正肝方对二乙基亚硝胺诱导的肝癌大鼠的抗癌作用及机制:基于激活Hippo/YAP通路  

Zheng Gan Decoction inhibits diethylnitrosamine-induced hepatocellular carcinoma in rats by activating the Hippo/YAP signaling pathway

作　　者：宋添力 王一民 孙童 刘绪 黄胜 冉云 SONG Tianli;WANG Yimin;SUN Tong;LIU Xu;HUANG Sheng;RAN Yun(Department of Medicine,Hubei Minzu University,Enshi 445000,China;Hubei Provincial Key Laboratory of Rheumatic Disease Occurrence and Intervention,Enshi 445000,China;Department of Hepatology,Shenzhen Hospital(Longgang)of Beijing University of Chinese Medicine,Shenzhen 518172,China)

机构地区：[1]湖北民族大学医学部,湖北恩施445000 [2]风湿性疾病发生与干预湖北省重点实验室,湖北恩施445000 [3]北京中医药大学深圳医院(龙岗)肝病科,广东深圳518172

出　　处：《南方医科大学学报》2025年第4期799-809,共11页Journal of Southern Medical University

基　　金：国家自然科学基金地区基金(32160141);湖北省自然科学基金(2022CFB173);湖北省教育厅科研项目(B2022101);深圳市科技计划项目(JCYJ20220530172804010)。

摘　　要：目的探讨正肝方通过调控Hippo/YES-相关蛋白同源癌蛋白(YAP)信号通路对二乙基亚硝胺(DEN)诱导的肝癌大鼠模型的抗癌作用及对肝癌潜在的抑制机制。方法将80只SD大鼠随机分为正常对照组(n=10)、造模组(n=70)。造模组通过连续12周腹腔注射DEN(50 mg/kg)建立肝癌模型。建模成功后,随机分为5组:模型对照组、阳性对照组(槐耳颗粒,4 g/kg)、正肝方低、中、高剂量组(2、4、8 g/kg),n=10。各组接受相应药物干预,1次/d,持续17周。正常对照组和模型组仅灌喂等量纯水。观察大鼠生存状况和生存率,并测定体质量、肝脏指数、脾脏指数和胸腺指数。观测大鼠肝脏组织的形态变化。通过HE染色观察组织病理变化。利用免疫组化技术检测组织中YAP及p-YAP的表达水平。Western blotting检测组织中Hippo/YAP通路中YAP、p-YAP、MST1、LATS1和p-LATS1等相关蛋白表达。结果与正常对照组相比,模型对照组大鼠生存状况较差,生存率降低,体质量、肝脏指数、脾脏指数升高(P<0.01),胸腺指数降低(P<0.05)。正肝方治疗后,肝脏指数、脾脏指数降低(P<0.05),胸腺指数升高(P<0.01)。大鼠肝脏组织形态及HE染色结果表明,模型组肝小叶结构受损,肝细胞排列紊乱,正肝方治疗后这些病理变化得到改善。免疫组化结果显示,与模型对照组相比,在正肝方低、中、高剂量组及阳性对照组中,YAP的阳性表达水平逐渐减弱,且与正肝方的剂量呈负相关(P<0.01);而p-YAP的阳性表达则逐渐增强(P<0.01),且与正肝方的剂量呈正相关(P<0.01)。Western blotting结果显示,与模型对照组相比,在接受正肝方治疗的各剂量组中,随着药物浓度的增加,YAP蛋白表达水平随着药物剂量的增加而降低(P<0.001),p-YAP的阳性表达水平逐渐升高,尤其在高剂量组中这一变化更为明显(P<0.001);正肝方各剂量组及槐耳颗粒组治疗组的组织中MST1蛋白表达明显升高(P<0.05)。同时,正�Objective To investigate the inhibitory effect of Zheng Gan Decoction(ZGF)on tumor progression in a rat model of diethylnitrosamine(DEN)-induced hepatocellular carcinoma(HCC)and explore the possible mechanism.Methods Seventy SD rats were subjected to regular intraperitoneal injections of DEN(50 mg/kg)for 12 weeks to induce HCC tumorigenesis,with another 10 rats receiving saline injections as the normal control.After successful modeling,the rats were randomized into 5 groups(n=10)for daily treatment with distilled water(model group),Huaier Granules(4 g/kg;positive control group),or ZGF at low,medium,and high doses(2,4,and 8 g/kg,respectively)via gavage for 17 weeks.Body weight changes of the rats were monitored,and after completion of the treatments,the rats were euthanized for measurement of liver,spleen and thymus indices and morphological and histopathological examinations of the liver tissues using HE staining.The expressions of YAP,p-YAP,MST1,LATS1 and p-LATS1 in the liver tissues were detected using immunohistochemistry and Western blotting.Results Compared with the normal control rats,the rat models with DEN-induced HCC exhibited much poorer general condition with a significantly reduced survival rate,increased body weight and liver and spleen indices,and a lowered thymus index.ZGF treatment obviously reduced liver and spleen indices,increased the thymus index,and improved pathologies of the liver tissues of the rat models.Immunohistochemistry and Western blotting showed a dose-dependent reduction of YAP expression and an increment of p-YAP expression in ZGF-treated rats,which also exhibited significantly upregulated hepatic expressions of MST1,LATS1 and p-LATS1.Conclusion ZGF inhibits DEN-induced HCC in rats by activating the Hippo/YAP pathway via upregulating MST1 and LATS1 expression,which promotes YAP phosphorylation and degradation to suppress proliferation and induce apoptosis of the tumor cells.

关 键 词：正肝方 肝癌 Hippo/YAP通路 作用机制 二乙基亚硝胺 

分 类 号：R285.5[医药卫生—中药学]