Differentiation of embryonic-like stem cells derived from human bone marrow into multinucleated fibers in vitro

AIM: To compare the capacity of in vitro differentiation into multinucleated fibers between embryonic-like stem cells (ELSCs) and mesenchymal stem cells (MSCs) derived from human bone marrow. METHODS: To isolate ELSCs, human bone marrow mononuclear cells were cultured in gelatin-coated flask with serum-free Knockout-DMEM medium designed for the expansion of human embryonic stem cells. MSCs were isolated from the same bone marrow by the traditional method. The morphological characters of both ELSCs and MSCs were observed under inverted phase-contrast microscope, and the expression of their multipotent antigen markers was identified by immunofluorescent staining. ELSCs and MSCs were cultured in myogenic differentiation medium. The protein levels of muscle-specific antigen markers myosin heavy chain (MHC), myogenin and MyoD were detected by the method of immunostaining. The mRNA expression of MHC, myogenin and MyoD was detected by RT-PCR. The capacity of in vitro differentiation into multinucleated fibers was compared between ELSCs and MSCs by calculating the proportion of MHC-positive multinucleated fibers. RESULTS: ELSCs, which weakly expressed the multipotential markers Oct-4, Nanog-3 and Sox-2, were isolated from bone marrow by the method of serum-free medium. ELSCs appeared smaller, slenderer and more homogeneous, and were morphologically different from MSCs derived from the same marrow. No multipotential marker in MSCs was expressed. ELSCs and MSCs were induced into long multinucleated fibers expressing MHC and myogenin at mRNA and protein levels by culturing in the myogenic differentiation medium. However, on the 10th day after induction, the proportion of the MHC-positive fibers in ELSCs was (25.7?4.1)%, and the proportion in MSCs was (15.8?7.6)%.The capacity for differentiation into muscle in ELSCs was significantly higher than that in MSCs (P<0.05). CONCLUSION: Bone marrow ELSCs are induced into multinucleated fibers and have the stronger myogenic differentiation capacity than MSCs derived from the same marrow. ELSCs are a more ideal candidate for muscular disease therapy.