亚精胺促进线粒体自噬缓解急性胰腺炎的分子机制研究  

Molecular mechanism of spermidine promoting mitochondrial autophagy in alleviating acute pancreatitis

作　　者：巴穆登 阿布力克木·吾拉音 吴朝阳[1] BA Mu-den;ABULIKMU Wulayin;WU Chao-yang(Department of Emergency Trauma Surgery,People’s Hospital of Xinjiang Uygur Autonomous Region,Urumqi 830001,China)

机构地区：[1]新疆维吾尔自治区人民医院急诊创伤外科,新疆乌鲁木齐830001

出　　处：《现代药物与临床》2023年第7期1575-1583,共9页Drugs & Clinic

基　　金：新疆维吾尔自治区自然科学基金资助项目(2022D01C107)。

摘　　要：目的探讨亚精胺促进线粒体自噬缓解急性胰腺炎的疗效及作用机制。方法将C57BL/6J小鼠随机分为对照组、模型组、亚精胺组、亚精胺+环孢菌素A组,每组5只。通过每小时向小鼠腹膜内注射1次50μg/kg的蛙皮素构建急性胰腺炎模型。对照组按照同模型组的处理方式注射等体积无菌生理盐水;亚精胺组腹膜内注射100 mmol的亚精胺;亚精胺+环孢菌素A组在给予亚精胺治疗的同时,腹膜内注射10 mg/kg的环孢菌素A。苏木精–伊红(HE)染色评估小鼠胰腺组织损伤程度。TUNEL染色评估胰腺腺泡细胞凋亡情况。ELISA法测定小鼠血清炎症因子白细胞介素-6(IL-6)、肿瘤细胞坏死因子-α(TNF-α)、白细胞介素-1β(IL-1β)、C反应蛋白(CRP)的水平。免疫组织化学染色(IHC)评估髓过氧化物酶(MPO)水平。JC-1染料测定胰腺组织提取的总线粒体的线粒体膜电势水平。Western blotting法测定胰腺组织细胞质基质型LC3(LC-3Ⅰ)、微管相关蛋白1轻链3Ⅱ(LC-3Ⅱ)、泛素结合蛋白(p62)、NOD样受体热蛋白结构域相关蛋白3(NLRP3)、半胱氨酸天冬氨酸蛋白酶(Caspase-1)、cleaved-Caspase-1、IL-1β、PTEN诱导假定激酶1(PINK1)、磷酸化Parkin(p-Parkin)和Parkin的表达水平。结果与模型组相比,亚精胺组胰腺组织几乎不存在中性粒细胞的大量浸润,腺泡细胞肿胀程度明显减弱,基本无腺泡细胞坏死;TUNEL染色阳性细胞数和MPO IHC阳性染色细胞数明显降低;小鼠血清IL-6、TNF-α、IL-1β、CRP的水平明显降低;胰腺组织LC-3Ⅰ/Ⅱ、NLRP3、IL-1β、cleaved-Caspase-1、PINK1和p-Parkin的表达水平明显降低,p62的表达水平明显增强;胰腺组织总线粒体膜电势水平明显增强(P<0.05)。与亚精胺组相比,亚精胺+环孢菌素A组完全逆转了亚精胺的治疗效果。结论亚精胺可以促进线粒体自噬缓解急性胰腺炎。Objective To explore the effect and mechanism of spermidine promoting mitochondrial autophagy in alleviating acute pancreatitis.Methods C57BL/6J mice were randomly divided into control group,model group,spermidine group,and spermidine+cyclosporin A group,with 5 mice in each group.Acute pancreatitis model was established by intraperitoneal injection of 50μg/kg of frogskin into mice every hour.Control group was injected with equal volume sterile saline according to the treatment method of the same model group.The spermidine group was given intraperitoneal injection dose of 100 mmol spermidine,Spermidine+cyclosporine A group was given 10 mg/kg cyclosporine A by intraperitoneal injection at the same time of spermidine treatment.Hematoxylin-eosin(HE)staining was used to evaluate the degree of pancreatic tissue injury in mice.The apoptosis of pancreatic acinar cells was evaluated by TUNEL staining.Serum levels of inflammatory factor IL-6,TNF-α,IL-1β,and CRP were determined by ELISA.Myeloperoxidase(MPO)levels were evaluated by immunohistochemical staining(IHC).The level of mitochondrial membrane potential of total mitochondria extracted from pancreatic tissue was determined by JC-1 dye.LC-3Ⅰ,LC-3Ⅱ,p62,NLRP3,Caspase-1,cleaved-Caspase-1,IL-1β,PINK1,p-Parkin,Parkin were determined by Western blotting method.Results Compared with the model group,there was almost no large infiltration of neutrophils in the pancreatic tissue of spermidine group,the swelling degree of acinous cells was significantly reduced,and there was basically no acinous cell necrosis.The number of TUNEL positive cells and MPO IHC positive cells decreased significantly.Serum levels of IL-6,TNF-α,IL-1β,and CRP were significantly decreased.The expression levels of LC-3-Ⅰ/Ⅱ,NLRP3,IL-1β,cleaved Caspase-1,PINK1,and p-Parkin in pancreatic tissue were significantly decreased,and the expression level of p62 was significantly increased.The total mitochondrial membrane potential of pancreatic tissue was significantly increased(P<0.05).Compared with th

关 键 词：亚精胺 环孢菌素A 急性胰腺炎 巨自噬 线粒体自噬 白细胞介素-6 肿瘤细胞坏死因子-α 

分 类 号：R965[医药卫生—药理学] R979.5[医药卫生—药学]