摘要
目的研究二甲双胍(Met)对伴刀豆球蛋白A(ConA)所致急性爆发型肝炎小鼠的保护作用并探索其机制。方法C57BL/6小鼠随机分为正常对照组(NC组)、ConA组、Met组,每组8只。后两组分别灌饲0.2 mL生理盐水和100 mg/kg Met连续5 d后,尾静脉注射0.1 mL ConA(25 mg/kg)建立小鼠急性爆发型肝炎模型,18 h后处死。全自动生化分析仪检测小鼠血清丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)及总胆红素(TB)水平;HE染色观察小鼠肝组织病理改变;免疫组织化学染色法检测肝组织巨噬细胞浸润;实时定量PCR检测肝脏组织白细胞介素1β(IL-1β)、IL-6及肿瘤坏死因子α(TNF-α)的mRNA表达;血细胞分析仪检测小鼠外周血白细胞(WBC)总数及其中淋巴细胞数量,流式细胞术检测脾细胞中Th17细胞比例;免疫荧光组织化学染色检测肝组织胱天蛋白酶3(caspase-3)的表达;Western blot法检测肝组织腺苷酸活化蛋白激酶(AMPK)和磷酸化的AMPK蛋白表达。结果与ConA组相比,Met组小鼠血清ALT、AST及TB明显下降,肝病变减轻,巨噬细胞浸润减少,外周血白细胞总数及其中淋巴细胞数量增多,脾脏中Th17淋巴细胞比例下调,IL-1β、IL-6及TNF-α的水平降低,凋亡下降,磷酸化的AMPK的表达升高。结论Met通过激活AMPK,降低Th17细胞比例,抑制肝脏炎细胞浸润和炎细胞因子产生,减轻ConA诱导的肝损伤。
Objective To investigate the protective effect of metformin(Met)on acute fulminant hepatitis induced by concanavalin A(ConA)in mice and explore its mechanism.Methods Twenty-four mice were randomly divided into normal group(NC),ConA group,and Met group,with 8 mice in each group.The latter two groups respectively were gavaged with 0.2 mL normal saline and metformin(100 mg/kg)for 5 days,followed by tail vein injection of 0.1 mL ConA(25 mg/kg)to establish the acute fulminant hepatitis model,and all the mice were sacrificed 18 hours later.The serum levels of alanine aminotransferase(ALT),aspartate aminotransferase(AST)and total bilirubin(TB)were detected;the pathological changes of mouse liver tissue were observed with HE staining;the macrophage infiltration in liver tissue was detected with immunohistochemistry.The mRNA of IL-1β,IL-6 and TNF-αin liver tissue were tested with real time quantitative PCR.The number of total white blood cells(WBC)and lymphocytes in the peripheral blood were recorded and the frequency of Th17 cells in the spleen was detected by flow cytometry.The expression of apoptosis protein caspase-3 in liver tissue was observed with immunofluorescence histochemistry and the expression of AMPK and phosphorylated AMPK(p-AMPK)were detected by Western blot analysis.Results Compared with the ConA group,the Met group showed significantly decreased serum ALT,AST and TB,improved liver tissue pathology,decreased macrophage infiltration and increased content of peripheral totalWBCs and lymphocytes,as well as decreased frequency of Th17 lymphocytes in the spleen.The expression of IL-1β,IL-6 and TNF-αand apoptosis also decreased in this group,along with the increased expression of p-AMPK.Conclusion Met has a significant protective effect on acute fulminant hepatitis mice,and its mechanism may be related to the activation of AMPK signal,thus reducing the frequency of Th17 lymphocytes and alleviating the infiltration of hepatic inflammatory cells and the production of pro-inflammatory cytokines.
作者
刘青青
田海霞
杨昊
康行
刘海霞
杨晓丹
罗旭光
樊卫平
LIU Qingqing;TIAN Haixia;YANG Hao;KANG Xing;LIU Haixia;YANG Xiaodan;LUO Xuguang;FAN Weiping(Department of Microbiology and Immunology,Shanxi Medical University,Taiyuan 030001;Linfen Central Hospital,Linfen 041000,China)
出处
《细胞与分子免疫学杂志》
CAS
CSCD
北大核心
2022年第4期302-307,共6页
Chinese Journal of Cellular and Molecular Immunology
基金
山西省省筹资金资助回国留学人员科研项目(2020-079)。
