持续牵张促进小鼠骨髓间充质干细胞成骨向分化机制的研究

目的:研究p38MAPK在持续性机械牵张力促进C57BL/6J小鼠骨髓间充质干细胞成骨分化过程中的作用机制。方法:C57BL/6J小鼠BMSCs被随机分为对照组和牵张力阻断组。牵张力阻断组预先用p38MAPK特异性阻断剂SB203580处理1 h,后用Flexercell加力仪加载0.5Hz,0.8%的牵张力。对照组不做特殊处理。分别对两组细胞施加0、30 min和60 min的牵张力。采用Western blot检测P-p38MAPK蛋白的表达情况,Real time-PCR检测成骨基因ALP、COL I、OCN mRNA的表达变化。结果:C57BL/6J小鼠BMSCs传代后细胞生长状态稳定,呈梭形,具有多向分化潜能。Real time-PCR结果显示对照组BMSCs中ALP、COL I、OCN mRNA表达在不同时间点(30 min与0 min,60 min与30 min)间差异均有统计学意义(P<0.05);对照组ALP、COLI、OCN的表达量均明显高于同一时间点(30 min和60 min)牵张力阻断组(P<0.05)。Western blot结果显示,对照组内BMSCs中P-p38MAPK的蛋白水平在不同时间点间(30 min与0 min,60 min与30 min)差异均有统计学意义(P<0.05);对照组P-p38MAPK蛋白的表达量均明显高于同一时间点(30 min和60 min)牵张力阻断组(P<0.05)。结论:所采用的0.5 Hz,0.8%的机械牵张力在持续牵张力下可通过p38MAPK 通路促进小鼠BMSCs向成骨细胞分化。

Mechanism of Osteogenesis of BMSCs under Continuous Mechanical Tension Force

Objective: To study the role of p38MAPK signal pathway in regulating osteogenic differentiation of C57BL/6J mouse BMSCs under the continuous mechanical tension force. Methods: The BMSCs of primary culture were divided randomly into control group and tension inhibitor group. The tension inhibitor group was pretreated for 1 hour with SB203580 which was one of p38MAPK pathway specific inhibitor, and then loaded with mechanical stimulation by Flexercell. The tension force (0.5Hz, 0.8%) was inflicted to both groups at 0, 30 and 60 min, respectively. p38MAPK and P-p38MAPK protein were detected by Western blot, and the expression of osteogenic genes ALP, COLI, OCN were detected by real time-PCR. Results: C57BL/6J mouse BMSCs grew well with typical shuttle shape and capacities of multi-differentiation. Real time-PCR and Western blot results showed that the expression of ALP, COLI, OCN mRNA and P-p38MAPK protein in the control group was significant different among different loading time points. The mRNA and P-p38MAPK protein expression in control group was markedly higher than those in the tension inhibitor group at different time points (P<0.05). Conclusion: The continuous tension force (0.5Hz, 0.8%) can promote osteogenesis of BMSCs through p38MAPK signal pathway.