不同牵张应力对大鼠BMSCs增殖和成骨分化作用

Proliferation and Osteogenic Differentiation of Rat BMSCs by Different Tension Stress

目的:考察牵张应力大小和作用时间对骨髓间充质干细胞(BMSCs)增殖和成骨分化的影响。方法:SPF级SD大鼠10只,分离培养并采用成骨和成脂分化法鉴定BMSCs。细胞分组设置为牵张应力加载大小(0.5%拉伸幅度,5%拉伸幅度,10%拉伸幅度、15%拉伸幅度)和作用时间(0.5h,6h,12h,24h,48h,72h)。CCK-8法测定BMSCs增殖。BCIP/NBT法测定细胞碱性磷酸酶活性。茜素红和油红O染色法检测细胞成骨和成脂分化。结果:10%拉伸幅度的牵张应力作用12h,可最大提高BMSCs增殖速度1.53倍,高于5%和15%拉伸幅度组,差异具有统计学意义(P<0.01)。5%拉伸幅度的牵张应力作用48h,可最大提高BMSCs碱性磷酸酶表达4.2倍,提高BMSCs矿化结节形成65%,减少BMSCs形成脂滴32%,高于10%和15%拉伸幅度组,差异具有统计学意义(P<0.05或P<0.01)。结论:牵张应力拉伸幅度为10%作用时间12h促进BMSCs增殖效果最佳。牵张应力拉伸幅度为5%作用时间48h提高BMSCs碱性磷酸酶和矿化结节形成最显著,进而对BMSCs发挥促进成骨分化作用。牵张应力促进BMSCs增殖和成骨分化所需牵张应力大小及作用时间不同。

Objective:To examine the effects of the magnitude and duration of drafting stress on the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells(BMSCs).Methods:Ten SPF-grade SD rats were isolated and cultured and BMSCs were identified by osteogenic and lipogenic differentiation methods.Cell grouping was set for the magnitude of drafting stress loading(0.5%stretch,5% stretch,10% stretch,15% stretch)and duration of action(0.5h,6h,12h,24h,48h,72h).Proliferation of BMSCs was measured by CCK-8 method.The BCIP/NBT method was used to determine cellular alkaline phosphatase activity.Cellular osteogenic and lipogenic differentiation was detected by alizarin red and oil red O staining.Results:Tensile stress at 10% stretch for 12h maximally increased the proliferation rate of BMSCs by 1.53-fold,which was higher than that of the 5% and 15% stretch groups,with statistically significant differences(P<0.01).tensile stress at 5% stretch for 48h maximally increased alkaline phosphatase expression of BMSCs by 4.2-fold,increased mineralized nodule formation of BMSCs by 65%,and decreased BMSCs formation of lipid droplets by 32%,which was higher than the 10% and 15% stretch amplitude groups,with statistically significant differences(P<0.05 or P<0.01).Conclusion:The best effect of tensile stress stretching at 10% for 12h was observed in promoting the proliferation of BMSCs.Draft stress of 5% for 48h had the most significant effect on the formation of alkaline phosphatase and mineralized nodules in BMSCs,which in turn promoted osteogenic differentiation of BMSCs.The magnitude and duration of tension stress required to promote the proliferation and osteogenic differentiation of BMSCs varied.