目的 基于Janus激酶1(Janus kinase 1,JAK1)和转录激活因子3(signal transducer and activator of transcription 3,STAT3)信号通路研究高车前素对四氯化碳(carbon tetrachloride,CCl₄)诱导肝纤维化(hepatic fibrosis,HF)小鼠的影响。方法 小鼠连续6周背部sc CCl₄溶液制备HF模型,将HF小鼠随机分为模型组、秋水仙碱(2 mg/kg)组和高车前素高、低剂量(30、10 mg/kg)组,各给药组连续4周ip相应药物,末次给药后24 h制备血清,计算肝脏指数;采用全自动生化分析仪测定血清中丙氨酸氨基转移酶(alanine aminotransferase,ALT)、天冬氨酸氨基转移酶(aspartate aminotransferase,AST)活性及总胆红素(total bilirubin,TBIL)水平;测定血清和肝组织中羟脯氨酸(hydroxyproline,Hyp)水平;测定肝组织中丙二醛(malondialdehyde,MDA)、还原型谷胱甘肽(glutathione,GSH)水平及超氧化物歧化酶(superoxide dismutase,SOD)活性;检测血清中透明质酸(hyaluronic acid,HA)、层黏连蛋白(laminin,LN)、III型前胶原(type III procollagen,PIIINP)、IV型胶原(type IV collagen,Col-IV)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-1β(interleukin-1β,IL-1β)和IL-6水平;采用苏木素-伊红(HE)染色观察肝组织病理改变;采用qRT-PCR法检测肝组织中尿激酶型纤溶酶原激活物(urokinase type plasminogen activator,uPA)和纤溶酶原激活物抑制因子-1(plasminogen activator inhibitor-1,PAI-1)mRNA表达;采用Western blotting检测肝组织中JAK1/STAT3信号通路中相关蛋白表达。结果 与模型组比较,高车前素组小鼠体质量显著增加(P<0.05、0.01),肝脏指数显著降低(P<0.05、0.01);血清中ALT、AST活性和TBIL水平显著降低(P<0.05、0.01);血清和肝组织中Hyp水平显著降低(P<0.05、0.01);肝组织中MDA水平显著降低(P<0.01),SOD活性显著升高(P<0.05、0.01);血清中LN、HA、PIIINP、Col-IV、TNF-α、IL-1β和IL-6水平均显著降低(P<0.05、0.01);光镜下可见肝组织细胞水肿、空泡样改变、炎细胞浸润明显减轻;肝组织中uPA mRNA表达水平显著升高(P<0.01),PAI-1 mRNA表达水平显著降低(P<0.01);肝组织中p-JAK1和p-STAT3蛋白表达水平显著降低(P<0.001)。结论 高车前素可减轻HF小鼠的肝损伤和炎症程度,其抗HF的作用机制可能与激活uPA纤溶酶系统和干预JAK1/STAT3信号转导通路有关。

Objective To study the effect of hispidulin on carbon tetrachloride (CCl₄)-induced hepatic fibrosis (HF) mice based on Janus kinase 1(JAK1) and signal transducer and activator of transcription 3(STAT3) signaling pathway.Methods HF model was prepared by sc CCl₄ solution on the back of mice for six consecutive weeks,HF mice were randomly divided into model group,colchicine (2 mg/kg) group and hispidulin high-and low-dose (30,10 mg/kg) groups,each administration group was ip corresponding drug for four consecutive weeks,serum was prepared at 24 h after the last administration,and liver index was calculated;Fully automated biochemical analyzer was used to measure alanine aminotransferase (ALT),aspartate aminotransferase (AST) activities and total bilirubin (TBIL) level in serum;Levels of hydroxyproline (Hyp) in serum and liver tissues were detected;Malondialdehyde (MDA),glutathione (GSH) levels and superoxide dismutase (SOD) activity in liver tissues were detected;Hyaluronic acid (HA),laminin (LN),type III procollagen (PIIINP),type IV collagen (Col-IV),tumor necrosis factor-α(TNF-α),interleukin-1β(IL-1β) and IL-6 levels in serum were detected;Hematoxylin-eosin (HE) staining was used to observe pathological changes in liver tissues;qRT-PCR was used to detect urokinase type plasminogen activator (uPA) and plasminogen activator inhibitor-1(PAI-1) mRNA expressions in liver tissues;Western blotting was used to detect the expressions of related proteins in JAK1/STAT3 signaling pathway in liver tissues.Results Compared with model group,body weight of mice in hispidulin group was significantly increased (P<0.05,0.01),liver index was significantly decreased (P<0.05,0.01),ALT,AST activities and TBIL level in serum were significantly decreased (P<0.05,0.01),Hyp levels in serum and liver tissue were significantly decreased (P<0.05,0.01),MDA level in liver tissue was significantly decreased (P<0.01),while SOD activity was significantly increased (P<0.05,0.01),levels of LN,HA,PIIINP,Col-IV,TNF-α,IL-1β and IL-6 in serum were significantly decreased (P<0.05,0.01);Hepatic tissue edema,vacuolar-like changes,and inflammatory cell infiltration were observed under light microscope;uPA mRNA expression in liver tissue was significantly increased (P<0.01),andPAI-1 mRNA expression was significantly decreased (P<0.01);p-JAK1 and p-STAT3 protein expression levels in liver tissue were significantly decreased (P<0.001).Conclusion Hispidulin can reduce liver injury and inflammation in HF mice,and its anti-HF mechanism may be related to the activation of uPA plasmin system and intervention of JAK1/STAT3 signal transduction pathway.

R285.5

重庆市自然科学基金面上项目(cstc2019jcyj-msxmX0044)