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目的:探讨机械牵拉后人肺微血管内皮细胞(human pul monary microvascular endothelial cells,HPMVECs)单层通透性、F-肌动蛋白(F-actin)的变化及它们之间的关系,并探讨Rho激酶抑制剂Y27632对上述变化的影响。方法:体外培养HPMVECs,磁扭力刺激(magnetic twisting sti mulation,MTS)实施机械牵拉,随机分为3组,对照组(C组):细胞不经MTS,在细胞培养箱中培养1h、2h;机械牵拉组(M组):MTS1h、2h;抑制剂组(Y组):细胞与Rho激酶抑制剂Y27632(10μmol/L)孵育2h后给予MTS1h、2h;用FITC标记的右旋糖苷在transwell模型中的通透率检测HPMVECs单层通透性,用免疫荧光染色检测HPMVECs的F-actin分布。结果:M组的HPMVECs单层通透性较C组显著增加(P<0.05),且MTS2h后HPMVECs单层通透性较MTS1h高(P<0.05);Y组则较M组显著降低(P<0.05)。C组细胞F-actin主要分布在细胞边缘;而MTS引起F-actin聚合形成应力纤维,而以刺激2h明显;而Y组应力纤维形成较M组明显减少。结论:机械牵拉可引起HPMVECs的单层通透性增高,F-actin聚集形成应力纤维,且HPMVECs单层通透性增高的机制可能与F-actin聚集成应力纤维密切相关。Y27632可通过抑制应力纤维形成而减轻机械牵拉对HPMVECs单层通透性的影响。
Objective: To investigate the changes of monolayer permeability and F-actin in human pulmonar microvascular endothelial cells (HPMVECs) after mechanical pulling and the relationship between them Effect of kinase inhibitor Y27632 on these changes. Methods: HPMVECs were cultured in vitro and mechanically pulled by magnetic twisting stimulation (MTS). The cells were randomly divided into 3 groups: control group (C group): cells were cultured for 1 h and 2 h in a cell incubator without MTS; (M group): MTS1h, 2h; inhibitor group (Y group): cells were incubated with Rho kinase inhibitor Y27632 (10μmol / L) for 2h and then MTS for 1h, 2h; FITC-labeled dextran in transwell The permeability of the model was used to detect the monolayer permeability of HPMVECs. The distribution of F-actin in HPMVECs was detected by immunofluorescence staining. Results: The monolayer permeability of HPMVECs in group M was significantly higher than that in group C (P <0.05), and the monolayer permeability of HPMVECs was higher in M group than that in MTS2h group (P <0.05) <0.05). F-actin in group C was mainly distributed on the periphery of cells. MTS induced the aggregation of F-actin to form stress fibers, which was obviously stimulated by 2 h. However, the stress fibers in group Y were significantly reduced compared with M group. CONCLUSION: Mechanical stretch can induce single layer permeability of HPMVECs to increase, F-actin aggregates to form stress fibers, and the mechanism of single layer permeability of HPMVECs may be closely related to the aggregation of F-actin into stress fibers. Y27632 can reduce the influence of mechanical stretch on the monolayer permeability of HPMVECs by inhibiting the formation of stress fibers.