Abstract
MicroRNAs (miRNAs, miRs) have the potential to control stem cells fate decisions. The cardiac- and skeletal-muscle-specific miRNA, miR-1, can regulate embryonic stem cells differentiation to cardiac lineage by suppressing gene expression of alternative lineages. Accordingly, we hypothesized that overexpression of miR-1 may also promote cardiac gene expression in mesenchymal stem cells. Since Notch signaling could inhibit muscle differentiation, a process in contrast with the effect of miR-1, miR-1-mediated repression of Notch signaling may contribute to the observed effects of miR-1 in mesenchymal stem cells. Thus, mesenchymal stem cells were infected by lentiviral vectors carrying miR-1, and cells expressing miR-1 were selected. Alterations in Notch signaling and cardiomyocyte markers, Nkx2.5, GATA-4, cTnT, and CX43, were identified by Western blot in the infected cells on days 1, 7, and 14. Our study showed that the downstream target molecule of Notch pathway, Hes-1, was obviously decreased in mesenchymal stem cells modified with miR-1, and overexpression of miR-1 promotes the specific cardiac gene expression in the infected cells. Knockdown of Hes-1 leads to the same effects on cell lineage decisions. Our results indicated that miR-1 promotes the differentiation of MSCs into cardiac lineage in part due to negative regulation of Hes-1.
Highlights
Stem cell transplantation has been extensively investigated as a therapy to regenerate cardiac tissue a er myocardial infarction
To study whether downregulation of Hes-1 protein by miR-1 could account for a subset of the effects of miR-1 on cell lineage decisions, we used shRNA constructs directed against distinct regions of Hes-1 to generate Hes-1shRNA cell line
By using Western blot methods, we show that the expression of Hes-1 is signi cantly decreased in MSCsHes-1-shRNA on days 7 and 14
Summary
Stem cell transplantation has been extensively investigated as a therapy to regenerate cardiac tissue a er myocardial infarction. Mesenchymal stem cells (MSCs) can be isolated and expanded, and they possess neurogenic, chondrogenic, adipogenic, osteogenic, and myogenic properties under speci c differentiating conditions [1, 2]. These cells have a stable genetic background and low risk of immune rejection. As a result, they are o en used as seeding cells in tissue engineering and stem cell therapy. Numerous signaling pathways, including Wnt, BMP, and Notch signaling pathways, regulate cell fate decisions during MSC differentiation and can be utilized to in uence lineage choices in vitro [3,4,5]. The Notch signaling pathway is comprised of ve transmembrane ligands: deltalike- (Dll-) 1, (Dll-), (Dll-), Jagged (Jag-1), and Jagged (Jag2), four transmembrane receptors: Notch-1, -2, -3, and -4, and downstream target genes, such as bHLH (basic helixloop-helix) proteins: Hes (hairy/enhancer of split) and Hey (Hes-related protein). e pathway is crucial for cell-tocell interaction during cardiovascular development and may in uence cardiac differentiation, proliferation, and apoptotic events [7,8,9]
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