猪骨髓间充质干细胞抑制脂多糖诱导炎症反应改善猪胰岛细胞功能损伤  

Bone marrow mesenchymal stem cells from pigs inhibit inflammatory response induced by lipopolysaccharide and improve islet cell dysfunction in pigs

作　　者：袁宝红[1] 赵维山[1] 王若天[1] 管傲然[1] 王珏[1] 李汝红[1] Yuan Baohong;Zhao Weishan;Wang Ruotian;Guan Aoran;Wang Yu;Li Ruhong(First Department of General Surgery,Yan’an Hospital of Kunming City(Affiliated Yan’an Hospital of Kunming Medical University),Kunming 650051,Yunnan Province,China)

机构地区：[1]昆明市延安医院(昆明医科大学附属延安医院)普通外科一科,云南省昆明市650051

出　　处：《中国组织工程研究》2021年第13期1969-1975,共7页Chinese Journal of Tissue Engineering Research

基　　金：云南省科技厅-昆明医科大学联合专项[2019FE001(-099)],项目负责人:袁宝红

摘　　要：背景:猪胰岛移植是治疗糖尿病的常用手段,但炎症反应和氧化应激会导致猪胰岛移植的远期效果欠佳。猪骨髓间充质干细胞已被证实与胰岛联合移植可改善移植物的功能。但关于猪骨髓间充质干细胞通过调节胰岛细胞中miR-299-5p的表达来调控胰岛β细胞功能和存活率还尚未被报道。目的:探讨猪骨髓间充质干细胞通过miR-299-5p/SIAH1分子轴调控JNK/C-Jun信号通路对猪胰岛细胞功能损伤的影响。方法:用脂多糖诱导猪胰岛细胞功能损伤,然后与骨髓间充质干细胞共培养24 h,采用ELISA检测猪胰岛细胞中白细胞介素6、白细胞介素1β、肿瘤坏死因子α、活性氧、超氧化物歧化酶及胰岛素水平,Hoechst 33258染色观察猪胰岛细胞凋亡情况,Annexin V-FITC/PI检测猪胰岛细胞凋亡率,Western blot检测SIAH1和JNK/C-Jun信号通路相关蛋白表达;qPCR检测猪胰岛细胞中与胰岛细胞功能损伤有关的miRNAs表达;双荧光素酶报告基因验证miR-299-5p和SIAH1的靶向关系。结果与结论:①猪骨髓间充质干细胞抑制脂多糖引起的白细胞介素6、白细胞介素1β、肿瘤坏死因子α、活性氧上调和促凋亡作用,同时也抑制脂多糖引起的超氧化物歧化酶、胰岛素分泌量下调作用;②猪骨髓间充质干细胞显著上调miR-299-5p表达,miR-299-5p可通过靶向下调SIAH1表达抑制JNK/C-Jun信号通路激活;③结果表明,猪骨髓间充质干细胞通过调控miR-299-5p/SIAH1分子轴抑制JNK/C-Jun信号通路激活,抑制脂多糖诱导的炎症反应、氧化应激和猪胰岛细胞凋亡,并促进胰岛素分泌,从而改善猪胰岛细胞功能。BACKGROUND:Porcine pancreatic islet transplantation is a common treatment for diabetes mellitus,but inflammatory reaction and oxidative stress can lead to poor long-term effect.It has been confirmed that transplantation of porcine bone marrow mesenchymal stem cells with islets can improve graft function.However,it has not been reported that porcine bone marrow mesenchymal stem cells regulate the expression of miR-299-5p in islet cells to regulate the function and survival rate of isletβcells.OBJECTIVE:To investigate the effects of bone marrow mesenchymal stem cells in pigs on functional impairment of islet in pig by regulating the JNK/C-Jun pathway through the miR-299-5p/SIAH1 molecular axis.METHODS:Lipopolysaccharide was used to induce functional impairment of porcine islet cells,which were co-cultured with bone marrow mesenchymal stem cells for 24 hours.Interleukin-6,interleukin-1β,tumor necrosis factor-α,reactive oxygen species,superoxide dismutase and insulin levels in islet cells were detected by ELISA.Hoechst 33258 staining was used to observe apoptosis.Annexin V-FITC/PI was used to detect the apoptosis rate of porcine islet cells.Western blotting was used to detect the expression of SIAH1 and proteins associated with JNK/C-Jun pathway.The expression of miRNAs in porcine islet cells was detected by qPCR.The dual-luciferase reporter gene assay verified the targeting relationship between miR-299-5p and SIAH1.RESULTS AND CONCLUSION:(1)Bone marrow mesenchymal stem cells in pigs inhibited the up-regulation of interleukin-6,interleukin-1β,tumor necrosis factor-α,reactive oxygen species and pro-apoptotic effects,and also inhibited the down-regulation of superoxide dismutase and insulin secretion induced by lipopolysaccharide.(2)Bone marrow mesenchymal stem cells of pigs significantly upregulated miR-299-5p expression.miR-299-5p inhibited activation of the JNK/C-Jun pathway by down-regulating SIAH1 expression.(3)Results indicate that bone marrow mesenchymal stem cells of pigs inhibit the activation of the JNK/

关 键 词：干细胞 骨髓间充质干细胞 胰岛细胞 脂多糖 炎症 氧化应激 猪 实验 

分 类 号：R459.9[医药卫生—治疗学] R394.2[医药卫生—临床医学]