Due to this, these -glucans are commercially available and consumed as food supplements

ssessed by a direct cell count. Data are mean SD of 4 independent experiments. Myotube width increased from day 4 to day 6 in myoblasts cultured on collagen I and muscle matrices, while widths MedChemExpress RS1 remained constant in myotubes formed on fibronectin over the same time period. Overall, there was no difference in myotube width between the three substrates. The number of nuclei per myofibre was also similar for all three substrates and by day 6 of differentiation, myotubes on all substrates PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19667322 developed striations. Striations were observed in myotubes on muscle matrix at day 4 of differentiation, whereas this was not the case when C2C12 cells were grown and differentiated on collagen I or fibronectin. 13 / 27 An Acellular Muscle Matrix Supports Myoblast Differentiation PubMed ID:http://www.ncbi.nlm.nih.gov/pubmed/19666601 Fig 6. Immunofluorescent staining of mature C2C12 myotubes. C2C12 myoblasts were cultured for 4 and 6 days in differentiation medium on collagen I, fibronectin and muscle matrix, and visualized using the mouse anti-MyHCB mAb and goat anti-mouse AF488. Arrows indicate striations. Nuclei are stained with DAPI. Scale bars are 50 m for ab, de, gh and g insert; and 25 m for c, f and i. Myotube width was measured using ImageJ software, values are the mean and SD of 50 myotubes.An Acellular Muscle Matrix Supports Myoblast Differentiation Gene expression of myogenic markers in C2C12 cells The expression of genes involved in different stages of myogenesis was assessed using qPCR. These genes were myogenic factor 5 and myogenin, both transcription factors belonging to the myogenic regulatory factor family; three isoforms of myosin heavy chain; and two muscle actin isoforms. Of the four reference genes tested, SDHA showed the most stable expression across all conditions and was used as the reference gene for normalization of data. Gene expression by C2C12 cells on days 1, 4 and 8 of differentiation was assessed when C2C12 myoblasts were cultured on collagen I, fibronectin and muscle matrix. Myf5 showed similar levels of expression at days 1, 4 and 8 of differentiation on collagen I. On fibronectin and muscle matrix surfaces, Myf5 levels at day 8 were significantly lower than levels at day 1. MYOG mRNA increased on muscle matrix coated surfaces by approximately 4-fold between days 1 and 8. In contrast, on collagen I and fibronectin coated surfaces, MYOG levels peaked at day 4 before declining. An increase in the expression of MYH1, MYH7 and ACTA1 was observed from differentiation day 1 to day 4, and maintained at day 8 on all matrices. MYH3 and ACTC1, which are expressed in immature muscle, increased from differentiation day 1 to day 4, before decreasing at day 8. A similar trend was seen on all matrices, although the decrease in MYH3 expression was most pronounced in cells on fibronectin and muscle matrix. These data, plus the fact that on muscle matrix MYH1 expression did not increase from day 4 to day 8, suggests that differentiation occurred earlier in cells cultured on muscle matrix compared to collagen I, but at about the same time as those on fibronectin. C2C12 cell adhesion on three dimensional muscle matrices Decellularized 3D rat muscle was used to examine how an intact muscle matrix affected cell adhesion and proliferation. Cell proliferation in serum-free cultures on 3D matrices was assessed using a Click-iT EdU assay, which labels recently synthesized DNA with the coloured thymidine analogue EdU. Many cells were proliferating at day three after seeding and the 3D colour coded imag