pathways, which are dependent on redox regulation. Crosstalk among transcription factors, microRNAs and redox signaling is essential for embryogenesis.
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Affiliation(s)
- Po-Hsiang Chen
- Graduate Institute of Health Industry Technology, College of Human Ecology, Chang Gung University of Science and Technology, Taoyuan 33303, Taiwan; (P.-H.C.); (W.-Y.T.); (D.T.-Y.C.)
- Research Center for Chinese Herbal Medicine, College of Human Ecology, Chang Gung University of Science and Technology, Taoyuan 33303, Taiwan
| | - Wen-Ye Tjong
- Graduate Institute of Health Industry Technology, College of Human Ecology, Chang Gung University of Science and Technology, Taoyuan 33303, Taiwan; (P.-H.C.); (W.-Y.T.); (D.T.-Y.C.)
- Research Center for Chinese Herbal Medicine, College of Human Ecology, Chang Gung University of Science and Technology, Taoyuan 33303, Taiwan
| | - Hung-Chi Yang
- Department of Medical Laboratory Science and Biotechnology, Yuanpei University of Medical Technology, Hsinchu 30015, Taiwan
- Correspondence: ; Tel.: +886-3-6108175; Fax: +886-3-6102327
| | - Hui-Ya Liu
- Department of Medical Biotechnology and Laboratory Sciences, College of Medicine, Chang Gung University, Taoyuan 33302, Taiwan;
| | - Arnold Stern
- Grossman School of Medicine, New York University, New York, NY 10016, USA;
| | - Daniel Tsun-Yee Chiu
- Graduate Institute of Health Industry Technology, College of Human Ecology, Chang Gung University of Science and Technology, Taoyuan 33303, Taiwan; (P.-H.C.); (W.-Y.T.); (D.T.-Y.C.)
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Lim J, Yoon J, Shin M, Lee KB, Choi JW. Biomolecular Electron Controller Composed of Nanobiohybrid with Electrically Released Complex for Spatiotemporal Control of Neuronal Differentiation. SMALL METHODS 2022; 6:e2100912. [PMID: 35174997 DOI: 10.1002/smtd.202100912] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/10/2021] [Revised: 11/11/2021] [Indexed: 06/14/2023]
Abstract
In vitro spatiotemporal control of cell differentiation is a critical issue in several biomedical fields such as stem cell therapy and regenerative medicine, as it enables the generation of heterogeneous tissue structures similar to those of their native counterparts. However, the simultaneous control of both spatial and temporal cell differentiation poses important challenges, and therefore no previous studies have achieved this goal. Here, the authors develop a cell differentiation biomolecular electron controller ("Biomoletron") composed of recombinant proteins, DNA, Au nanoparticles, peptides, and an electrically released complex with retinoic acid (RA) to spatiotemporally control SH-SY5Y cell differentiation. RA is only released from the Biomoletron when the complex is electrically stimulated, thus demonstrating the temporal control of SH-SY5Y cell differentiation. Furthermore, by introducing a patterned Au substrate that allows controlling the area where the Biomoletron is immobilized, spatiotemporal differentiation of the SH-SY5Y cell is successfully achieved. Therefore, the proposed Biomoletron-mediated differentiation method provides a promising strategy for spatiotemporal cell differentiation control with applications in regenerative medicine and cell therapy.
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Affiliation(s)
- Joungpyo Lim
- Department of Chemical & Biomolecular Engineering, Sogang University, Mapo-gu, Seoul, 04107, Republic of Korea
| | - Jinho Yoon
- Department of Chemical & Biomolecular Engineering, Sogang University, Mapo-gu, Seoul, 04107, Republic of Korea
- Department of Chemistry and Chemical Biology, Rutgers, The State University of New Jersey, Piscataway, NJ, 08854, USA
| | - Minkyu Shin
- Department of Chemical & Biomolecular Engineering, Sogang University, Mapo-gu, Seoul, 04107, Republic of Korea
| | - Ki-Bum Lee
- Department of Chemistry and Chemical Biology, Rutgers, The State University of New Jersey, Piscataway, NJ, 08854, USA
| | - Jeong-Woo Choi
- Department of Chemical & Biomolecular Engineering, Sogang University, Mapo-gu, Seoul, 04107, Republic of Korea
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Kim HJ, Kim G, Lee J, Lee Y, Kim JH. Secretome of Stem Cells: Roles of Extracellular Vesicles in Diseases, Stemness, Differentiation, and Reprogramming. Tissue Eng Regen Med 2022; 19:19-33. [PMID: 34817808 PMCID: PMC8782975 DOI: 10.1007/s13770-021-00406-4] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/05/2021] [Revised: 10/05/2021] [Accepted: 10/13/2021] [Indexed: 12/16/2022] Open
Abstract
Increasing evidence suggests that stem cells or stem cell-derived cells may contribute to tissue repair, not only by replacing lost tissue but also by delivering complex sets of secretory molecules, called secretomes, into host injured tissues. In recent years, extracellular vesicles (EVs) have gained much attention for their diverse and important roles in a wide range of pathophysiological processes. EVs are released from most types of cells and mediates cell-cell communication by activating receptors on target cells or by being taken up by recipient cells. EVs, including microvesicles and exosomes, encapsulate and carry proteins, nucleic acids, and lipids in the lumen and on the cell surface. Thus, EV-mediated intercellular communication has been extensively studied across various biological processes. While a number of investigations has been conducted in different tissues and body fluids, the field lacks a systematic review on stem cell-derived EVs, especially regarding their roles in stemness and differentiation. Here, we provide an overview of the pathophysiological roles of EVs and summarize recent findings focusing on EVs released from various types of stem cells. We also highlight emerging evidence for the potential implication of EVs in self-renewal, differentiation, and reprograming and discuss the benefits and limitations in translational approaches.
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Affiliation(s)
- Hyo Jin Kim
- Laboratory of Stem Cells and Tissue Regeneration, Department of Biotechnology, College of Life Sciences and Biotechnology, Korea University, 145, Anam-ro, Seongbuk-gu, West building of Life Sciences, Seoul, 02841, South Korea
| | - Gyeongmin Kim
- Laboratory of Stem Cells and Tissue Regeneration, Department of Biotechnology, College of Life Sciences and Biotechnology, Korea University, 145, Anam-ro, Seongbuk-gu, West building of Life Sciences, Seoul, 02841, South Korea
| | - Jihun Lee
- Laboratory of Stem Cells and Tissue Regeneration, Department of Biotechnology, College of Life Sciences and Biotechnology, Korea University, 145, Anam-ro, Seongbuk-gu, West building of Life Sciences, Seoul, 02841, South Korea
| | - Youngseok Lee
- Laboratory of Stem Cells and Tissue Regeneration, Department of Biotechnology, College of Life Sciences and Biotechnology, Korea University, 145, Anam-ro, Seongbuk-gu, West building of Life Sciences, Seoul, 02841, South Korea
| | - Jong-Hoon Kim
- Laboratory of Stem Cells and Tissue Regeneration, Department of Biotechnology, College of Life Sciences and Biotechnology, Korea University, 145, Anam-ro, Seongbuk-gu, West building of Life Sciences, Seoul, 02841, South Korea.
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Jo HY, Seo HH, Gil D, Park Y, Han HJ, Han HW, Thimmulappa RK, Kim SC, Kim JH. Single-Cell RNA Sequencing of Human Pluripotent Stem Cell-Derived Macrophages for Quality Control of The Cell Therapy Product. Front Genet 2022; 12:658862. [PMID: 35173760 PMCID: PMC8841343 DOI: 10.3389/fgene.2021.658862] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2021] [Accepted: 12/07/2021] [Indexed: 01/28/2023] Open
Abstract
Macrophages exhibit high plasticity to achieve their roles in maintaining tissue homeostasis, innate immunity, tissue repair and regeneration. Therefore, macrophages are being evaluated for cell-based therapeutics against inflammatory disorders and cancer. To overcome the limitation related to expansion of primary macrophages and cell numbers, human pluripotent stem cell (hPSC)-derived macrophages are considered as an alternative source of primary macrophages for clinical application. However, the quality of hPSC-derived macrophages with respect to the biological homogeneity remains still unclear. We previously reported a technique to produce hPSC-derived macrophages referred to as iMACs, which is amenable for scale-up. In this study, we have evaluated the biological homogeneity of the iMACs using a transcriptome dataset of 6,230 iMACs obtained by single-cell RNA sequencing. The dataset provides a valuable genomic profile for understanding the molecular characteristics of hPSC-derived macrophage cells and provide a measurement of transcriptomic homogeneity. Our study highlights the usefulness of single cell RNA-seq data in quality control of the cell-based therapy products.
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Affiliation(s)
- Hye-Yeong Jo
- Division of Intractable Diseases Research, Department of Chronic Diseases Convergence Research, Korea National Institute of Health, Cheongju, South Korea
- Korea National Stem Cell Bank, Cheongju, South Korea
- Division of Healthcare and AI, Center for Precision Medicine, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, South Korea
| | - Hyang-Hee Seo
- Division of Intractable Diseases Research, Department of Chronic Diseases Convergence Research, Korea National Institute of Health, Cheongju, South Korea
- Korea National Stem Cell Bank, Cheongju, South Korea
| | - Dayeon Gil
- Division of Intractable Diseases Research, Department of Chronic Diseases Convergence Research, Korea National Institute of Health, Cheongju, South Korea
- Korea National Stem Cell Bank, Cheongju, South Korea
| | | | - Hyeong-Jun Han
- Division of Intractable Diseases Research, Department of Chronic Diseases Convergence Research, Korea National Institute of Health, Cheongju, South Korea
- Korea National Stem Cell Bank, Cheongju, South Korea
| | - Hyo-Won Han
- Division of Intractable Diseases Research, Department of Chronic Diseases Convergence Research, Korea National Institute of Health, Cheongju, South Korea
- Korea National Stem Cell Bank, Cheongju, South Korea
| | - Rajesh K. Thimmulappa
- Department of Biochemistry, Center of Excellence in Molecular Biology and Regenerative Medicine, JSS Medical College, JSS Academy of Higher Education & Research, Mysuru, India
| | - Sang Cheol Kim
- Division of Healthcare and AI, Center for Precision Medicine, Korea National Institute of Health, Korea Centers for Disease Control and Prevention, Cheongju, South Korea
- *Correspondence: Jung-Hyun Kim, ; Sang Cheol Kim,
| | - Jung-Hyun Kim
- Division of Intractable Diseases Research, Department of Chronic Diseases Convergence Research, Korea National Institute of Health, Cheongju, South Korea
- Korea National Stem Cell Bank, Cheongju, South Korea
- *Correspondence: Jung-Hyun Kim, ; Sang Cheol Kim,
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Qiao L, Dho SH, Kim JY, Kim LK. SEPHS1 is dispensable for pluripotency maintenance but indispensable for cardiac differentiation in mouse embryonic stem cells. Biochem Biophys Res Commun 2022; 590:125-131. [PMID: 34974300 DOI: 10.1016/j.bbrc.2021.12.091] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2021] [Accepted: 12/23/2021] [Indexed: 11/28/2022]
Abstract
Embryonic stem cells (ESCs) are derived from the inner cell mass of developing blastocysts, which have self-renewal ability and have the potential to develop or reconstitute into all embryonic lineages. Selenophosphate synthetase 1 (SEPHS1) is an essential protein in mouse early embryo development. However, the role of SEPHS1 in mouse ESCs remains to be elucidated. In this study, we generated Sephs1 KO ESCs and found that deficiency of SEPSH1 has little effect on pluripotency maintenance and proliferation. Notably, SEPHS1 deficiency impaired differentiation into three germ layers and gastruloid aggregation in vitro. RNA-seq analysis showed SEPHS1 is involved in cardiogenesis, verified by no beating signal in Sephs1 KO embryoid body at d10 and low expression of cardiac-related and contraction markers. Taken together, our results suggest that SPEHS1 is dispensable in ESC self-renewal, but indispensable in subsequent germ layer differentiation especially for functional cardiac lineage.
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Affiliation(s)
- Lu Qiao
- School of Biological Sciences, College of Natural Sciences, Seoul National University, Seoul, 08826, South Korea
| | - So Hee Dho
- Severance Biomedical Science Institute, Graduate School of Medical Science, Brain Korea 21 Project, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, 06230, South Korea
| | - Ji Young Kim
- Severance Biomedical Science Institute, Graduate School of Medical Science, Brain Korea 21 Project, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, 06230, South Korea
| | - Lark Kyun Kim
- Severance Biomedical Science Institute, Graduate School of Medical Science, Brain Korea 21 Project, Gangnam Severance Hospital, Yonsei University College of Medicine, Seoul, 06230, South Korea.
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Thambyrajah R, Bigas A. Notch Signaling in HSC Emergence: When, Why and How. Cells 2022; 11:cells11030358. [PMID: 35159166 PMCID: PMC8833884 DOI: 10.3390/cells11030358] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2021] [Revised: 01/17/2022] [Accepted: 01/19/2022] [Indexed: 02/07/2023] Open
Abstract
The hematopoietic stem cell (HSC) sustains blood homeostasis throughout life in vertebrates. During embryonic development, HSCs emerge from the aorta-gonads and mesonephros (AGM) region along with hematopoietic progenitors within hematopoietic clusters which are found in the dorsal aorta, the main arterial vessel. Notch signaling, which is essential for arterial specification of the aorta, is also crucial in hematopoietic development and HSC activity. In this review, we will present and discuss the evidence that we have for Notch activity in hematopoietic cell fate specification and the crosstalk with the endothelial and arterial lineage. The core hematopoietic program is conserved across vertebrates and here we review studies conducted using different models of vertebrate hematopoiesis, including zebrafish, mouse and in vitro differentiated Embryonic stem cells. To fulfill the goal of engineering HSCs in vitro, we need to understand the molecular processes that modulate Notch signaling during HSC emergence in a temporal and spatial context. Here, we review relevant contributions from different model systems that are required to specify precursors of HSC and HSC activity through Notch interactions at different stages of development.
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Affiliation(s)
- Roshana Thambyrajah
- Program in Cancer Research, Institut Hospital del Mar d’Investigacions Mèdiques, CIBERONC, 08003 Barcelona, Spain
- Correspondence: (R.T.); (A.B.); Tel.: +34-933160437 (R.T.); +34-933160440 (A.B.)
| | - Anna Bigas
- Program in Cancer Research, Institut Hospital del Mar d’Investigacions Mèdiques, CIBERONC, 08003 Barcelona, Spain
- Josep Carreras Leukemia Research Institute, 08003 Barcelona, Spain
- Correspondence: (R.T.); (A.B.); Tel.: +34-933160437 (R.T.); +34-933160440 (A.B.)
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Shukla AK, Gao G, Kim BS. Applications of 3D Bioprinting Technology in Induced Pluripotent Stem Cells-Based Tissue Engineering. MICROMACHINES 2022; 13:155. [PMID: 35208280 PMCID: PMC8876961 DOI: 10.3390/mi13020155] [Citation(s) in RCA: 14] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 12/24/2021] [Revised: 01/17/2022] [Accepted: 01/18/2022] [Indexed: 02/01/2023]
Abstract
Induced pluripotent stem cells (iPSCs) are essentially produced by the genetic reprogramming of adult cells. Moreover, iPSC technology prevents the genetic manipulation of embryos. Hence, with the ensured element of safety, they rarely cause ethical concerns when utilized in tissue engineering. Several cumulative outcomes have demonstrated the functional superiority and potency of iPSCs in advanced regenerative medicine. Recently, an emerging trend in 3D bioprinting technology has been a more comprehensive approach to iPSC-based tissue engineering. The principal aim of this review is to provide an understanding of the applications of 3D bioprinting in iPSC-based tissue engineering. This review discusses the generation of iPSCs based on their distinct purpose, divided into two categories: (1) undifferentiated iPSCs applied with 3D bioprinting; (2) differentiated iPSCs applied with 3D bioprinting. Their significant potential is analyzed. Lastly, various applications for engineering tissues and organs have been introduced and discussed in detail.
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Affiliation(s)
- Arvind Kumar Shukla
- School of Biomedical Convergence Engineering, Pusan National University, Yangsan 50612, Korea;
| | - Ge Gao
- Institute of Engineering Medicine, Beijing Institute of Technology, Beijing 100081, China
- Department of Medical Technology, Beijing Institute of Technology, Beijing 100081, China
| | - Byoung Soo Kim
- School of Biomedical Convergence Engineering, Pusan National University, Yangsan 50612, Korea;
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50
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Šafaříková E, Ehlich J, Stříteský S, Vala M, Weiter M, Pacherník J, Kubala L, Víteček J. Conductive Polymer PEDOT:PSS-Based Platform for Embryonic Stem-Cell Differentiation. Int J Mol Sci 2022; 23:ijms23031107. [PMID: 35163031 PMCID: PMC8835127 DOI: 10.3390/ijms23031107] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2021] [Revised: 01/13/2022] [Accepted: 01/17/2022] [Indexed: 01/12/2023] Open
Abstract
Organic semiconductors are constantly gaining interest in regenerative medicine. Their tunable physico-chemical properties, including electrical conductivity, are very promising for the control of stem-cell differentiation. However, their use for combined material-based and electrical stimulation remains largely underexplored. Therefore, we carried out a study on whether a platform based on the conductive polymer poly(3,4-ethylenedioxythiophene):polystyrene sulfonate (PEDOT:PSS) can be beneficial to the differentiation of mouse embryonic stem cells (mESCs). The platform was prepared using the layout of a standard 24-well cell-culture plate. Polyethylene naphthalate foil served as the substrate for the preparation of interdigitated gold electrodes by physical vapor deposition. The PEDOT:PSS pattern was fabricated by precise screen printing over the gold electrodes. The PEDOT:PSS platform was able to produce higher electrical current with the pulsed-direct-current (DC) electrostimulation mode (1 Hz, 200 mV/mm, 100 ms pulse duration) compared to plain gold electrodes. There was a dominant capacitive component. In proof-of-concept experiments, mESCs were able to respond to such electrostimulation by membrane depolarization and elevation of cytosolic calcium. Further, the PEDOT:PSS platform was able to upregulate cardiomyogenesis and potentially inhibit early neurogenesis per se with minor contribution of electrostimulation. Hence, the present work highlights the large potential of PEDOT:PSS in regenerative medicine.
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Affiliation(s)
- Eva Šafaříková
- Institute of Biophysics of the Czech Academy of Sciences, Královopolská 135, 612 65 Brno, Czech Republic; (E.Š.); (L.K.)
- Department of Experimental Biology, Faculty of Science, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic;
| | - Jiří Ehlich
- Faculty of Chemistry, Brno University of Technology, Purkyňova 118, 612 00 Brno, Czech Republic; (J.E.); (S.S.); (M.V.); (M.W.)
| | - Stanislav Stříteský
- Faculty of Chemistry, Brno University of Technology, Purkyňova 118, 612 00 Brno, Czech Republic; (J.E.); (S.S.); (M.V.); (M.W.)
| | - Martin Vala
- Faculty of Chemistry, Brno University of Technology, Purkyňova 118, 612 00 Brno, Czech Republic; (J.E.); (S.S.); (M.V.); (M.W.)
| | - Martin Weiter
- Faculty of Chemistry, Brno University of Technology, Purkyňova 118, 612 00 Brno, Czech Republic; (J.E.); (S.S.); (M.V.); (M.W.)
| | - Jiří Pacherník
- Department of Experimental Biology, Faculty of Science, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic;
| | - Lukáš Kubala
- Institute of Biophysics of the Czech Academy of Sciences, Královopolská 135, 612 65 Brno, Czech Republic; (E.Š.); (L.K.)
- Department of Experimental Biology, Faculty of Science, Masaryk University, Kamenice 5, 625 00 Brno, Czech Republic;
- International Clinical Research Center, St. Anne’s University Hospital Brno, Pekařská 53, 656 91 Brno, Czech Republic
| | - Jan Víteček
- Institute of Biophysics of the Czech Academy of Sciences, Královopolská 135, 612 65 Brno, Czech Republic; (E.Š.); (L.K.)
- Correspondence: ; Tel./Fax: +420-541-517104; Fax: +420-541-517104
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