Obtaining spontaneously beating cardiomyocyte-like cells from adipose-derived stromal vascular fractions cultured on enzyme-crosslinked gelatin hydrogels

Abstract

Heart failure often develops after acute myocardial infarction because the injured myocardial tissue fails to recover or regenerate. Stem cell transplantation using adult cell sources, such as adipose-derived stromal vascular fraction (SVF), draws extensive attention. In this study, SVF cells were isolated from rat adipose tissue and cultivated on enzyme-crosslinked gelatin hydrogels. Morphological features of cell development and spontaneous beating behavior from these cells were observed and recorded. Cardiac phenotypes were characterized via immunofluorescence staining, and the expression of cardiac-specific genes was measured via RT-PCR. The functional assessment of SVF-derived cardiomyocyte-like cells (SVF-CMs) was performed by detecting cellular calcium transient activities and pharmacological responses. Results showed that most SVF-CMs exhibited elongated myotubule shapes and expressed cardiac troponin I strongly. SVF-CMs expressed cardiac-specific RNA (including transcription factors GATA binding protein 4) and myocyte enhancer factor 2c, as well as the structural proteins, namely, sarcomere actinin alpha 2, cardiac troponin I type 3, cardiac troponin T type 2, and cardiac gap junction protein alpha 1. Their beating mode, calcium activities, and pharmacological responses were similar to those of native CMs. Spontaneously beating SVF-CMs can be derived from adipose tissue-derived SVFs, and enzyme-crosslinked gelatin hydrogel promoted the cardiac differentiation of SVF cells.

Transcriptional regulation profiling reveals disrupted lipid metabolism in failing hearts with a pathogenic phospholamban mutation

“…49 A previous study showed the possibility of differentiating adipose-derived stromal vascular fraction cells into cardiomyocytelike cells. 50 Another study showed the transduction of cardiac mesenchymal progenitors into adipocytes. 51 Additionally, fibroblasts can also be differentiated into adipocytes, directly and indirectly, promote the differentiation of adipocytes indirectly via the adipose progenitor cells.…”

Myocyte-specific enhancer factor 2c triggers transdifferentiation of adipose tissue-derived stromal cells into spontaneously beating cardiomyocyte-like cells

“…Planat-Bénard et al reported transdifferentiation of Ad-SVF into beating CMs using semi-liquid medium and supplementation with cytokines and growth factors 17 . The use of semisolid culture with enzyme-crosslinked gelatine hydrogels obviates the need for supplements to induce differentiation of SVF into beating CMs 18 . By contrast, we achieved differentiation of SVF into spontaneously beating CMs using an uncoated culture dish and standard liquid medium supplemented only with a reducing agent.…”

“…As clinical trials have shown, somatic stem cells, including bone-marrow-derived stromal cells (BMSCs) and adipose tissue-derived stromal vascular fraction (Ad-SVF), have advantages in terms of safety and availability over iPS cells 2 , 15 , but they rarely differentiate into spontaneously beating cardiomyocytes. Several groups have achieved transdifferentiation of bone-marrow or Ad-SVF into spontaneously beating cardiomyocyte-like cells (CMs); however, supplementation with DNA methyltransferase 1 inhibitor, growth factors, or cytokines, or use of a semi-solid medium, was required 16 – 18 . Additionally, because a small number of CMs was produced, their characteristics could not be evaluated in detail.…”

Improved Efficiency of Cardiomyocyte-Like Cell Differentiation from Rat Adipose Tissue-Derived Mesenchymal Stem Cells with a Directed Differentiation Protocol

“…So far, only two studies and ours quantify the cardiomyocyte differentiation efficiency by flow cytometry [7, 16]. The great majority of ASC-derived cardiomyocyte protocols count the number of beating clones or the percentage of positive expressing cells for cardiac markers in a qualitative manner [6, 14, 17–19, 23, 29–31].…”