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目的观察转化生长因子(TGF)β_1刺激后肝星状细胞迁移和细胞骨架的变化,并观察TGFβ_1刺激对肝星状细胞内Rho三磷酸鸟苷(GTP)酶表达的影响。方法分离原代大鼠肝星状细胞,用Transwell Chamber观察不同浓度TGFβ_1直接及趋化刺激后细胞迁移改变,FITC标记的鬼笔环肽标记F-actin,共聚焦激光扫描显微镜观察细胞骨架的变化,GST pull-down分析检测不同浓度TGFβ_1刺激后活性RhoA、Racl及Cdc42的表达。结果FGFβ_1直接及趋化刺激后肝星状细胞迁移均比对照组增加,≥5 ng/ml浓度时增加明显(直接刺激后：130．90±7．64比102．93±1．01,趋化刺激后205．17±10．78比102．93±1．01,P＜0．05)；TGFβ_1刺激后细胞形态改变,刺激5 min后层状伪足出现,并随时间推移应力纤维逐渐增多；不同浓度TGFβ_1刺激后活性Cdc42、RhoA表达较对照组均增强,≥5 ng/ml浓度时表达明显增加(GTP-Cdc42：0．273±0．024比0．176±0．001,P＜0．05；GTP-RhoA：0．176±0．005比0．096±0．004,P＜0．05),而活性的Rac1表达无明显改变。结论TGFβ_1可导致肝星状细胞骨架改变,并可通过激活Cdc42、RhoA GTP酶信号途径,增加细胞迁移。
Objective To observe the changes of hepatic stellate cell migration and cytoskeleton after stimulated by transforming growth factor β1 (TGFβ1), and observe the effect of TGFβ1 on the expression of GTPase in hepatic stellate cells. Methods Primary rat hepatic stellate cells were isolated. Transwell chamber was used to observe the direct and chemotactic effects of different concentrations of TGFβ 1. The FITC-labeled phalloidin labeled F-actin and confocal laser scanning microscope were used to observe the change of cytoskeleton GST pull-down assay was used to detect the expression of active RhoA, Racl and Cdc42 after stimulation with different concentrations of TGFβ 1. Results FGFβ 1 directly and chemotactically stimulated the hepatic stellate cells to migrate more than the control group, and increased obviously at the concentration of 5 ng / ml (after direct stimulation: 130.90 ± 7.64 vs 102.93 ± 1.01, 205.17 ± 10.78 vs 102.93 ± 1.01 after stimulation (P <0.05). The morphological changes of cells after stimulation with TGFβ_1 and the appearance of lamellipodia appeared 5 min after stimulation, and the stress fibers gradually increased with time (P <0.05). Compared with the control group, the expression of active Cdc42 and RhoA in TGFβ_1 group increased significantly (P <0.05), and the expression of RhoA and Cdc42 in the group of GTP-Cdc42: 0.273 ± 0.024 and 0.176 ± 0.001 <0.05; GTP-RhoA: 0.176 ± 0.005 vs. 0.096 ± 0.004, P <0.05), whereas the active Rac1 expression did not change significantly. Conclusion TGFβ 1 can lead to changes of hepatic stellate cytoskeleton and increase cell migration by activating Cdc42 and RhoA GTPase signaling pathways.