消渴通痹方含药血清对高糖损伤后HUVEC细胞骨架及RhoA/ROCK信号通路的影响  

Xiaoke Tongbi Formula(消渴通痹方)Regulates the RhoA/ROCK Signaling Pathway and Cytoskeleton of High Glucose-Induced Human Umbilical Vein Endothelial Cells

作　　者：张欢 丁纪茹[1] 袁有才 段玉红[2] 张效科[2] ZHANG Huan;DING Jiru;YUAN Youcai;DUAN Yuhong;ZHANG Xiaoke(Affiliated Hospital of Chengdu University of Traditional Chinese Medicine,Chengdu 610075;Shaanxi University of Chinese Medicine,Xianyang 712046)

机构地区：[1]成都中医药大学附属医院,成都610075 [2]陕西中医药大学,咸阳712046

出　　处：《中药药理与临床》2023年第1期25-28,共4页Pharmacology and Clinics of Chinese Materia Medica

基　　金：国家自然科学基金资助项目(编号:81774304)。

摘　　要：目的:观察消渴通痹方含药血清对高糖损伤后人脐静脉内皮细胞(Human Umbilical Vein Endothelial Cells, HUVEC)F肌动蛋白(F-actin)、Ras基因家族成员A(Ras homolog gene family member A,RHOA)、Rho相关蛋白激酶(Rho-associated coiled-coil containing kinase, ROCK)蛋白表达的影响。方法:将高糖损伤后HUVEC模型分为高糖模型组、22.5 g/kg消渴通痹方5%、10%、15%含药血清组,另设正常对照组,应用免疫荧光法检测F-actin的动态变化,Western blot检测F-actin、ROCK、p-ROCK和RhoA蛋白表达。结果:高糖刺激HUVEC骨架蛋白F-actin聚集到细胞中央,形成大量的应力纤维;22.5 g/kg消渴通痹方5%、10%含药血清干预后能够改善F-actin的聚集状态,使F-actin重新分布在细胞周边。与正常对照组比较,高糖模型组F-actin积分光密度值升高,F-actin、p-ROCK、RHOA蛋白表达明显上调(P<0.05或P<0.01),ROCK蛋白表达明显下调(P<0.05);与高糖模型组比较,22.5 g/kg消渴通痹方5%含药血清组F-actin积分光密度值明显降低,F-actin、p-ROCK、RHOA蛋白表达明显下调(P<0.05),ROCK蛋白表达明显上调(P<0.05);10%含药血清组F-actin积分光密度值明显降低、F-actin蛋白表达明显下调(P<0.01或P<0.05),RHOA、ROCK蛋白表达明显上调(P<0.05)。结论:益气活血中药消渴通痹方可能通过抑制RHOA/ROCK信号通路的异常激活稳定或保护内皮细胞骨架。Objective:To observe the protective effect of Xiaoke Tongbi Formula(消渴通痹方)on the protein levels of F-actin, Ras homolog gene family member A(RhoA),Rho-associated coiled-coil containing kinase(ROCK) in high glucose-induced human umbilical vein endothelial cells(HUVECs). Methods:High glucose-induced HUVECs were assigned into a normal control group, a high-glucose model group, and groups of serum containing 5%,10%,and 15% Xiaoke Tongbi Formula administrated at a dose of 22.5 g/kg. Immunofluorescence assay was employed to detect the expression of F-actin, and Western blotting to determine the protein levels of F-actin, ROCK,p-ROCK,and RhoA. Results:High glucose stimulated the aggregation of the cytoskeleton protein F-actin to the center of HUVECs and the formation of a large number of stress fibers. The treatment with the serum containing 5% and 10% Xiaoke Tongbi Formula administrated at a dose of 22.5 g/kg alleviated the aggregation of F-actin and redistributed it around the periphery of the cell. Compared with the normal control group, the modeling increased the integrated optical density(IOD) of F-actin, up-regulated the protein levels of F-actin, p-ROCK,and RhoA(P<0.05 or P<0.01),and down-regulated the protein level of ROCK(P<0.05). Compared with the high glucose model group, the serum containing 5% drug decreased the IOD,down-regulated the protein levels of F-actin, p-ROCK,and RhoA(P<0.05),and up-regulated the protein level of ROCK(P<0.05). The serum containing 10% drug decreased the IOD of F-actin, down regulated the protein level of F-actin(P<0.05 or P<0.01),and up-regulated the protein levels of ROCK and RhoA(P<0.05). Conclusion:Xiaoke Tongbi Formula can stabilize the cytoskeleton by inhibiting the RhoA/ROCK signaling pathway, which provides a theoretical basis for the prevention and treatment of diabetic peripheral neuropathy.

关 键 词：消渴通痹方 糖尿病周围神经病变 人脐静脉内皮细胞 细胞骨架 Ras基因家族成员A/Rho相关蛋白激酶信号通路 

分 类 号：R285.5[医药卫生—中药学]