Dataset Information

IGFBP2 Mediates Human iPSC-Cardiomyocyte Proliferation in a Cellular Contact-Dependent Manner

ABSTRACT: Inducing cardiomyocyte proliferation in situ presents a promising approach for cardiac regeneration following myocardial injury; however, mature cardiomyocytes exhibit inhibitory mechanisms that suppress proliferative signaling. This study investigates the role of cell-cell contact as a key regulator of cardiomyocyte proliferation. Using human iPSC-derived cardiomyocytes (hiPSC-CMs), the findings demonstrate that proliferation initially increases with cell density but is markedly suppressed upon the establishment of intercellular contacts. Phosphoproteomic and phenotypic analyses reveal that cell-cell contact promotes adherens junction formation, enhances sarcomere organization, and increases contractility, collectively contributing to the suppression of proliferation.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Pluripotent Stem Cell, Stem Cell, Cardiac Muscle Cell

SUBMITTER: Sean Humphrey

LAB HEAD: Sean Humphrey

PROVIDER: PXD061405 | Pride | 2025-12-15

REPOSITORIES: Pride

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	20231023_ASTR1_SeHu_EasyPhos_Hudson_1-1.raw	Raw
	20231023_ASTR1_SeHu_EasyPhos_Hudson_1-2.raw	Raw
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IGFBP2 Mediates Human iPSC-Cardiomyocyte Proliferation in a Cellular Contact-Dependent Manner.

Lee Soah S Heinrich Paul P Lee Daniel D Kang Yongwon Y Robinson Harley R HR Humphrey Sean J SJ Yun Jihye J Goodyer William R WR Buikema Jan W JW Paik David T DT Galdos Francisco X FX Kim Boyoung B Belbachir Nadjet N Min Sungjin S Cho Seung-Woo SW Lee Jaecheol J Moretti Alessandra A Wu Joseph C JC Hudson James E JE Wu Sean M SM

Circulation research 20251001 10

<h4>Background</h4>Induction of cardiomyocyte proliferation in situ represents a promising strategy for myocardial regeneration following injury. However, cardiomyocytes possess intrinsic inhibitory mechanisms that attenuate pro-proliferative signaling and constrain their expansion. We hypothesized that cell-cell contact is a key suppressor of cardiomyocyte proliferation. We aimed to delineate the underlying molecular pathways to enable sustained proliferation in 3-dimensional contexts.<h4>Metho ...[more]

PMID: 41031396

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Project description:Rationale: Inducing cardiomyocyte proliferation in situ is an attractive approach for achieving cardiac regeneration after myocardial injury. However, numerous inhibitory mechanisms are present in cardiomyocytes that silence the pro-proliferative signals for their expansion. We hypothesized that cell-cell contact exerts a major suppressive effect on cardiomyocyte proliferation. Objectives: This study aims to uncover the molecular mechanisms underlying cell contact-mediated inhibition of cardiomyocyte proliferation and leverage this knowledge to sustain proliferation of cardiomyocytes in 3D despite cell-cell contact. Methods and Results: Using human iPSC-derived cardiomyocytes (hiPSC-CMs) as a model system, we found that the proliferative capacity of hiPSC-CMs is initially increased proportional to cell density until cells form intercellular contacts. We found that cell-cell contact exerts a strong inhibitory effect on hiPSC-CM proliferation as cell density increases. Phosphoproteomics analysis and cellular phenotyping revealed that cell-cell contact accompanies adherens junction formation, enhanced sarcomere organization, and increased contractility. Disruption of adherens junctions or sarcomere assembly via siRNA-mediated knockdown of N-cadherin or ɑ-actinin, respectively, resulted in increased cell cycle activation of hiPSC-CMs. Furthermore, disruption of cell-cell contact enhanced nuclear translocation of β-catenin and TCF/LEF transcriptional activity that contributed to hiPSC-CM proliferation. However, this was not sufficient to drive division of hiPSC-CMs. Additional screening for putative secreted growth factors in the conditioned media from sparsely plated hiPSC-CMs revealed the enrichment of IGFBP2, which was sufficient to drive hiPSC-CM division in the presence of cell-cell contact in 3D constructs. Conclusions: Our findings demonstrate that cell-cell contact inhibits hiPSC-CM proliferation through adherens junction formation, sarcomeric assembly, and reduced IGFBP2 secretion. Importantly, exogenous supplementation of IGFBP2 can overcome cell contact-mediated inhibition of hiPSC-CM proliferation and facilitate the growth of 3D cardiac tissue. These insights provide valuable implications for advancing cardiac tissue engineering and regenerative therapies.

Project description:Background: There is a limited capacity to repair damage in the mammalian heart after birth, which is primarily due to the inability of cardiomyocytes to proliferate after birth. This is in contrast to zebrafish and salamander, in which cardiomyocytes retain the ability to proliferate throughout life and can regenerate their heart after significant damage. Recent studies in zebrafish and rodents implicate microRNAs (miRNAs) in the regulation of genes responsible for cardiac cell cycle progression and regeneration, in particular, miR-133a, the miR-15 family, miR-199a and miR-590. However, the significance of these miRNAs and miRNA in general in the regulation of cardiomyocyte proliferation in large mammals, including humans, where the timing of heart development relative to birth is very different than in rodents, is unclear. To determine the involvement of miRNAs in the down-regulation of cardiomyocyte proliferation occurring before birth in large mammals, we investigated miRNA and target gene expression in sheep hearts before and after birth. The experimental approach included targeted transcriptional profiling of miRNA and target mRNA previously identified in rodent studies as well as genome-wide miRNA profiling using microarrays. Results: The cardiac expression of miR-133a increased and its target gene IGF1R decreased with increasing age, reaching their respective maximum and minimum abundance when the majority of ovine cardiomyocytes were quiescent. The expression of the miR-15 family members was variable with age, however, four of their target genes decreased with age. These latter profiles are inconsistent with the direct involvement of this family of miRNA in cardiomyocyte quiescence in late gestation sheep. The expression patterns of ‘pro-proliferative’ miR-199a and miR-590 were also inconsistent with their involvement in cardiomyocyte quiescence. Consequently, miRNA microarray analysis was undertaken, which identified six discrete clusters of miRNA with characteristic developmental profiles. The functions of predicted target genes for the miRNA in four of the six clusters were enriched for aspects of cell division and regulation of cell proliferation suggesting a potential role of these miRNA in regulating cardiomyocyte proliferation. Conclusion: The results of this study show that the expression of miR-133a and one of its target genes is consistent with it being involved in the suppression of cardiomyocyte proliferation, which occurs across the last third of gestation in sheep. The expression patterns of the miR-15 family, miR-199a and miR-590 were inconsistent with direct involvement in the regulation cardiomyocyte proliferation in sheep, despite studies in rodents demonstrating that their manipulation can influence the degree of cardiomyocyte proliferation. miRNA microarray analysis suggests a coordinated and potentially more complex role of multiple miRNA in the regulation of cardiomyocyte quiescence and highlights significant differences between species that may reflect their substantial differences in the timing of this developmental process. RNA, (2 µg) from three heart samples from each of the 91 d (fetus), 141 d (fetus), 5 d (postnatal) and 173 d (postnatal) groups, was used for analysis according to standard procedures. The experimental design included three biological replicates from each of the four normal developmental ages in sheep and thus a total of 12 samples was analyzed.