急性胰腺炎大鼠肠黏膜屏障损伤、MLCK/p-MLC信号通路活性变化及其与抗菌肽的关系  

Intestinal mucosal barrier damage and activity change of MLCK/p-MLC2 signaling pathway in rats with acute pancreatitis and their relationships with antimicrobial peptides

作　　者：马凤雨 曾家月 文琳 陈霞 MA Fengyu;ZENG Jiayue;WEN Lin;CHEN Xia(Department of Gastroenterology,The Affiliated Hospital of Southwest Medical University,Luzhou 646000,China;不详)

机构地区：[1]西南医科大学附属医院消化内科,四川泸州646000 [2]成都医学院第一附属医院消化内科

出　　处：《山东医药》2023年第2期28-32,共5页Shandong Medical Journal

基　　金：国家自然科学基金项目青年科学基金(81600420);四川省泸州市人民政府—西南医科大学科技战略合作项目(2019LZXNYDJ48)。

摘　　要：目的观察急性胰腺炎(AP)大鼠肠黏膜屏障损伤情况和MLCK/p-MLC2信号通路活性变化,探讨MLCK/p-MLC2信号通路激活与抗菌肽的关系。方法将24只SD大鼠分为对照组6只和AP组18只,采用胰胆管逆行注射5%牛磺胆酸钠溶液进行AP造模,对照组仅翻动胰腺后复位。AP组于造模后6、12、24 h各取6只大鼠进行解剖和取材,对照组于造模后6 h取材。采用ELISA法检测血清淀粉酶、脂肪酶水平,HE染色法观察回肠组织病理改变,RT-qPCR法检测回肠组织潘氏细胞分泌的抗菌肽REG3γ、α-防御素5、溶菌酶mRNA表达水平,Western blotting法检测回肠组织紧密连接蛋白ZO-1、Claudin-1及MLCK/p-MLC2信号通路相关蛋白MLCK、p-MLC2蛋白表达水平。结果AP组造模后6、12、24 h血清淀粉酶、脂肪酶水平均高于对照组,其中12 h和24 h血清淀粉酶、脂肪酶水平高于6 h,且24 h高于12 h(P均<0.05)。对照组回肠黏膜完整,绒毛结构规整正常;AP组回肠黏膜出现不同程度的断裂缺失、间质水肿、炎性细胞浸润。AP组造模后6、12、24 h回肠组织病理学评分均高于对照组,且24 h高于6 h和12 h,12 h高于6 h(P均<0.05)。AP组造模后6、12、24 h回肠组织REG3γ、α-防御素5、溶菌酶mRNA及ZO-1、Claudin-1蛋白表达水平均低于对照组,且24 h低于6 h和12 h,12 h低于6 h(P均<0.05),造模后6、12、24 h回肠组织MLCK、p-MLC2蛋白表达水平均高于对照组,且12、24 h高于6 h,24 h高于12 h(P均<0.05)。AP组回肠组织MLCK蛋白表达水平与REG3γ、α-防御素5、溶菌酶mRNA水平呈负相关(r分别为-0.96、-0.95、-0.93,P均<0.05),p-MLC2蛋白表达水平与REG3γ、α-防御素5、溶菌酶mRNA水平亦呈负相关(r分别为-0.96、-0.94、-0.96,P均<0.05)。结论AP大鼠存在肠黏膜屏障损伤,MLCK/p-MLC2信号通路激活参与了AP肠黏膜屏障损伤的过程,该作用可能与潘氏细胞分泌抗菌肽减少相关。Objective To observe the intestinal barrier injury and activity change of MLCK/p-MLC2 signaling pathway in acute pancreatitis(AP)rats,and to explore their relationships with antimicrobial peptides.Methods Twentyfour SD rats were divided into the control group(n=6)and AP group(n=18),and 5%taurocholic acid was retrogradely injected into the pancreaticobiliary duct to establish the rat model of AP.For the rats in the control group,we only turned the pancreases and then reset them.In the AP group,6 samples each were collected at 6,12 and 24 h after modeling,respectively.The rats were selected in the control group at 6 h after modeling.ELISA was performed to detect the serum levels of amylase and lipase,HE staining was performed to observe the ileal pathology,RT-qPCR was performed to detect the mRNA expression levels of REG3γ,α-defencin 5 and lysozyme,and Western blotting was performed to detect the protein expression levels of ZO-1,Claudin-1,MLCK/p-MLC2 signaling pathway-related MLCK,p-MLC2.Results Compared with the control group,the levels of serum amylase and lipase were elevated in the AP group at 6,12 and 24 h after modeling,and those at 12 and 24 h were higher than those at 6 h,and those at 24 h were higher than those at 12 h(all P<0.05).In the control group,the ileum mucosa was intact and the villi structure was regular.In the AP group,ileal mucosa showed different degrees of fracture deletion,interstitial edema and inflammatory cell infiltration.The ileal histopathological scores of the AP group at 6,12 and 24 h after modeling were higher than those of the control group,and those at 24h were higher than those at 6 and 12 h,and those at 12 h were higher than those at 6 h(all P<0.05).The expression levels of REG3γ,α-defensin 5,lysozyme mRNA and ZO-1 and Claudin-1 protein in ileum of the AP group at 6,12 and 24 h after molding were lower than those of the control group,and those at 24 h were lower than those at 6 and 12 h,and those at 12 h were lower than those at 6 h(all P<0.05).The protein expression levels

关 键 词：MLCK/p-MLC2信号通路 抗菌肽 紧密连接蛋白 肠黏膜屏障损伤 急性胰腺炎 

分 类 号：R576[医药卫生—消化系统]