高剂量西罗莫司可能通过促进细胞自噬对老年小鼠肝脏缺血-再灌注损伤起保护作用  

High-dose sirolimus protects hepatic ischemia-reperfusion injury probably through promoting cellular autophagy in aged mice

作　　者：滕旭 丁凡[2,3] 陈文捷 许赤[2] 

机构地区：[1]深圳市龙岗区第六人民医院,518117 [2]中山大学附属第三医院肝脏外科暨肝移植中心 [3]广东省肝脏疾病重点实验室生物治疗中心

出　　处：《器官移植》2017年第5期349-354,共6页Organ Transplantation

基　　金：深圳市科技计划项目(JCYJ20150402112934552)

摘　　要：目的探讨高剂量西罗莫司(雷帕霉素)对老年小鼠肝脏缺血-再灌注损伤(HIRI)的保护作用及其作用机制。方法 20只C57BL/6系老年小鼠按随机数字表法随机分为4组:缺血-再灌注组(IRI组)、低剂量雷帕霉素预处理组(rpm组)、高剂量雷帕霉素预处理组(RPM组)、对照组(Sham组),每组5只。Sham组仅开腹、关腹;其余3组建立老年小鼠70%HIRI模型,缺血时间均为60 min;rpm组和RPM组分别在术前1 h腹腔注射雷帕霉素1 mg/kg和5 mg/kg。再灌注12 h后,采用苏木素-伊红(HE)染色检测各组小鼠肝组织病理学变化,并采用Suzuki评分法进行病理学评分;比较各组小鼠血清丙氨酸转氨酶(ALT)和天冬氨酸转氨酶(AST)水平、血清肿瘤坏死因子(TNF)-α和白细胞介素(IL)-10水平以及肝组织中细胞自噬相关蛋白LC3B-Ⅱ蛋白水平。结果小鼠肝组织HE染色示Sham组为正常肝组织;IRI组和rpm组肝细胞损伤严重,炎症细胞大量浸润;RPM组可见肝窦轻度淤血,炎症细胞轻度浸润。病理学评分示RPM组为5(4~6)分,均低于rpm组[7(5~8)分]及IRI组[8(7~10)分](Z=-2.554、-2.731,均为P<0.05)。各组小鼠术后肝功能结果显示,RPM组AST[(691±207)U/L]显著低于IRI组[(2 032±575)U/L]、rpm组[(1 817±777)U/L](t=4.90、3.13,均为P<0.05);RPM组ALT[(996±584)U/L]均显著低于IRI组[(2 992±992)U/L]、rpm组[(2 373±687)U/L](t=3.86、3.41,均为P<0.05),而IRI组和rpm组的AST、ALT比较,差异均无统计学意义(均为P>0.05)。各组小鼠血清中的TNF-α和IL-10水平比较差异均无统计学意义(均为P>0.05)。Western-blot法显示RPM组肝组织LC3B-Ⅱ蛋白的相对表达量明显高于Sham组、IRI组、rpm组(均为P<0.05)。结论高剂量雷帕霉素可能通过促进细胞自噬对老年小鼠HIRI起保护作用。Objective To investigate the effect and mechanism of high-dose sirolimus （rapamycin） upon protecting the hepatic ischemia-reperfusion injury （HIRI） in aged mice. Methods Twenty C57BL/6 aged mice were randomly and evenly divided into the ischemia-reperfusion injury group （IRI group）, low-dose rapamycin pretreatment group （rpm group）, high-dose rapamycin pretreatment group （RPM group） and control group （Sham group） using the random number table method （ 5 mice in each group）. In the Sham group, abdominal cavity was incised and sutured alone. In the other three groups, aged mouse 70% HIRI models were established. The ischemia time was 60 min. At preoperative 1 h, rapamycin at a dose of 1 mg/kg and 5 mg/kg was administered via intraperitoneal injection in the rpm and RPM groups. At 12 h post- reperfusion, hematoxylin-eosin （HE） staining was performed to observe the histological changes in the mouse liver. Suzuki grading method was adopted to evaluate the pathological score. The serum levels of alanine aminotransferase （ALT） and aspartate aminotransferase （AST）, tumor necrosis factor （TNF）-α and interleukin （IL）-10 and the LC3B-Ⅱ protein level in the liver tissues were quantitatively measured and statistically compared among different groups. Results HE staining of the liver tissues revealed normal liver tissues in the Sham group, severe liver cellular injury accompanied with a large quantity of inflammatory cellular infiltration in the IRI and rpm groups. Mild sinusoidal congestion and slight inflammatory cellular infiltration were observed in the RPM group. The pathological score was 5 （4-6） in the RPM group, significantly lower than 7 （5-8） and 8 （7-10） in the rpm and IRI groups （Z=-2.554 and -2.731, both P〈0.05）. In terms of postoperative liver function parameters, the AST level was （691±207） U/L in the RPM group, significantly lower compared with （2 032 ± 575） U/L and （1 817 ± 777） U/L in the IRI and rpm groups （t=-4.90 and 3.13

关 键 词：缺血-再灌注损伤 肝移植 小鼠 老年 西罗莫司(雷帕霉素) 细胞自噬 丙氨酸转氨酶 天冬氨酸转氨酶 肿瘤坏死因子 白细胞介素 LC3B-Ⅱ蛋白 

分 类 号：R617[医药卫生—外科学] R329.2[医药卫生—临床医学]