1-磷酸神经鞘氨醇对大鼠骨骼肌成肌细胞分化的影响

Effects of Sphingosine 1-phosphate on Differentiation of Skeletal Myoblast Cell in Rats

【目的】观察1-磷酸神经鞘氨醇(S1P)对大鼠骨骼肌成肌细胞向成熟肌细胞分化、形成肌管的作用,探索定向诱导肌分化的途径及其分子机制。【方法】分离和培养大鼠骨骼肌成肌细胞,含不同浓度S1P的高糖DMEM培养基培养,收集细胞,在相差显微镜和共焦显微镜下观察形态变化,并通过免疫细胞化学方法分别检测成肌细胞分化的早期特异性标志—肌球蛋白和生肌素表达的改变。使用磷酸神经鞘氨醇激酶(SphK)活性抑制剂DMS和HACPT后观察S1P对骨骼肌成肌细胞的生肌素表达的作用。【结果】S1P能明显促进骨骼肌成肌细胞细胞形成肌小管,诱导2～3 d开始有肌管形成,之后肌管越来越多,到6～7 d达高峰;同对照组相比随S1P浓度增高,细胞核生肌素阳性率逐渐增高(P<0.01);SphK活性抑制剂抑制S1P促进骨骼肌成肌细胞生肌素阳性率的增高(P<0.01)。【结论】体外S1P可能通过提高SphK活性调节成肌细胞向成熟肌细胞分化。

[Objective] To explore the effects and mechanism of sphingosine 1-phosphate （SIP） on differentiation of skeletal myoblast cells into mature myocytes and consequent formation of myotubes in rats. [Methods] Primary skeletal myoblast cells in rats were separated and cultured, here we described the roles of S1P in the regulation of differentiation by observing alteration of their appearance under phase contrast microscope and detected for the expression change of muscle-associated gene myosine and myogenin by immunocytochemistry. Sphingosine kinase inhibitors were used to during S1P on differentiation of myoblast cells, [Results] S1P remarkably promoted myotude formation of myoblasts. Induced with S1P for 2-3 days, myotubes started to form. Later on, more and more myotubes appeared, and at the peak on 6-7 days. S1P induced myogenin positive nucleis higher with concentration-dependent form compare with control （P 〈 0.01）. But SphK inhibitors reduce SIP effect on differentiation of myoblast cells （P 〈 0.01）. [Conclusion] S1P may through sphingosine kinase promote differentiation of skeletal myoblast ceils into mature myocytes and formation of myotube.