选择性环氧合酶-2抑制剂通过Rho/ROCK通路对2型糖尿病合并非酒精性脂肪性肝炎大鼠肝脏保护作用机制的研究  

Mechanism study of the protective effects of selective cyclooxygenase-2 enzyme inhibitors on the liver of rats with type 2 diabetes mellitus combined with nonalcoholic steatohepatitis via Rho/ROCK pathway

作　　者：袁玲[1] 田丰[2] Yuan Ling;Tian Feng(Department of Gastroenterology,Qinghai Provincial Peoples's Hospital,Xining 810000,China;Department of Gastroenterology,Shengjing Hospital Affiliated to China Medical University,Shenyang 110004,China)

机构地区：[1]青海省人民医院消化内科,西宁810000 [2]中国医科大学附属盛京医院消化内科,沈阳110000

出　　处：《中华肝脏病杂志》2022年第1期74-80,共7页Chinese Journal of Hepatology

摘　　要：目的探讨选择性环氧合酶-2抑制剂塞来昔布是否通过抑制Rho/Rho相关蛋白激酶(ROCK)通路的表达对2型糖尿病(T2DM)合并非酒精性脂肪性肝炎(NASH)大鼠肝脏具有保护作用。方法雄性SD大鼠40只,随机等分为T2DM合并非酒精性脂肪性肝炎(T2DM-NASH)组、T2DM-NASH+塞来昔布组、对照组、对照+塞来昔布组四组,其中T2DM-NASH及T2DM-NASH+塞来昔布组予高糖高脂饲料喂养,对照组及对照+塞来昔布组予基础饲料(25 kJ/kg)喂养,4周后T2DM-NASH组及T2DM-NASH+塞来昔布组腹腔注射链脲佐菌素(STZ,30mg/kg)诱导T2DM模型,而对照组及对照+塞来昔布组腹腔注射等体积柠檬酸-柠檬酸钠缓冲液,STZ注射4周后T2DM-NASH+塞来昔布组及对照+塞来昔布组予塞来昔布(10 mg·kg^(-1)·d^(-1))溶于生理盐水中灌胃4周,剩余两组大鼠予等体积生理盐水灌胃4周。第12周末处死动物,取肝脏组织,行HE染色观察肝脏病理改变;免疫组织化学及蛋白质印迹法检测RhoA、Rho相关蛋白激酶1(ROCK1)及Rho相关蛋白激酶2(ROCK2)蛋白在肝脏的表达,实时定量PCR法检测RhoA、ROCK1及ROCK2 mRNA在肝脏的表达情况,并采用方差分析及t检验对比各组间蛋白及mRNA的表达差异。结果与对照组及对照组+塞来昔布组相比,T2DM-NASH组及T2DM-NASH+塞来昔布组光镜下肝组织呈重度脂肪变性,部分可见炎症细胞浸润,且肝组织RhoA、ROCK1及ROCK2的蛋白及mRNA表达水平均显著升高(P<0.05),而T2DM-NASH+塞来昔布组较T2DM-NASH组肝细胞脂肪变性有所减轻且肝组织RhoA、ROCK1及ROCK2蛋白及mRNA表达水平均降低(P<0.05)。结论选择性环氧合酶-2抑制剂塞来昔布对T2DM-NASH大鼠肝脏具有保护作用,而其作用可能是通过抑制Rho/ROCK通路的表达而达到的。Objective To investigate whether the selective cyclooxygenase-2 enzyme inhibitors celecoxib has protective effect on the liver of rats with type 2 diabetes mellitus(T2DM)combined with nonalcoholic steatohepatitis(NASH)via inhibiting the expression of Rho/ROCK pathway.Methods Forty male SD rats were randomly divided into four groups:type 2 diabetes mellitus combined with nonalcoholic steatohepatitis(T2DM-NASH)group,T2DM-NASH+celecoxib group,control group,and control+celecoxib group.The T2DM-NASH and T2DM-NASH+celecoxib groups were fed with high-sugar and fat diet,and the control group and control+celecoxib group were fed with basal diet(25 kJ/kg).Four weeks later,streptozotocin(STZ,30 mg/kg)was intraperitoneally injected into the NASH group and T2DM-NASH+celecoxib group to induce T2DM model,and the control group and control+celecoxib group were intraperitoneally injected with isovolumic citric acid-sodium citrate buffer.Four weeks after STZ injection,the T2DM-NASH+celecoxib group and the control+celecoxib group were gavaged with celecoxib(10 mg·kg·d)dissolved in nonrial saline for 4 weeks,and the remaining two groups of rats were gavaged with isovolumic normal saline for 4 weeks.Animals were sacrificed at the end of the 12-weeks,and the liver tissue was collected.Liver pathological changes were observed by HE staining.The expressions of RhoA,RhoA,ROCK1 and ROCK2 proteins in liver were detected by immunohistochemistry and western blot.The expressional condition of RhoA,ROCK1 and ROCK2 mRNA in liver were detected by real-time quantitative PCR.The differences were compared between protein and mRNA expression among the groups by analysis of variance and t-test.Results Compared with the control group and the control+celecoxib group,the liver tissue of the T2DM-NASH group and the T2DM-NASH+celecoxib group had severe steatosis,and there was partial inflammatory cell infiltration under the light microscope.The expression levels of RhoA,ROCK1 and ROCK2 protein and mRNA were significantly increased(P<0.05)in each liver ti

关 键 词：非酒精性脂肪性肝炎 2型糖尿病 选择性环氧合酶-2抑制剂 RHOA Rho相关蛋白激酶 

分 类 号：R587.2[医药卫生—内分泌] R575.5[医药卫生—内科学]