Human Mesenchymal Stem Cells in Cardiomyocyte Co-Culture
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE32171 | GEO | 2011/09/16
SECONDARY ACCESSION(S): PRJNA147425
REPOSITORIES: GEO
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Project description:Evaluate the change in transcription factors that have a role in human mesenchymal stem cell (hMSC) commitment to a cardiomyocyte lineage when co-cultured for 4 days with rat neonatal cardiomyocytes and before acquiring a recognizable cardiac phenotype. A myocardial microenvironment was generated by dissociating neonatal rat hearts and establishing cardiomyocyte primary cultures. HumanMSCs constitutively labeled with dsRed localized to the cell's mitochondria were either grown separately (control) or added to the cardiomyocyte primary cultures and grown for 4 days. dsRed fluorescent hMSCs were harvested from co-cultures at 4 days using a FACscan flow cytometer. The RNA for the microarray analysis was prepared from three biologically separate samples of hMSCs co-cultured for 4 days and from hMSCs grown separately for 4 days (control).
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Project description:Heart muscle cells, cardiomyocytes, are highly differentiated cells that usually do not proliferate . During the non-proliferative state, extracellular signals control cardiomyocyte contractile function. However, during development and regeneration, cardiomyocytes enter the cell cycle and divide. It is unknown how cardiomyocytes modify their intracellular signaling to direct the cell cycle program. Here, we show that the nuclear lamina protein Lamin B2 (Lmnb2) regulates cardiomyocyte cell cycle activity using a gatekeeper mechanism. We identified Lmnb2 as a candidate for regulating intracellular signaling with deep transcriptional profiling of single cardiomyocytes. Lmnb2 was sufficient and necessary for cardiomyocyte cycling in the presence of serum. Lmnb2 increased the nucleoporin NUP98 and permeability of the nuclear membrane for phosphorylated ERK1/2. In vivo, the Lmnb2 gene was required for cardiomyocyte cell cycle activity during development. Increasing the expression of Lmnb2 in neonatal mice promoted cardiomyocyte M-phase and cytokinesis. LmnB2 gene transfer in neonatal mice that received a myocardial injury increased cardiomyocyte division and myocardial function in the injury border zone, indicating that the regenerated cardiomyocytes were functionally integrated. We propose a gatekeeper function of Lmnb2 that can be targeted to increase cardiomyocyte regeneration without the administration of exogenous growth factors.
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Project description:Cardiomyocyte poly(A) RNA was sequenced from purified bulk cardiomyocytes collected from one male and female murine heart at postnatal day 2 (P2). Neonatal cardiomyocytes were isolated and purified (96% cardiomyocytes at P2) by Langendorff retrograde perfusion and immunomagnetic cell separation, respectively. We found evidence of sexual dimorphism with 9 differentially expressed genes (FDR<0.05) encoded on XY chromosomes in this RNA-Seq dataset.
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Project description:Cardiac maturation lays the foundation for postnatal heart development and disease, yet little is known about the contributions of the microenvironment to cardiomyocyte maturation. By integrating single-cell RNA-sequencing data of mouse hearts at multiple postnatal stages, we construct cellular interactomes and regulatory signaling networks. Here we report switching of fibroblast subtypes from a neonatal to adult state and this drives cardiomyocyte maturation. Molecular and functional maturation of neonatal mouse cardiomyocytes and human embryonic stem cell-derived cardiomyocytes are considerably enhanced upon coculture with corresponding adult cardiac fibroblasts. Further, single-cell analysis of in vivo and in vitro cardiomyocyte maturation trajectories identify highly conserved signaling pathways, pharmacological targeting of which substantially delays cardiomyocyte maturation in postnatal hearts, and markedly enhances cardiomyocyte proliferation and improves cardiac function in infarcted hearts. Together, we identify cardiac fibroblasts as a key constituent in the microenvironment promoting cardiomyocyte maturation, providing insights into how the manipulation of cardiomyocyte maturity may impact on disease development and regeneration.
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Project description:Since the proliferative capacity of cardiomyocytes is extremely limited in the adult mammalian hearts, the irreversible loss of cardiomyocytes following cardiac injury markedly reduces cardiac function, leading to cardiac remodeling and heart failure. However, the early neonatal mice have a strong ability in cardiomyocyte proliferation and cardiac regeneration after heart damage such as apical resection. Besides of cardiomyocytes, non-myocytes in heart tissue also play important roles in the regeneration process. Previous studies showed that cardiac macrophages, regulatory T cells and CD4+ T cells are all involved in regulating the myocardial regeneration process. However, the roles of other cardiac immune cells in cardiac regeneration remains to be elucidated. B cells is a prominent immune cell in injured heart; here we discovered the indispensable function of cardiac B cells in improving cardiomyocyte proliferation and heart regeneration in neonatal mice.
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Project description:The acquisition of knowledge pertaining to the molecular mechanisms that govern cardiomyocyte (CM) proliferation would enable the creation of novel approaches to promote cardiac regeneration in adult individuals, a crucial therapeutic objective that has yet to be accomplished. The limited regenerative response observed in adult myocardium may be attributed, in part, to the relatively low proliferative capacity of cardiomyocytes. The comprehension of cardiomyocyte division is a complex undertaking, as cardiomyocytes are capable of entering the cell cycle but are unable to complete cell division. Our research revealed that the induction of cardiac cytoskeleton changes by Blebbistatin result in the entry of cardiomyocytes entry into the cell cycle but, followed by cell cycle arrest at the G2/M phase. Sirt1 is identified as a potential mediator of the binucleation process in neonatal rat cardiomyocytes, which is regulated by the cardiomyocyte cytoskeleton. Furthermore, the distribution of H3K9me3 in cardiomyocytes with varying nuclear numbers and treatments indicates a significant involvement in the formation of binucleated cardiomyocytes.
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Project description:We examine the role of G3bp1, a RNA binding protein and site specific endoribonuclease in gene expression in isolated neonatal cardiomyocytes. RNAseq data from cardiomyocytes were infected with adenoviruses expressing shRNA against G3bp1 (ad-siG3bp1) or Luciferase (ad-siLUC, control) showed significant decrease in transcript abudnnace of cardiac-enriched genes involved in Calcium handling, contraction, action potential and sacromere function. On the other hand increase was observed in genes that regulate Hippo, TNF and TGFb signaling. Knockdown of G3bp1 inhibited endothelin-1 induced cardiomyocyte hypertrophy.
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