摘要
目的:观察舒肝化癥方对四氯化碳(CCl4)诱导的肝纤维化模型大鼠的治疗作用,并探究其是否通过缺氧诱导因子-1α/血管内皮生长因子/转化生长因子-β1(HIF-1α/VEGF/TGF-β1)通路发挥作用。方法:将54只雄性SPF级SD大鼠按照随机原则分为6组:空白组、模型组、秋水仙碱组(0.2 mg·kg^(-1))、舒肝化癥方高、中、低剂量组(29.52、14.76、7.38 g·kg^(-1)),每组9只。造模过程为每周3次,持续8周。首次注射次日开始给药,药物干预为每日1次,持续8周。末次给药后当天,大鼠禁食禁水,次日处死大鼠,期间动态监测各组大鼠的生理状况。苏木素-伊红(HE)染色观察肝脏的病理学形态改变,酶联免疫吸附测定法(ELISA)检测肝组织中羟脯氨酸(HYP)和血管紧张素Ⅱ(AngⅡ)的含量,实时荧光定量聚合酶链式反应(Real-time PCR)测定肝组织中HIF-1α、VEGF和TGF-β1的mRNA表达水平,免疫组化法(IHC)和蛋白免疫印迹法(Western blot)检测肝脏组织中HIF-1α、VEGF和TGF-β1的蛋白表达水平。结果:与空白组比较,模型组大鼠整体状况明显下降;结缔组织明显增生,肝细胞间脂肪细胞明显增加;HYP、AngⅡ的含量升高;HIF-1α、VEGF和TGF-β1的mRNA和蛋白表达均出现不同程度地增高(P<0.05)。与模型组比较,秋水仙碱和舒肝化癥方组大鼠肝组织中结缔组织增生减少,炎性细胞浸润减少;HYP、AngⅡ的含量降低;HIF-1α、VEGF、TGF-β1的mRNA和蛋白表达水平均降低,其中以秋水仙碱和舒肝化癥方高剂量最为明显(P<0.05)。结论:舒肝化癥方对CCl4诱导的肝纤维化模型大鼠具有明显的治疗效果,其治疗机制可能与HIF-1α/VEGF/TGF-β1信号通路的调节有关,也与改善肝组织缺氧状况、血管重构和肝纤维化气虚血瘀证候有关。
Objective:To observe the therapeutic effect of Shugan Huazheng prescription on hepatic fibrosis model rats induced by carbon tetrachloride(CCl4)and explore whether it plays its role through hypoxiainduced factor-1α/vascular endothelial growth factor/transforming growth factor-β1(HIF-1α/VEGF/TGF-β1)pathway.Method:A total of 54 male SPF SD rats were randomly divided into six groups:blank group,model group,colchicine group(0.2 mg·kg^(-1)),and high-,medium-,and low-dose groups(29.52,14.76,and 7.38 g·kg^(-1))of Shugan Huazheng prescription,with nine rats in each group.The molding was conducted three times a week for eight weeks.Administration began the day after the first injection,and the drug intervention was once a day for eight weeks.On the day after the last administration,the rats were deprived of food and water,and they were killed the next day,during which the physiological status of each group of rats was dynamically monitored.The pathological changes in the liver were observed by hematoxylin-eosin(HE)staining,and the content of hydroxyproline(HYP)and angiotensinⅡ(AngⅡ)in liver tissue were detected by enzyme-related immunosorbent assay(ELISA).Real-time fluorescent quantitative PCR(Real-time PCR)was used to determine the mRNA expression levels of HIF-1α,VEGF,and TGF-β1 in liver tissue,and immunohistochemical method(IHC)and Western blot were used to detect the protein expression levels of HIF-1α,VEGF,and TGF-β1 in liver tissue.Result:Compared with the blank group,the overall condition of rats in the model group decreased significantly.The proliferation of connective tissue and the increase in adipose cells between hepatocytes were obvious.The content of HYP and Ang was increased.The mRNA and protein expressions of HIF-1α,VEGF,and TGF-β1 were increased to varying degrees(P<0.05).Compared with the model group,the proliferation of connective tissue and inflammatory cell infiltration in the liver tissue of colchicine and Shugan Huazheng prescription groups were reduced.The content of HYP and Ang was decreased.The mRNA and protein expression levels of HIF-1α,VEGF,and TGF-β1 were decreased,and the colchicine group and high-dose group of Shugan Huazheng prescription were the most significant(P<0.05).Conclusion:Shugan Huazheng prescription has an obvious therapeutic effect on CCl4-induced hepatic fibrosis model rats.Its therapeutic mechanism may be related to the regulation of the HIF-1α/VEGF/TGF-β1 signaling pathway and the improvement of hepatic hypoxia,vascular remodeling,and the syndrome of Qi deficiency and blood stasis in hepatic fibrosis.
作者
邢安丽
赵鲲鹏
张秋菊
李婕妠
陈世玉
郭嘉琪
张铭
XING Anli;ZHAO Kunpeng;ZHANG Qiuju;LI Jiena;CHEN Shiyu;GUO Jiaqi;ZHANG Ming(Clinical College of Traditional Chinese Medicine,Gansu University of Chinese Medicine,Lanzhou 730000,China)
出处
《中国实验方剂学杂志》
CAS
CSCD
北大核心
2024年第8期57-65,共9页
Chinese Journal of Experimental Traditional Medical Formulae
基金
国家自然科学基金项目(81660772)。
