microRNAs (miRs) have emerged as critical modulators of various physiological processes including stem cell differentiation. Indeed, miR-1 has been reported to play an integral role in the regulation of cardiac muscle progenitor cell differentiation. However, whether overexpression of miR-1 in embryonic stem (ES) cells (miR-1-ES cells) will enhance cardiac myocyte differentiation following transplantation into the infarcted myocardium is unknown. In the present study, myocardial infarction (MI) was produced in C57BL/6 mice by left anterior descending artery ligation. miR-1-ES cells, ES cells, or culture medium (control) was transplanted into the border zone of the infarcted heart, and 2 wk post-MI, cardiac myocyte differentiation, adverse ventricular remodeling, and cardiac function were assessed. We provide evidence demonstrating enhanced cardiac myocyte commitment of transplanted miR-1-ES cells in the mouse infarcted heart as compared with ES cells. Assessment of apoptosis revealed that overexpression of miR-1 in transplanted ES cells protected host myocardium from MI-induced apoptosis through activation of p-AKT and inhibition of caspase-3, phosphatase and tensin homolog, and superoxide production. A significant reduction in interstitial and vascular fibrosis was quantified in miR-1-ES cell and ES cell transplanted groups compared with control MI. However, no statistical significance between miR-1-ES cell and ES cell groups was observed. Finally, mice receiving miR-1-ES cell transplantation post-MI had significantly improved heart function compared with respective controls (P < 0.05). Our data suggest miR-1 drives cardiac myocyte differentiation from transplanted ES cells and inhibits apoptosis post-MI, ultimately giving rise to enhanced cardiac repair, regeneration, and function.

Cardiac myocyte differentiation reported thus far is from iPS cells generated from mouse and human fibroblasts. However, there is no article on the generation of iPS cells from cardiac ventricular specific cell types such as H9c2 cells. Therefore, whether transduced H9c2 cells, originally isolated from embryonic cardiac ventricular tissue, will be able to generate iPS cells and have the potential to repair and regenerate infarcted myocardium remains completely elusive. We transduced H9c2 cells with four stemness factors, Oct3/4, Sox2, Klf4, and c-Myc, and successfully reprogrammed them into iPS cells. These iPS cells were able to differentiate into beating cardiac myocytes and positively stained for cardiac specific sarcomeric α-actin and myosin heavy chain proteins. Following transplantation in the infarcted myocardium, there were newly differentiated cardiac myocytes and formation of gap junction proteins at 2 weeks post-myocardial infarction (MI), suggesting newly formed cardiac myocytes were integrated into the native myocardium. Furthermore, transplanted iPS cells significantly (p < 0.05) inhibited apoptosis and fibrosis and improved cardiac function compared with MI and MI+H9c2 cell groups. Moreover, our iPS cell derived cardiac myocyte differentiation in vitro and in vivo was comparable to embryonic stem cells in the present study. In conclusion we report for the first time that we have H9c2 cell-derived iPS cells which contain the potential to differentiate into cardiac myocytes in the cell culture system and repair and regenerate infarcted myocardium with improved cardiac function in vivo.

The purpose of this study was to define the proteomic and phosphoproteomic landscape of circulating extracellular vesicles (EVs) in people with normal glucose tolerance (NGT), prediabetes (PDM), and diabetes (T2DM). Archived serum samples from 30 human subjects (n = 10 per group, ORIGINS study, NCT02226640) were used. EVs were isolated using EVtrap®. Mass spectrometry-based methods were used to detect the global EV proteome and phosphoproteome. Differentially expressed features, correlation, enriched pathways, and enriched tissue-specific protein sets were identified using custom R scripts. Phosphosite-centric analyses were conducted using directPA and PhosR software packages. A total of 2372 unique EV proteins and 716 unique EV phosphoproteins were identified among all samples. Unsupervised clustering of the differentially expressed (fold change ≥ 2, p < 0.05, FDR < 0.05) proteins and, particularly, phosphoproteins showed excellent discrimination among the three groups. CDK1 and PKCδ appear to drive key upstream phosphorylation events that define the phosphoproteomic signatures of PDM and T2DM. Circulating EVs from people with diabetes carry increased levels of specific phosphorylated kinases (i.e., AKT1, GSK3B, LYN, MAP2K2, MYLK, and PRKCD) and could potentially distribute activated kinases systemically. Among characteristic changes in the PDM and T2DM EVs, “integrin switching” appeared to be a central feature. Proteins involved in oxidative phosphorylation (OXPHOS), known to be reduced in various tissues in diabetes, were significantly increased in EVs from PDM and T2DM, which suggests that an abnormally elevated EV-mediated secretion of OXPHOS components may underlie the development of diabetes. A highly enriched signature of liver-specific markers among the downregulated EV proteins and phosphoproteins in both PDM and T2DM groups was also detected. This suggests that an alteration in liver EV composition and/or secretion may occur early in prediabetes. This study identified EV proteomic and phosphoproteomic signatures in people with prediabetes and T2DM and provides novel insight into the pathobiology of diabetes.

A maladaptive increase in anaplerosis via ME1 in TAC is associated with reduced GSH content. Suppressing increased ME1 expression in hypertrophied rat hearts, which is also elevated in failing human hearts, reduced pyruvate carboxylation thereby normalizing anaplerosis, restoring GSH content, and reducing lactate accumulation. Reducing ME1 induced favorable metabolic shifts for carbohydrate oxidation, improving intracellular redox state and enhanced cardiac performance in pathological hypertrophy.