Academic Journal
Enhancing cardiomyocyte reprogramming efficiency by targeting cellular senescence is mediated via Rb1 gene.
| Title: | Enhancing cardiomyocyte reprogramming efficiency by targeting cellular senescence is mediated via Rb1 gene. |
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| Authors: | Fang J; Department of Cardiology, Guangdong Cardiovascular Institute, Guangdong Provincial People's Hospital (Guangdong Academy of Medical Sciences), Southern Medical University, Guangzhou, 510080, China.; Department of Cardiology, Experimental Cardiology Laboratory, University Medical Center Utrecht, Utrecht, The Netherlands.; Circulatory Health Laboratory, UMC Utrecht, Regenerative Medicine Center Utrecht, University Utrecht, Utrecht, 3508 GA, The Netherlands., Yang Q; Department of Cardiology, Experimental Cardiology Laboratory, University Medical Center Utrecht, Utrecht, The Netherlands.; CDL Research, University Medical Center Utrecht, Utrecht, The Netherlands., Maas RGC; Department of Cardiology, Experimental Cardiology Laboratory, University Medical Center Utrecht, Utrecht, The Netherlands.; Circulatory Health Laboratory, UMC Utrecht, Regenerative Medicine Center Utrecht, University Utrecht, Utrecht, 3508 GA, The Netherlands., Vader P; Department of Cardiology, Experimental Cardiology Laboratory, University Medical Center Utrecht, Utrecht, The Netherlands.; Circulatory Health Laboratory, UMC Utrecht, Regenerative Medicine Center Utrecht, University Utrecht, Utrecht, 3508 GA, The Netherlands.; CDL Research, University Medical Center Utrecht, Utrecht, The Netherlands., Mokry M; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Heidelberglaan 100, Utrecht, 3508 GA, The Netherlands., van den Dungen NAM; Central Diagnostic Laboratory, University Medical Center Utrecht, Utrecht University, Heidelberglaan 100, Utrecht, 3508 GA, The Netherlands., Qian L; McAllister Heart Institute, University of North Carolina, Chapel Hill, NC, USA., Xiao J; Cardiac Regeneration and Ageing Lab, Institute of Cardiovascular Sciences, School of Life Science, Shanghai University, Shanghai, 200444, China., Schiffelers R; CDL Research, University Medical Center Utrecht, Utrecht, The Netherlands., Lei Z; Department of Cardiology, Experimental Cardiology Laboratory, University Medical Center Utrecht, Utrecht, The Netherlands. Z.Lei-3@umcutrecht.nl.; Circulatory Health Laboratory, UMC Utrecht, Regenerative Medicine Center Utrecht, University Utrecht, Utrecht, 3508 GA, The Netherlands. Z.Lei-3@umcutrecht.nl.; CDL Research, University Medical Center Utrecht, Utrecht, The Netherlands. Z.Lei-3@umcutrecht.nl., Sluijter JPG; Department of Cardiology, Experimental Cardiology Laboratory, University Medical Center Utrecht, Utrecht, The Netherlands. J.Sluijter@umcutrecht.nl.; Circulatory Health Laboratory, UMC Utrecht, Regenerative Medicine Center Utrecht, University Utrecht, Utrecht, 3508 GA, The Netherlands. J.Sluijter@umcutrecht.nl. |
| Source: | Stem cell research & therapy [Stem Cell Res Ther] 2025 Dec 29; Vol. 16 (1), pp. 685. Date of Electronic Publication: 2025 Dec 29. |
| Publication Type: | Journal Article |
| Language: | English |
| Journal Info: | Publisher: BioMed Central Country of Publication: England NLM ID: 101527581 Publication Model: Electronic Cited Medium: Internet ISSN: 1757-6512 (Electronic) Linking ISSN: 17576512 NLM ISO Abbreviation: Stem Cell Res Ther Subsets: MEDLINE |
| Imprint Name(s): | Original Publication: London : BioMed Central |
| MeSH Terms: | Cellular Senescence*/genetics , Myocytes, Cardiac*/metabolism , Myocytes, Cardiac*/cytology , Cellular Reprogramming*/genetics , Ubiquitin-Protein Ligases*/genetics , Ubiquitin-Protein Ligases*/metabolism , Retinoblastoma Binding Proteins*/genetics , Retinoblastoma Binding Proteins*/metabolism, Fibroblasts/metabolism ; Fibroblasts/cytology ; GATA4 Transcription Factor/metabolism ; GATA4 Transcription Factor/genetics ; MEF2 Transcription Factors/metabolism ; MEF2 Transcription Factors/genetics ; Induced Pluripotent Stem Cells/metabolism ; Induced Pluripotent Stem Cells/cytology ; Animals ; Mice ; Humans ; Apoptosis ; Cells, Cultured |
| Abstract: | Introduction: Direct reprogramming of fibroblasts into cardiomyocytes by overexpressing cardiac transcription factors Gata4, Mef2c, and Tbx5 (GMT) is a promising way for cardiac repair, however, the low reprogramming efficiency remains a significant challenge. Cellular senescence, an irreversible cell-cycle arrest occurring in mitotic cells, has been reported to influence the efficiency of induced pluripotent stem cell (iPSC) reprogramming. Methods: We established an inducible GMT expression system in mouse embryonic fibroblasts (MEFs) and human fetal cardiac fibroblasts (hFCFs) using the PiggyBac transposon system. RNA sequencing was performed to identify genes associated with cellular senescence during reprogramming. Selected senescence-related genes were knocked down using shRNA, and their impact on reprogramming efficiency was assessed via flow cytometry, gene expression analysis, and staining for senescence and apoptosis markers. Results: Direct cardiac reprogramming induced cellular senescence and apoptosis, evidenced by enhanced β-Gal staining, elevated expression of senescence markers P16 and GLB1, and increased apoptosis rates. RNA sequencing and gene set enrichment analysis (GSEA) revealed significant upregulation of senescence-related genes (RB1, RBBP4, RBBP7, CBX8, and CDKN1B). Knockdown of these genes, particularly RB1, significantly enhanced reprogramming efficiency, increasing the proportion of GFP + cells in MEFs and α-actinin + cells in hFCFs. RB1 inhibition also reduced senescence marker levels and upregulated endogenous cardiac transcription factors GATA4 and MEF2C. Conclusions: Our findings demonstrate that cellular senescence might serves as a barrier to direct cardiac reprogramming and offer novel insights into the regulatory mechanisms involved in this process. (© 2025. The Author(s).) |
| Competing Interests: | Declarations. Ethics approval and consent to participate: Human fetal heart tissue was obtained by individual permission using standard written informed consent procedures and prior approval of the ethics committee of the Leiden University Medical Center, the Netherlands. This is in accordance with the principles outlined in the Declaration of Helsinki for the use of human tissue or subjects. All experiments were conducted according to the criteria of the code of proper use of human tissue used in the Netherlands. Consent for publication: Not applicable. Competing interests: The authors declare no competing interests. |
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| Grant Information: | 201906210082 Chinese Scholarship Council (CSC) fellowship program; 82400345 National Natural Science Foundation of China; 2025A04J4716 Basic and Applied Basic Research Foundation of Guangdong Province; 116002016 ZonMw-TAS program; 725229 Project EVICARE of the European Research Council (ERC) |
| Contributed Indexing: | Keywords: Direct cardiac reprogramming; RB1; Senescence |
| Substance Nomenclature: | 0 (GATA4 Transcription Factor) 0 (MEF2 Transcription Factors) EC 2.3.2.27 (Ubiquitin-Protein Ligases) 0 (Retinoblastoma Binding Proteins) 0 (RB1 protein, human) |
| Entry Date(s): | Date Created: 20251230 Date Completed: 20251230 Latest Revision: 20260101 |
| Update Code: | 20260130 |
| PubMed Central ID: | PMC12751188 |
| DOI: | 10.1186/s13287-025-04776-7 |
| PMID: | 41462332 |
| Database: | MEDLINE |
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