川陈皮素经AKT/GSK3β/NF-κB通路抑制Kupffer细胞活化减轻大鼠原位肝移植后缺血再灌注损伤  

Nobiletin alleviates OLT-induced liver ischemia-reperfusion by inhibiting activation of Kupffer cell through AKT/GSK3β/NF-κB pathway in rats

作　　者：王敬元 李生伟(指导)[1] 吴涯昆 WANG Jing-Yuan;LI Sheng-Wei;WU Ya-Kun(Department of Hepatobiliary Surgery,Second Affiliated Hospital of Chongqing Medical University,Chongqing 400010,China)

机构地区：[1]重庆医科大学附属第二医院肝胆外科,重庆400010

出　　处：《中国免疫学杂志》2021年第9期1047-1052,共6页Chinese Journal of Immunology

基　　金：宜宾市科学技术局重点科技项目(No.17PJ112)。

摘　　要：目的:探讨川陈皮素(nobiletin)对大鼠原位肝移植后缺血再灌注损伤(IRI)的作用及机制。方法:提取大鼠Kupffer细胞(KCs),分为空白对照组、LPS组、LPS+川陈皮素组,ELISA检测炎症因子IL-1β、TNF-α和IL-6浓度,Western blot检测AKT/GSK3β/NF-κB通路蛋白的表达。动物实验中构建大鼠肝移植后缺血再灌注模型,分为假手术组(Sham组)、肝移植组(LT组)、LT+川陈皮素组。分别于再灌注3 h、6 h、12 h、24 h后获取各组外周血并检测血清转氨酶水平,ELISA检测血清炎症因子水平,获取肝脏组织,提取肝组织及KCs总蛋白,检测炎症因子、凋亡蛋白、AKT/GSK3β/NF-κB通路蛋白水平,HE染色检测肝脏组织病理改变,TUNEL检测肝细胞凋亡情况,流式检测各组CD68和CD86阳性KCs比例。结果:川陈皮素抑制LPS激活的KCs促炎因子IL-1β、TNF-α和IL-6的表达(P<0.05),促进抑炎因子IL-10的表达(P<0.05),同时川陈皮素抑制AKT/GSK3β/NF-κB通路磷酸化水平(P<0.05),动物实验中川陈皮素抑制AKT/GSK3β/NF-κB通路活性,LT+川陈皮素组肝脏损伤程度明显低于LT组(P<0.05),且LT+川陈皮素组肝细胞凋亡水平及M1型KCs比例均低于LT组(P<0.05)。结论:川陈皮素通过AKT/GSK3β/NF-κB通路抑制KCs炎症水平,减轻大鼠肝移植后肝脏缺血再灌注损伤。Objective:To investigate the effect and mechanisms of nobiletin on ischemia-reperfusion injury(IRI)after orthotopic liver transplantation(OLT)in rats.Methods:Kupffer cells(KCs)isolated from normal rats were divided into control group,LPS group,LPS+nobiletin,ELISA assay was used to detect the concentrations of IL-1β,TNF-αand IL-6.Western blot was used to detect the protein levels of AKT/GSK3β/NF-κB pathway.The animals were divided into sham group,LT group,LT+nobiletin group.After3,6,12,24 h of reperfusion,the peripheral blood of each group was obtained for the measurement of serum ALT,AST,IL-1β,TNF-αand IL-6.Total protein of liver tissue and KCs was extracted for the measurement of protein levels of inflammatory factors,apoptotic protein,and AKT/GSK3β/NF-κB pathway,HE staining was used to detect liver tissue pathological changes and TUNEL assay was used to detect hepatocyte apoptosis,and the proportion of M1 type KCs was detected by flow cytometry.Results:Nobiletin inhibited the expression of pro-inflammatory factors IL-1β,TNF-αand IL-6,but promoted the expression of IL-10,and inhibited the activation of AKT/GSK3β/NF-κB pathway in KCs that stimulated by LPS(P<0.05).Nobiletin also inhibited AKT/GSK3β/NF-κB pathway activity in animal experiments.The degree of liver injury in the LT+nobiletin group was significantly lower than that in the LT group(P<0.05),what’s more,the level of hepatocellular apoptosis and the proportion of M1 type KCs in the LT+nobiletin group were lower than those in the LT group(P<0.05).Conclusion:Nobiletin inhibits the level of inflammation KCs by down regulating AKT/GSK3β/NF-κB pathway,which contributes to alleviate OLT-induced ischemia reperfusion injury in rats.

关 键 词：川陈皮素 肝移植 缺血再灌注损伤 KUPFFER细胞 

分 类 号：R617[医药卫生—外科学]