由BMP-4基因转入C2C12细胞引起的成肌细胞到成骨细胞的表型转变

目的:研究BMP-4基因对C2C12细胞分化的影响。方法:BMP-4基因在pCI-neo质粒中构建后转入C2C12细胞。转入pCI-neo-BMP-4重组质粒的细胞、转入pCI-neo质粒的细胞及没有转染的细胞在相同的条件下培养。结果:转入pCI-neo质粒的细胞及没有转染的细胞在增殖期和分化期均不表达BMP-4,在未分化期表现为典型的星形形态,并相互融合成细长而多核的肌小管。而转入pCI-neo-BMP-4重组质粒的细胞在增殖期和分化期均大量表达BMP-4,形态上不形成肌小管,而是形成类似成骨细胞的形态,并高水平表达碱性磷酸酶、在培养基中分泌骨钙素。结论:表明将BMP-4基因转入C2C12细胞可改变其肌源的分化途径,而分化为成骨细胞系,且这种分化转向是基因水平上的。

Phenotypic transformation of myoblast to osteoblast induced by introduction of bone morphogenetic protein-4 coding sequence into C2C12 cells

AIM: To study the effect of introduced bone morphogenetic protein-4 (BMP-4) gene on the differentiation of C2C12 cells. METHODS: BMP-4 gene was constructed into pCI-neo plasmid, which was then introduced into C2C12 cells. Cells transfected with pCI-neo-BMP-4 recombinant plasmid or pCI-neo plasmid alone and non-transfected cells were cultured in the same condition and studied. RESULTS: It was demonstrated that cells with/without pCI-neo plasmid showed no expression of BMP-4 both in their proliferating and differentiating stages. Cells transfected with pCI-neo-BMP-4 showed abundant BMP-4 expression both in their proliferating and differentiating stages. The undifferentiated C2C12 cells with/without pCI-neo plasmid displayed the characteristic stellate morphology, and fused with each other to form characteristic thin, elongated multinucleated myotubes. In contrast, the cells transfected with pCI-neo-BMP-4, subject to both proliferation or differentiation stages did not form myotubes, instead they formed osteoblast-like morphology with high level expression of alkaline phosphatase (ALP) in the cells and also secreted osteocalcin in their culture medium. CONCLUSION: The introduction of BMP-4 gene into C2C12 cells could convert their committed myogenic differentiation pathway into that of osteoblast lineage, and the conversion of differentiation pathway is heritable.