模拟微重力对人牙髓干细胞细胞骨架及迁移能力影响的研究

目的:初步研究模拟微重力(simulated microgravity,SMG)对人牙髓干细胞(human dental pulp stem cells,hDPSCs)细胞骨架及迁移能力影响的信号通路机制。方法:采用酶消化法分离培养hDPSCs,并将其接种于PLGA支架上进行模拟微重力培养及普通重力培养。72 h后收集细胞,行Realtime-PCR检测LARG(leukemia-associated Rho GEF factor)基因的mRNA相对定量表达;拖拽实验检测RhoA-GTP表达情况;western blot检测RhoA及下游信号分子LIMK-P蛋白表达情况。结果:模拟微重力下培养的hDPSCs细胞LARG基因的mRNA表达下调(P<0.05);微重力下RhoA-GTP、RhoA及LIMK-P蛋白表达均较对照组下调(P<0.05)。结论:模拟微重力下人牙髓干细胞细胞骨架的重排,细胞迁移能力的改变可能与RhoA/ROCK信号通路有关。

Study of Simulated Microgravity on Cytoskeleton and Migration Ability of hDPSCs.

Objective: To investigate the effect of RhoA/ROCK signaling pathway on cytoskeleton and migration ability of human dental pulp stem cells (hDPSCs) in simulated microgravity (SMG). Methods: hDPSCs were seeded on PLGA scaffolds, and were cultured both on the conditions of conventional environment and SMG for 72 hours. The expression levels of LARG and GAPDH mRNA were measured using the real-time quantitative PCR method. Pull-down assay and western-blot were conducted to detect the RhoA-GTP,RhoA,LIMK-P expression levels. Results: The gene expression of LARG, a member of the Rho guanine nucleotide exchange factor (GEF) family, is observed to be downregulated under SMG(P<0.05);The RhoA-GTP,RhoA,LIMK-P protein expression levels are downregulated under microgravity(P<0.05). Conclusion: Simulated microgravity can inhibit RhoA/ROCK signaling pathway to change human dental pulp stem cells (hDPSCs) cytoskeleton structure and enhance the adhesion ability, reduce the ability to migrate.