肝X受体负性调控枯否细胞中Toll样受体4通路减轻移植肝脏缺血再灌注损伤  

Liver X receptor ameliorate ischemia-reperfusion injury of transplanted livers in rats by down-regulating Toll-like receptor 4 signaling pathway in Kupffer cells

作　　者：刘双权 薛强[1] 成名翔[2] 夏丰[1] 龚建平[1] 涂兵[1] 

机构地区：[1]重庆医科大学附属第二医院肝胆外科,重庆400010 [2]重庆医科大学附属第一医院肝胆外科,重庆400016

出　　处：《中国新药与临床杂志》2014年第6期440-445,共6页Chinese Journal of New Drugs and Clinical Remedies

基　　金：重庆市卫生局医学科研项目(2011-20148)

摘　　要：目的探讨肝X受体(LXR)负性调控大鼠枯否细胞(KCs)中Toll样受体4(TLR4)通路对移植肝脏缺血再灌注损伤的影响。方法密度梯度离心法分离培养SD大鼠肝脏KCs,将获得的细胞随机分为脂多糖(LPS)组、LPS+GW3965组、LPS+LXR拮抗剂组。动物实验选用雄性SD大鼠96只,随机分为GW3965预处理组(GW3965 0.3 mg·kg-1于手术前30 min经尾静脉注射处理供体)、生理盐水对照组(给予等体积生理盐水)。两组均采用改进的Kamada两袖套法进行肝移植。Western blot测定各组细胞、移植物中白细胞介素-1受体相关激酶4(IRAK-4)、干扰素调节因子-3(IRF3)蛋白表达,EMSA测定各组细胞NF-κB的相对活性,酶联免疫吸附法检测KCs培养上清液肿瘤坏死因子α(TNF-α)、干扰素β(IFN-β),于移植肝脏再灌注后3、6、24 h测定血清丙氨酸转氨酶(ALT),光镜观察肝脏组织的形态学变化。结果 IRAK-4和IRF3蛋白表达、NF-κB相对活性、TNF-α和IFN-β水平在LPS+LXR拮抗剂组最高,而在LPS+GW3965组最低(均P<0.05)。GW3965预处理组各时间点组织的IRAK-4和IRF3蛋白表达量以及血清ALT活性明显低于对照组(P<0.05),GW3965预处理组各时间点肝组织的损伤均明显轻于对照组(P<0.05)。结论激活LXRs能负性调控KCs中TLR4信号通路的IRAK-4、IRF3的蛋白表达,抑制NF-κB的活化和炎症因子的释放,可减轻肝移植缺血再灌注损伤。AIM To investigate the protective effects of liver X receptor in ischemia-repeffusion injury of rat liver grafts by inhibiting Toll-like receptor 4 （TLR4） signaling in Kupffer cells （KCs）. METHODS KCs were isolated from male Sprague Dawley （SD） rats by density gradient centrifugation, and were cultured and divided into LPS treated group, LPS ＋ GW3965 （the LXR agonist） group and LPS ＋ LXR antagonist group. Ninety-six male SD rats were used as donors （n = 48） and recipients （n = 48） of liver transplantation. They were randomly assigned to GW3965 group （n = 24） with donor injected with GW3965 （0.3 mg·kg^-1） through the caudal vein thirty minutes before operation, control group （n = 24） with donor injected the same amount of saline. The protein expressions of the IRAK-4 and IRF3 in KCs and liver tissue were detected by Western blot, and the activity of NF- κB in KCs was determined by electrophoretic mobility shift assay （EMSA） . The concentration of TNF- α and IFN- β in supernatants were evaluated by enzymelinked immunosorbent assay （ELISA）. The serum levels of alanine aminotransferase （ALT） in each group were assessed with automatic biochemistry analyzer at 3, 6, 24 h after the transplantation. The morphological observation of hepatic tissue was made under light microscope. RESULTS The protein expression levels of IRAK-4 and IRF3, the activity of NF-κB and the concentration of TNF-α and IFN-β in the LPS ＋ GW3965 group were the lowest, as compared to the other two groups （P 〈 0.05） . At each time point, serum levels of ALT and the protein expression levels of IRAK-4 and IRF3 in the GW3965 group were significantly lower than those in the control group （P 〈 0.05）. And hepatic injury in the GW3965 group was significantly lighter than that in the control group. CONCLUSION Activating LXR can effectively downregulate the levels of IRAK- 4 and IRF3 and suppress activity of NF-κB, which inhibit inflammatory after liver transplantation. reaction and pal

关 键 词：肝X受体 枯否细胞 再灌注损伤 TOLL样受体4 

分 类 号：R617[医药卫生—外科学]