利胆合剂抑制钙敏感受体介导的肝细胞凋亡改善幼龄大鼠肝内胆汁淤积研究  

Study on Lidan Mixture improving intrahepatic cholestasis in young rats by inhibiting hepatocyte apoptosis-mediated by calcium-sensitive receptor

作　　者：丘燕燕 鄢素琪[3] 秦欢 周俪姗[2] 熊小丽[2] 江治霞[2] QIU Yan-yan;YAN Su-qi;QIN Huan;ZHOU Li-shan;XIONG Xiao-li;JIANG Zhi-xia(Hubei University of Chinese Medicine,Wuhan 430065,China;Wuhan Children's Hospital,Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430016,China)

机构地区：[1]湖北中医药大学,武汉430065 [2]华中科技大学同济医学院附属武汉儿童医院,武汉430016 [3]华中科技大学同济医学院附属武汉儿童医院中西医结合科

出　　处：《中华中医药杂志》2022年第4期2264-2269,共6页China Journal of Traditional Chinese Medicine and Pharmacy

基　　金：国家自然科学基金面上项目(No.81574024);湖北省卫计委2016-2017年度中医药中西医结合科研项目(No.2017HZ-Y35)。

摘　　要：目的:研究钙敏感受体(CaSR)在胆汁淤积幼龄大鼠模型中的表达及利胆合剂的干预作用。方法:将40只SD幼龄大鼠随机分为正常组、模型组、熊去氧胆酸(UDCA)组、利胆合剂组。正常组和模型组灌胃0.9%氯化钠溶液(10 mL·kg^(-1)·d^(-1))6 d,UDCA组和利胆合剂组分别灌胃UDCA(60 mg·kg^(-1)·d^(-1))和利胆合剂(40 g·kg^(-1)·d^(-1))6 d。除正常组外,其余各组在第5天给予α-异硫氰酸奈酯(ANIT)50 mg·kg^(-1)·d^(-1)灌胃1次建立肝内胆汁淤积模型。ANIT灌胃后48 h处死各组大鼠,采集标本。采用酶比色法检测TBIL、DBIL、TBA、ALP、γ-GT、ALT、AST;采用HE染色在光镜下观察各组肝脏组织病理学的变化;TUNEL检测肝组织的细胞凋亡情况;Western Blot检测肝组织CaSR及Bax、Bcl-2、Cytochrome-C、Cleaved caspase-3、Caspase-8、p-ERK、ERK、P38、p-P38、JNK、p-JNK表达。结果:模型组大鼠TBIL、DBIL、TBA、ALP、γ-GT、ALT、AST水平均较正常组显著升高(P<0.01);UDCA组、利胆合剂组此7项指标水平均较模型组显著降低(P<0.01),且利胆合剂组低于UDCA组(P<0.05,P<0.01)。UDCA组、利胆合剂组肝脏病理、肝细胞凋亡较模型组有明显改善,而利胆合剂组改善最为明显。模型组CaSR、Bax、Cytochrome-C、Cleaved caspase-3、Caspase-8、p-P38和p-JNK水平较正常组显著增加(P<0.01),Bcl-2和p-ERK水平显著降低(P<0.01);UDCA组、利胆合剂组CaSR、Bax、Cytochrome-C、Cleaved caspase-3、Caspase-8、p-P38和p-JNK水平较模型组显著降低(P<0.01),Bcl-2和p-ERK水平显著升高(P<0.01);利胆合剂组CaSR、Bax、Cleaved caspase-3、Caspase-8、p-JNK水平较UDCA组显著降低(P<0.01,P<0.05)。结论:利胆合剂可缓解ANIT诱导的胆汁淤积幼龄大鼠的胆汁淤积及肝损伤程度,保护肝细胞,疗效优于UDCA,其作用机制可能是通过线粒体途径和MAPK途径抑制CaSR介导的肝细胞凋亡。Objective:To investigate the expression of calcium-sensitive receptor(CaSR)in cholestasis young rat model and the intervention effect of Lidan Mixture(LDHJ).Methods:Forty SD young rats were randomly divided into normal group,model group,ursodeoxycholic acid(UDCA)group and LDHJ group.The normal group and model group were administrated with 0.9%sodium chloride solution(10 mL·kg^(-1)·d^(-1))by gavage for 6 days,UDCA group and LDHJ group were administrated with UDCA(60 mg·kg^(-1)·d^(-1))and LDHJ(40 g·kg^(-1)·d^(-1))by gavage for 6 days respectively.Except the normal group,other groups were administrated withα-isothiocyanate(ANIT,50 mg·kg^(-1)·d^(-1))by gavage one time on the fifth day to establish intrahepatic cholestasis model.48 hours after ANIT intragastric administration,rats in each group were sacrificed and specimens were collected.TBIL,DBIL,TBA,ALP,γ-GT,ALT and AST were measured by enzyme colorimetry.HE staining was used to observe the histopathological changes of liver in each group under light microscope.TUNEL was used to detect the apoptosis rate of liver tissue.Western Blot was used to detect CaSR,Bax,Bcl-2,Cytochrome-C,Cleaved caspase-3,Caspase-8,p-ERK,ERK,P38,p-P38,JNK,and p-JNK in liver tissues.Results:The levels of TBIL,DBIL,TBA,ALP,γ-GT,ALT,AST in model group were significantly higher than those in normal group(P<0.01);the levels of TBIL,DBIL,TBA,ALP,γ-GT,ALT and AST in UDCA group and LDHJ group were significantly lower than those in model group(P<0.01),and those in LDHJ group those in UDCA group(P<0.05,P<0.01).UDCA group and LDHJ group had significantly improve in liver pathology compared with that in the model group which has the most serious pathological,and LDHJ group showed the most significant improvement.The model group showed the most severe hepatocyte apoptosis,and the hepatocyte apoptosis in UDCA group and LDHJ group showed significant improvement compared with the model group,and the improvement in LDHJ group was the most obvious.Compared with the normal group,the levels of CaSR

关 键 词：利胆合剂 胆汁淤积 钙敏感受体 肝细胞凋亡 

分 类 号：R285.5[医药卫生—中药学]