腹腔穿刺引流对重症急性胰腺炎大鼠胰腺细胞凋亡的影响及机制  

Effect of APD on pancreatic apoptosis in rats with severe acute pancreatitis

作　　者：罗晨 黄启林 杨屹 原小惠 孙红玉 汤礼军 Luo Chen;Huang Qi-Lin;Yang Yi;Yuan Xiao-Hui;Sun Hong-Yu;Tang Li-Jun(Department of Hepatobiliary Surgery,Affiliated Hospital of Southwest Medical University,Luzhou,Sichuan 646000,China;Department of General Surgery&Pancreatic Injury and Repair Key Laboratory of Sichuan Province,General Hospital of Western Theater Command,Chengdu 610083,China)

机构地区：[1]西南医科大学附属医院肝胆外科,四川泸州646000 [2]西部战区总医院全军普通外科中心,四川省胰腺损伤与修复重点实验室,成都610083

出　　处：《解放军医学杂志》2020年第9期897-903,共7页Medical Journal of Chinese People's Liberation Army

基　　金：国家自然科学基金面上项目(81772001);国家临床重点专科项目(41732113)。

摘　　要：目的探讨腹腔穿刺引流(APD)对重症急性胰腺炎(SAP)大鼠胰腺细胞凋亡的影响,并探讨其可能的分子机制。方法将成年雄性SD大鼠随机分为假手术(SO)组、SAP组、APD组,每组18只。SAP组、APD组大鼠经胰胆管逆行微量泵入5%牛磺胆酸钠制备SAP模型,APD组进一步由右下腹导入一胃管进行引流。每组设置6、12、24 h时间点,收集腹主动脉血和胰腺组织。酶联免疫吸附法(ELISA)检测血清淀粉酶、炎性因子及内毒素的变化;HE染色观察胰腺组织损伤情况;原位末端转移酶标记法(TUNEL)检测胰腺组织细胞凋亡情况;Western blotting和免疫组化法检测凋亡相关蛋白及磷脂酰肌醇3-激酶/蛋白激酶B(PI3K/AKT)信号通路蛋白的表达。结果APD组大鼠胰腺组织坏死、水肿情况明显好于SAP组,病理学评分下降(P<0.05)。APD组大鼠血清淀粉酶、肿瘤坏死因子α(TNF-α)、白介素(IL)1β、IL-6及内毒素水平明显低于SAP组(P<0.05),胰腺细胞凋亡数量明显高于SAP组(P<0.05),胰腺组织凋亡蛋白裂解的半胱氨酸天冬氨酸蛋白酶-3(cleaved-caspase-3)和Bax表达水平明显高于SAP组(P<0.05),抗凋亡蛋白B细胞淋巴瘤-2(Bcl-2)表达水平明显低于SAP组(P<0.05)。相较于SAP组,APD组大鼠胰腺组织PI3K/AKT信号通路关键分子p-PI3K、p-AKT和核因子-κB(NF-κB)p65表达水平在各时间点均明显下降(P<0.05)。结论APD通过增强胰腺细胞凋亡减轻SAP的严重程度,其作用可能与抑制PI3K/AKT信号通路有关。Objective To investigate the effect of abdominal paracentesis drainage(APD)on pancreatic cell apoptosis in severe acute pancreatitis.Methods Male adult SD rats were randomized into the sham operation(SO)group,SAP group,and APD group,with 18 rats in each group.In the SAP group,5%sodium sulfonate was pumped into the retrograde pancreatic bile duct to prepare the SAP model.On this basis,a gastric tube was introduced into the right lower abdomen for drainage,namely the APD group.Blood from the abdominal aorta and pancreatic tissues were collected at 6,12,and 24 h time points in each group.The changes of serum amylase,inflammatory factor,and endotoxin were detected by ELISA.The HE staining was used to evaluate the pancreatic tissue injury.The apoptosis of pancreatic tissue was detected by TUNEL.Western blot and immunohistochemistry were used to detect the expression of apoptosis-related proteins and PI3K/AKT signaling pathway.Results Pancreatic tissue necrosis and edema were significantly lower in the APD group than in the SAP group,and the pathological score was decreased(P<0.05).Serum amylase,TNF-α,IL-1β,IL-6,and endotoxin levels in the APD group were significantly lower than those in the SAP group(P<0.05).The number of pancreatic cell apoptosis in the APD group was significantly higher than that in the SAP group(P<0.05),and the expression levels of pancreatic apoptotic proteins cleaved-caspase-3 and Bax were significantly increased in the APD group,while the expression levels of anti-apoptotic protein Bcl-2 were significantly decreased(P<0.05).Compared with the SAP group,the expression levels of PI3K/AKT signaling pathway key molecules p-PI3K,p-AKT,and NF-kB p65 were significantly decreased in the APD group(P<0.05).Conclusions Our data indicate that APD attenuates the severity of SAP by enhancing cell apoptosis via suppressing PI3K/AKT signaling pathway.This study provides a new theoretical basis for the treatment of severe acute pancreatitis with APD technology.

关 键 词：腹腔穿刺引流 急性重症胰腺炎 细胞凋亡 cleaved-caspase-3 PI3K/AKT信号通路 

分 类 号：R576[医药卫生—消化系统]