Differentiation of human adipose‐derived stem cells into beating cardiomyocytes

Abstract

Human adipose-derived stem cells (ASCs) may differentiate into cardiomyocytes and this provides a source of donor cells for tissue engineering. In this study, we evaluated cardiomyogenic differentiation protocols using a DNA demethylating agent 5-azacytidine (5-aza), a modified cardiomyogenic medium (MCM), a histone deacetylase inhibitor trichostatin A (TSA) and co-culture with neonatal rat cardiomyocytes. 5-aza treatment reduced both cardiac actin and TropT mRNA expression. Incubation in MCM only slightly increased gene expression (1.5- to 1.9-fold) and the number of cells co-expressing nkx2.5/sarcomeric α-actin (27.2%versus 0.2% in control). TSA treatment increased cardiac actin mRNA expression 11-fold after 1 week, which could be sustained for 2 weeks by culturing cells in cardiomyocyte culture medium. TSA-treated cells also stained positively for cardiac myosin heavy chain, α-actin, TropI and connexin43; however, none of these treatments produced beating cells. ASCs in non-contact co-culture showed no cardiac differentiation; however, ASCs co-cultured in direct contact co-culture exhibited a time-dependent increase in cardiac actin mRNA expression (up to 33-fold) between days 3 and 14. Immunocytochemistry revealed co-expression of GATA4 and Nkx2.5, α-actin, TropI and cardiac myosin heavy chain in CM-DiI labelled ASCs. Most importantly, many of these cells showed spontaneous contractions accompanied by calcium transients in culture. Human ASC (hASC) showed synchronous Ca2+ transient and contraction synchronous with surrounding rat cardiomyocytes (106 beats/min.). Gap junctions also formed between them as observed by dye transfer. In conclusion, cell-to-cell interaction was identified as a key inducer for cardiomyogenic differentiation of hASCs. This method was optimized by co-culture with contracting cardiomyocytes and provides a potential cardiac differentiation system to progress applications for cardiac cell therapy or tissue engineering.

Gene and Protein Expression Analysis of Mesenchymal Stem Cells Derived From Rat Adipose Tissue and Bone Marrow

“…22-24 On the other hand, ASC are also reported to be able to differentiate into cardiomyocytes. 25 Therefore, ASC and BM-MSC both might improve cardiac function by supplementing cardiomyocytes, as well as in a paracrine manner, although we did not investigate differences in differentiation into cardiomyocytes between them.…”

Epimuscular Fat in the Human Rotator Cuff Is a Novel Beige Depot

“…Adipose tissue biopsies and lipoaspirate were digested to isolate the ASC population, as previously described [24]. In brief, the biopsies were rinsed during transportation from the operating room in Dulbecco's modified Eagle's medium (DMEM; low glucose), transferred to a digestive solution composed of 0.3% collagenase type I, 60 U/ml dispase II, and 1% penicillin/ streptomycin in low-glucose DMEM, minced with forceps, and allowed to incubate at 37°C for 30 minutes.…”

Non-classical Signalling Mechanisms in Stem Cells

“…Previous studies by Muller-Borer et al (2004) suggested that synchronized Ca 2+ signalling via GJIC co-cultured with neonatal cardiomyocytes induces rat liver stem cells to express cardiac transcription factors and acquire a phenotype resembling cardiomyocytes (Anderson et al, 2007;Muller-Borer et al, 2004). Similarly, we have recently shown gap junctions established between human adiposederived MSC and neonatal rat cardiomyocytes in co-culture, which induces the expression of cardiac genes and spontaneous cardiomyocyte-like contractions in the human cells (Choi et al, 2010a) (Figure 2). All this points to a growing appreciation of the role of gap-junctionmediated Ca 2+ waves in modulating gene expression and promoting differentiation of stem cells.…”