MiR‐590 Promotes Transdifferentiation of Porcine and Human Fibroblasts Toward a Cardiomyocyte‐Like Fate by Directly Repressing Specificity Protein 1

Abstract

BackgroundReprogramming of cardiac fibroblasts into induced cardiomyocyte‐like cells represents a promising potential new therapy for treating heart disease, inducing significant improvements in postinfarct ventricular function in rodent models. Because reprogramming factors effective in transdifferentiating rodent cells are not sufficient to reprogram human cells, we sought to identify reprogramming factors potentially applicable to human studies.Methods and ResultsLentivirus vectors expressing Gata4, Mef2c, and Tbx5 (GMT); Hand2 (H), Myocardin (My), or microRNA (miR)‐590 were administered to rat, porcine, and human cardiac fibroblasts in vitro. induced cardiomyocyte‐like cell production was then evaluated by assessing expression of the cardiomyocyte marker, cardiac troponin T (cTnT), whereas signaling pathway studies were performed to identify reprogramming factor targets. GMT administration induced cTnT expression in ≈6% of rat fibroblasts, but failed to induce cTnT expression in porcine or human cardiac fibroblasts. Addition of H/My and/or miR‐590 to GMT administration resulted in cTNT expression in ≈5% of porcine and human fibroblasts and also upregulated the expression of the cardiac genes, MYH6 and TNNT2. When cocultured with murine cardiomyocytes, cTnT‐expressing porcine cardiac fibroblasts exhibited spontaneous contractions. Administration of GMT plus either H/My or miR‐590 alone also downregulated fibroblast genes COL1A1 and COL3A1. miR‐590 was shown to directly suppress the zinc finger protein, specificity protein 1 (Sp1), which was able to substitute for miR‐590 in inducing cellular reprogramming.ConclusionsThese data support porcine studies as a surrogate for testing human cardiac reprogramming, and suggest that miR‐590‐mediated repression of Sp1 represents an alternative pathway for enhancing human cardiac cellular reprogramming.

Cardiac reprogramming factor Gata4 reduces postinfarct cardiac fibrosis through direct repression of the profibrotic mediator snail

“…Although we were able to demonstrate differences between groups despite this variability, we do not have a good explanation for this variability between groups. In this context, we would note that we assayed RNA expression at 14 days after reprogramming factor infection based upon original research in this field suggesting that cellular reprogramming occurs over several weeks, 31,32 but additional studies demonstrated similar expression patterns as early as 5 days post-infection (data not shown).…”

Understanding the Role of the Transcription Factor Sp1 in Ovarian Cancer: from Theory to Practice

“…Similarly, MiR-590 a direct repressor of Sp1 has also been studied in conversion studies. When miR-590-mediated repression of Sp1 was done on human cardiac cells, it was found that it significantly upregulated the associated genes and promoted cardiac cellular reprogramming, showing that Sp1 may be an intermediary in this step [106] The E-Ras/JNK signalling is a critical mechanism to generate iPS cells by transduction of 4 factors. E-Ras was found to enhance binding of Sp1 on the cyclins to promote cell proliferation and reprogramming.…”

“…showing that Sp1 may be an intermediary in this step [106] The E-Ras/JNK signalling is a critical mechanism to generate iPS cells by transduction of 4 factors. E-Ras was found to enhance binding of Sp1 on the cyclins to promote cell proliferation and reprogramming.…”

Direct Cardiac Reprogramming: Progress and Promise

“…Zhao et al used a combination of GMHT, miRNA-1, miRNA-133, miRNA-208, miRNA-499, Y-27632, and A83-01 in MEFs and mouse adult fibroblasts to achieve ~60% cardiac troponin T+ and 60% α -actinin+ iCMs [ 29 ]. miRNA-590, a miRNA that can induce adult cardiomyocyte proliferation, was recently shown to be able to replace HAND2 and MYOCD in GMT direct reprogramming experiments using human and porcine fibroblasts [ 38 , 39 ]. While GMT was initially shown to be sufficient for cardiac reprogramming, further studies have indicated that a multiprong approach may be necessary to enhance reprogramming and could hold great promise for future in vivo clinical application.…”