摘要
目的研究脂联素(APN)介导APPL1/AMPK信号通路对小鼠肾脏急性缺血再灌注损伤后期纤维化的影响。方法雄性C57BL/6小鼠48只,随机分为假手术组(Sham组)、肾脏急性缺血再灌注组(IRI组)、肾脏急性缺血再灌注+APN组(APN/IRI组)和肾脏急性缺血再灌注+生理盐水组(NS/IRI组),每组各12只。APN/IRI组在术前15 min、术后每24 h分别经腹腔注射APN蛋白球状片段5μg/g,共3次;NS/IRI组在相应时点经腹腔注射等量生理盐水。IRI组均行双侧夹闭肾蒂30 min后开放肾灌注,Sham组分离肾蒂但不夹闭。各组大鼠分别在术后2 d随机各取6只小鼠,留取血液及肾组织标本。PAS染色观察肾组织病理变化;全自动生化分析仪检测血尿素氮(BUN)及血肌酐(Scr)含量;Western blot检测肾脏组织APPL1蛋白表达和AMPK磷酸化水平。术后14 d每组取剩余6只小鼠肾脏标本,天狼星红苦味酸(Sirus red)染色观察肾组织病理变化,real time PCR检测α平滑肌肌动蛋白(α-SMA)和转化生长因子β_1(TGF-β_1)的mRNA表达。结果术后2 d,与Sham组相比,IRI组、APN/IRI组和NS/IRI组小鼠肾组织小管间质损害加重,血BUN、Scr升高(P<0.05),APPL1蛋白表达和AMPK磷酸化水平明显增高(P<0.05);与IRI组相比,APN/IRI组小鼠肾小管损害明显减轻,血BUN、Scr明显降低(P<0.05),APPL1蛋白表达和AMPK磷酸化水平明显增高(P<0.05)。术后14 d,与Sham组相比,IRI组、APN/IRI组和NS/IRI组小鼠肾组织纤维化程度明显增高,α-SMA和TGF-β_1的mRNA的表达明显增多(P<0.05);与IRI组相比,APN/IRI组小鼠肾组织纤维化程度明显降低,α-SMA和TGF-β_1的mRNA的表达量明显减少(P<0.05)。结论 APN介导APPL1/AMPK信号通路减轻小鼠肾脏急性缺血再灌注损伤后期纤维化。
Objective To investigate the attenuation by APN on late fibrosis induced by acute ischemia reperfusion injury in mice via APPL1 / AMPK signaling pathway. Methods Forty- eight male C57BL /6 mice were randomly divided into sham operation group（ Group Sham）,acute ischemia- reperfusion group（ Group IRI）,acute ischemia- reperfusion plus APN group（ Group APN / IRI） and acute ischemia- reperfusion plus NS group（ Group NS / IRI）. The g APN（ 5 μg/g） and physiological saline were injected intraperitoneally 15 min before operation and every 24 h after operation in Group APN / IRI and Group NS / IRI,respectively. Renal acute ischemia reperfusion group were performed open renal perfusion after bilateral renal pedicle clip for 30 min was performed in Group IRI,as separation of renal pedicle without clip was performed in Group Sham. On the 2-（nd）day after the operation,6 mice were randomly selected from each groups were sacrificed for blood and kidney samples. The renal histopathological changes were observed through Schiff periodic acid staining（ PAS）. The contents of BUN and Scr were determined. The APPL1 protein expression and the phosphorylate level of AMPK were detected by Western blot. On the 14thday after the operation,6 mice from each group were sacrificed for kidney samples. The α smooth muscle actin（ α- SMA） and transforming growth factor beta 1（ TGF- β1） mRNA were determined by real- time PCR. The degree of kidney fibrosis was observed through Sirus red staining. Results Compared with Group Sham,tubular interstitial damage were significantly aggravated and the levels of BUN and Scr were increased in Group IRI,APN / IRI group and NS / IRI group on the 2-（nd）day after the operation（ P〈0. 05）,so were the increases APPL1 protein and the phosphorylate level of AMPK（ P〈0. 05）. Compared with Group IRI,tubular interstitial damage were significantly attenuated and the levels of BUN and Scr were significantly reduced in Group APN / IRI group on the 2-（nd）day after the
出处
《广东医学》
CAS
北大核心
2016年第16期2384-2388,共5页
Guangdong Medical Journal
基金
广东省科技计划项目(编号:2014A020212612)
