一贯煎联合骨髓间充质干细胞调控RhoA/ROCK1通路抑制大鼠肝纤维化的实验研究  

Yiguanjian Combined with Bone Marrow Mesenchymal Stem Cells Treats Liver Fibrosis in Rats via RhoA/ROCK1 Signaling Pathway

作　　者：李汶航 张睿[1] 崔欣怡 唐佐青[2] 油红捷[2] 杨铮[1] 付修文[1] 马赟[1] 刘文兰[1] LI Wenhang;ZHANG Rui;CUI Xinyi;TANG Zuoqing;YOU Hongjie;YANG Zheng;FU Xiuwen;MA Yun;LIU Wenlan(School of Traditional Chinese Medicine,Capital Medical University,Beijing 100069,China;School of Basic Medical Sciences,Capital Medical University,Beijing 100069,China)

机构地区：[1]首都医科大学中医药学院,北京100069 [2]首都医科大学基础医学院,北京100069

出　　处：《世界中医药》2022年第18期2563-2568,共6页World Chinese Medicine

基　　金：国家自然科学基金项目(81573879)。

摘　　要：目的:探究一贯煎联合骨髓间充质干细胞在治疗肝纤维化过程中对RhoA/ROCK1通路的作用。方法:将72只SD大鼠随机分为正常对照组、模型对照组、秋水仙碱组、一贯煎高剂量组、一贯煎中剂量组、一贯煎低剂量组、干细胞组、一贯煎+干细胞组;四氯化碳腹腔注射6周制备肝纤维化动物模型,干预4周后取材。记录大鼠阴虚表征情况;检测血清谷丙转氨酶(GPT)、谷草转氨酶(GOT)含量;HE、Masson染色观察病理改变;实时PCR、蛋白质印迹法检测肝组织中RhoA、Rho相关激酶1(ROCK1)、α-SMA和COL-1 mRNA及蛋白表达;IHC检测α-SMA、COL-1阳性面积。结果:与正常对照组比较,模型对照组大鼠具有明显阴虚表征,肝脏出现明显炎症浸润和纤维增生,肝组织RhoA、ROCK1、α-SMA、胶原蛋白-1(COL-1)mRNA与蛋白表达显著升高(P<0.05),α-SMA、COL-1阳性面积显著增加(P<0.05);与模型对照组比较,一贯煎高剂量组、一贯煎中剂量组、一贯煎低剂量组、骨髓干细胞、一贯煎+干细胞组大鼠的阴虚表征、肝脏炎症和纤维化程度减轻,RhoA、ROCK1、α-SMA和COL-1 mRNA与蛋白表达显著降低(P<0.05),α-SMA、COL-1阳性面积显著减少(P<0.05),其中一贯煎+干细胞组较一贯煎中剂量组、干细胞组效果更优(P<0.05)。结论:一贯煎与骨髓干细胞可通过调控RhoA/ROCK1通路抑制肝星状细胞活化从而发挥抗纤维化的作用,二者协同作用效果优于各自单独作用效果。Objective:To observe the effect of Yiguanjian(YGJ)combined with bone marrow mesenchymal stem cells(BMSCs)on Ras homolog gene family,member A(RhoA)/Rho-associated coiled-coil containing protein kinase 1(ROCK1)signaling pathway in the treatment of liver fibrosis.Methods:Seventy-two SD rats were randomly assigned into a normal group,a model group,a colchicine group,high-,medium,and low-dose YGJ(YGJ-H,YGJ-M,and YGJ-L,respectively)groups,a BMSCs group,and a YGJ+BMSCs group.The rat model of liver fibrosis was established by intraperitoneal injection of carbon tetrachloride(CCl_(4))for 6 weeks,and the rats were sacrificed after 4 weeks of treatment.Before the rats were sacrificed,we scored their behavior of yin deficiency.We then measured the content of glutamic-pyruvic transaminase(GPT)and glutamic-oxaloacetic transaminase(GOT)in the serum and stained the liver with hematoxylin-eosin(HE)and Masson to observe the pathological changes.RT-PCR and Western blotting were respectively employed to determine the mRNA and protein levels of RhoA,ROCK1,α-smooth muscle actin(α-SMA),and collagen type I(COL-1).Immunohistochemistry(IHC)was adopted to detectα-SMA and COL-1 positive areas in liver tissue sections.Results:Compared with the normal group,the modeling of liver fibrosis elevated the behavior scores of yin deficiency,caused liver inflammation and fibroplasia,up-regulated the mRNA and protein levels of RhoA,ROCK1,α-SMA,and COL-1(P<0.05),and increased the positive areas ofα-SMA and COL-1(P<0.05).Compared with the model group,YGJ-H,YGJ-M,YGJ-L,BMSCs,and YGJ+BMSCs relieved the behaviors of yin deficiency,liver inflammation,and fibrosis(P<0.05),down-regulated the mRNA and protein levels of RhoA,ROCK1,α-SMA,and COL-1(P<0.05),and decreased the positive areas ofα-SMA and COL-1(P<0.05).Among different treatment groups,YGJ+BMSCs group demonstrated the best performance(P<0.05).Conclusion:YGJ and BMSCs can inhibit the activation of hepatic stellate cells by inhibiting RhoA/ROCK1 signaling pathway to treat liver fibrosis,and their

关 键 词：一贯煎 肝纤维化 骨髓干细胞 冻干粉 大鼠 阴虚 行为学 RhoA/ROCK1信号通路 

分 类 号：R285[医药卫生—中药学]