Yocyte genes for example -MyHC and ANP. Similarly, addition of FGF-2, BMP-2, and combinations of both evoked an expression of cTnI (Fig. 1C) and a limited set of cardiac marker genes including Nkx-2.5 and GATA-4 in mBM-MASC1 and mBM-MASC2 (data not shown). According to the person experiment, in between 8 and 15 (n = 7) of mesenchymal adult stem cells (MASCs) stained optimistic for either Nkx-2.5 or GATA-4. However, close inspection of cTnI-positive cells revealed a highly aberrant cellular morphology (i.e., no crossstriations, flat shape, irregular size) that was not characteristic of cardiomyocytes (Fig. 1C). Moreover, we never spotted cells with an organized contractile apparatus or cells that underwent spontaneous contractions, that are hallmarks of functional cardiac muscle cells. We did not observe an initiation from the cardiac and PDGF-DD Proteins Biological Activity skeletal muscle program in mBM-MASCs when cells had been treated with conditioned medium from Wnt-expressing cells (data not shown), indicating that either direct cell-to-cell contacts are NT-4/5 Proteins Biological Activity needed or that the concentration of biologically active Wnt molecules in the medium didn’t suffice to stimulate expression of myogenic markers. Comparable final results had been obtained working with mesenchymal stem cells that were derived from different other tissues which includes heart and skeletal muscle. Though these cells displayed minor variations in the expression of stem cell marker molecules, they showed virtually precisely the same capabilities as bone-marrow-derived cells analyzed within this study (F. Belema-Bedada, in prep.).Fusion to myotubes or cardiomyocytes will be the predominant mechanism of mBM-MASCs to achieve complete myogenic differentiationSeveral groups have claimed to receive contracting heart muscle cells and functional skeletal muscle cells in vitro immediately after cocultivation of stem cells from different sources with bona fide muscle cells or following injection into regenerating host tissues in vivo. These reports are in apparent conflict to our observations, which indicated only a partial activation from the heart and skeletal muscle programs. It seems possible, nevertheless, that cocultivation of differentiated cells with stem cells may well give rise to a mixture of cells displaying a partially differentiated phenotype too as at least some semifunctional hybrid cells, which are derived from a fusion of uncommitted stem cells with totally differentiated cells. Actually, our cocultures of skeletal muscle cells or cardiomyocytes with MASCs labeled either with DiI or by virus-mediated delivery from the GFP gene also yielded labeled spontaneously contracting cardiomyocytes and myotubes that were apparently derived from labeled MASCs (information not shown). Such cells stained positive for each the cell tracking dye (GFP or DiI) and for musclecell-specific sarcomeric proteins (MyHC and cTnI inside the case of cardiomyocytes) (Fig. 2; Supplementary Fig. 1), raising the possibility that either differentiation of MASCs or fusion with C2C12 myoblasts had occurred. The appearance of double-labeled skeletal muscle cells was not a uncommon event. In selected experiments up to five.9 1.3 (n = 6) of all labeled MASCs stained positive with MyHC myotubes, although this ratio varied significantly (two- to threefold) amongst individual tests.GENES DEVELOPMENTSchulze et al.mation of hybrid myotubes was not entirely abrogated (Fig. 2F), while substantially fewer double-labeled cells were observed (0.6 0.three , n = 4). These data strongly suggest that the generation of functional heart and sk.
