茯苓酸调节JAK2/STAT3/SOCS3信号通路对四氯化碳诱导的大鼠肝纤维化的影响

Effect of pachymic acid on CCl4-induced liver fibrosis in rats by regulating JAK2/STAT3/SOCS3 signal pathway

目的探讨茯苓酸(PA)调节JAK2/信号转导与转录激活因子3(STAT3)/细胞因子信号转导抑制因子3(SOCS3)信号通路对四氯化碳(CCl4)诱导的大鼠肝纤维化的影响。方法随机选取10只SD大鼠设为对照组(NC组),其余43只大鼠采用CCl4灌胃法(2 mL/kg)构建肝纤维化模型。造模后随机选取3只大鼠验证造模成功。将其余40只肝纤维化大鼠随机分为肝纤维化组、PA组、香豆霉素A1(C-A1)组、PA+C-A1组,每组均10只大鼠。各组均每日干预1次,连续7 d。采用ELISA法检测各组的血清炎症因子、肝功能标志物及肝纤维化标志物水平。分别采用H-E和Masson染色法观察各组肝组织病理形态变化和肝纤维化情况。采用蛋白质印迹法检测各组肝组织中JAK2/STAT3/SOCS3信号通路相关蛋白、α-平滑肌肌动蛋白(α-SMA)和Ⅲ型胶原(COL3)蛋白表达水平。结果NC组大鼠肝脏结构正常,染色红润,胶原沉积较少,未见胶原纤维增生,肝小叶结构和肝窦均无明显病变,肝窦无扩张或充血,门静脉区无炎症细胞浸润。与NC组相比,肝纤维化组的肝小叶结构紊乱,肝细胞脂肪变性坏死,炎症细胞浸润,肝脏肿胀,表面有许多小结节,胶原容积分数、IL-2、IL-6、IL-17A、ALT、AST、碱性磷酸酶(ALP)、透明质酸(HA)、层粘连蛋白(LN)、Ⅲ型前胶原(PCⅢ)、Ⅳ型胶原(Ⅳ-C)水平,以及α-SMA、COL3、磷酸化JAK2(p-JAK2)/JAK2、p-STAT3/STAT3、SOCS3蛋白表达水平均显著升高(P均<0.05)。与肝纤维化组相比,PA组的肝损伤和肝纤维化情况均有改善,并且上述指标水平均显著降低(P均<0.05),而C-A1组则均显著升高(P均<0.05)。与PA组相比,PA+C-A1组的肝细胞坏死、炎症细胞浸润情况和肝纤维化程度均加重,并且上述指标水平均显著升高(P均<0.05)。结论PA可能通过抑制JAK2/STAT3/SOCS3信号通路对大鼠肝纤维化起到改善作用。

Objective This paper aims to investigate the effect of pachymic acid(PA)on CCl4-induced liver fibrosis in rats by regulating the JAK2/signal transducer and activator of transcription 3(STAT3)/suppressor of cytokine signaling 3(SOCS3)signal pathway.Methods Ten SD rats were randomly selected and assigned to the control group(NC group),and the remaining 43 rats were given CCl4(2 mL/kg)by gavage to build the liver fibrosis model.After modeling,three rats were randomly selected to verify successful modeling.The remaining 40 rats with liver fibrosis were randomly assigned to the liver fibrosis group,the PA group,the coumermycin A1(C-A1)group,and the PA+C-A1 group,with 10 rats in each group.Each group was intervened once a day for 7 consecutive days.ELISA was used to detect the levels of serum inflammatory factors,liver function markers,and liver fibrosis markers while H-E and Masson staining were used to observe the pathological changes and liver fibrosis of liver tissue in each group.The protein expression levels of JAK2/STAT3/SOCS3 signal pathway related proteins,α-smooth muscle actin(α-SMA),and typeⅢcollagen(COL3)proteins in liver tissues of each group were detected using Western blotting.Results The NC group has normal cell structure,with red staining and less collagen deposition.No collagen fiber proliferation is observed,and there are no obvious lesions in the liver lobule structure and sinusoids.The sinusoids are not dilated or congested,and there is no inflammatory cell infiltration in the portal vein area.Compared with the NC group,the liver fibrosis group has disordered liver lobular structure,the liver cells are steatosis and necrosis,with inflammatory cell infiltration,liver swelling,and many small nodules on the surface.The collagen volume fraction,the levels of IL-2,IL-6,IL-17A,ALT,AST,alkaline phosphatase(ALP),hyaluronic acid(HA),laminin(LN),typeⅢprocollagen(PCⅢ),typeⅣcollagen(Ⅳ-C),and the expression levels ofα-SMA,COL3,p-JAK2/JAK2,p-STAT3/STAT3,and SOCS3 proteins are significantly increased(P<0.05).Compared with the liver fibrosis group,the PA group shows improvement in liver injury and fibrosis,and the levels of the above indicators are significantly reduced(P<0.05),while the C-A1 group shows significant increases(P<0.05).Compared with the PA group,the liver cell necrosis,inflammatory cell infiltration,and degree of liver fibrosis are aggravated in the PA+C-A1 group,and the levels of the above indicators are significantly increased(P<0.05).Conclusion PA may improve liver fibrosis in rats by inhibiting the JAK2/STAT3/SOCS3 signal pathway.