秦皮素通过调节JAK1/STAT3信号通路抑制肝纤维化

Anti-fibrotic effect of fraxetin on liver fibrosis in rats by targeting the JAK1/STAT3 signaling pathway

目的探讨秦皮素对四氯化碳(CCl4)诱导的大鼠肝纤维化的防治作用及其机制。方法将48只成年雄性SD大鼠随机均分为4组,即正常对照组、模型组、秦皮素低剂量组和秦皮素高剂量组。除正常对照组外,其余3组大鼠给予50%CCl4花生油混合液(2 mL·kg-1)灌胃,每周2次,建立肝纤维化模型,持续8周。给药组造模的同时每日给予秦皮素干预,剂量分别为25 mg·kg-1和50 mg·kg-1。正常对照组给予等量的生理盐水。8周后处死各组大鼠,称量体重和肝重;全自动生化分析仪检测血清谷丙转氨酶(ALT)、谷草转氨酶(AST)和总胆红素(TBIL);放射性免疫法测定血清透明质酸(HA)、层黏蛋白(LN)、Ⅳ型胶原(CⅣ)和Ⅲ型前胶原肽(PⅢNP)。HE和Masson染色考察肝组织病理学改变,免疫组织化学染色检测肝组织α-SMA的表达;Western blot检测肝脏组织中α-SMA和JAK1/STAT3信号通路关键蛋白的表达水平;Q-PCR检测肝窦毛细血管化调控因子(VEGF、β-FGF和PD-ECGF)基因的表达水平。结果与模型组相比,秦皮素给药组中肝纤维化大鼠肝功能指标(ALT、AST、TBIL)和纤维化指标(HA、LN、CⅣ、PⅢNP)血清水平明显减低。HE和Masson染色结果显示秦皮素能显著改善肝纤维化大鼠肝脏组织病理学结构和胶原沉积程度。与此同时,秦皮素能够抑制肝星状细胞激活标志物(α-SMA)以及肝窦毛细血管化调控因子(VEGF、β-FGF、PD-ECGF)的表达。与模型组相比,秦皮素给药组大鼠肝脏中p-JAk1和p-STAT3蛋白的表达水平显著降低。结论秦皮素可改善大鼠肝纤维化,其机制可能与抑制JAK1/STAT3信号通路有关。

Objective To determine the anti-fibrotic effect and potential mechanism of fraxetin on liver fibrosis induced by CCl4 in rats. Methods Totally 48 male Sprague-Dawley(SD) rats were randomly divided into a normal group, a model group, a fraxetin low-dose group(25 mg·kg^-1) and a fraxetin high-dose group(50 mg·kg^-1).Except for the normal group liver fibrosis model was established in other groups by intragastric administration with the mixture solution(CCl4∶peanut oil = 1∶1) twice a week for 8 weeks. Synchronously, the rats in the 2 treatment groups were given fraxtin 25 mg·kg^-1 and 50 mg·kg^-1 respectively by intragastric administration once daily. As a control, the rats in the normal group were given the same volume of saline. After 8 weeks, all rats were sacrificed.The body and liver were weighted, and then the liver function(ALT, AST, and TBIL) and hepatic fibrosis markers(HA, LN, CⅣ, and PⅢNP) were detected by the automatic biochemistry analyzer and radioimmunoassay. HE and Masson’s trichrome staining were performed to evaluate the changes of morphology and fibration in liver tissue. Furthermore, immunohistochemical staining was employed to visualize the expression of α-SMA. Western blot was used to determine the expression of α-SMA and the key signaling proteins in the JAK1/STAT3 pathway.Q-PCR was used to detect the mRNA levels of regulatory factors of hepatic sinus capillary vascularization. Results Compared with the model group, the serological levels of the liver function(ALT, AST, and TBIL) and hepatic fibrosis markers(HA, LN, CⅣ, and PⅢNP) were significantly dereased in the fraxetin treatment groups. HE and Masson’s trichrome staining reconfirmed that fraxetin alleviated the liver histopathological structure and collagen deposition in the liver fibrotic rats. Concomitantly, the expression of HSCs activation makers(α-SMA) and hepatic sinus capillary vascularization factors(VEGF,β-FGF, and PD-ECGF) were significantly inhibited in the 2 fraxetin treatment goups. Moreover, compared with the model group, fraxetin significantly inhibited the expression of p-JAK1 and p-STAT3 proteins. Conclusion Fraxetin can alleviate liver fibrosis induced by CCl4 in rats and the mechanism may be closely related to inhibiting the JAK1/STAT3 signaling pathway.