与机械牵张盆底肛提肌卫星细胞间接共培养对大鼠骨髓间充质干细胞成肌分化抗原和结蛋白表达的影响

Effect of mechanical stretch on the expressions of MyoD and Desmin in rat BMSCs with rat anus levator satellite cell co-culture

目的研究与机械牵张的盆底肛提肌卫星细胞间接共培养对大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)成肌分化的影响;探索在体外条件下诱导BMSCs向肌细胞分化的微环境的创建以及成肌分化抗原(myogenic differentiation antigen,MyoD)和结蛋白(des minfilament,Desmin)蛋白的表达。方法选择7 d龄SD大鼠,利用密度梯度离心法体外分离纯化大鼠BMSCs。取大鼠盆底肛提肌组织,自行分离传代,获得大鼠盆底肛提肌卫星细胞。对第3代盆底肛提肌卫星细胞施以10%形变,1 Hz,12 h的周期性单向机械牵张刺激,机械牵张后的盆底肛提肌卫星细胞与纯化培养后的第4代BMSCs间接共培养6d。采用实时定量RT-PCR方法检测共培养后BMSCs中MyoD和Desmin mRNA的表达情况;Western-blot法检测MyoD和Desmin的蛋白含量。结果与对照组相比,共培养组和机械牵张共培养组BMSCs的MyoD和Desmin mRNA和蛋白的表达水平均有显著增高(P<0.05);与共培养组相比,机械牵张共培养组BMSCs的MyoD和Desmin mRNA和蛋白的表达量也有显著增高(P<0.05)。结论与机械牵张刺激的盆底肛提肌卫星细胞间接共培养可以促进BMSCs合成MyoD和Desmin,诱导BMSCs向肌细胞分化。

Objective This study aims to evaluate the effect of mechanical stretch on the differentiation of bone marrow mesehchymal stem cells （BMSCs） with a co - culture with anus levator satellite cell, and the expressions of MyoD and Desmin. Methods BMSCs were isolated and identified from 7 d SPF SD rats. Rat anus levator satellite cells were obtained from rat anus levator. The 3rd passage of anus levator satellite cells were subjected to 10% deformation with 1 Hz, 12 h periodic one - way mechanical stretch stimulation, and the cells were then co - cultured with BMSCs for 6 d. Results The co - culture and co - culture with mechanical stretch groups induced more expression of MyoD and Desmin, compared to the groups without stimulation. Conclusion These results suggest that in an indirect co - culture system, anus levator satellite cell with mechanical stretch stimulation can promote BMSCs differentiation, reflecting in increased expression of MyoD and Desmin.