Mir-21调控TIPE2-NFκB信号通路在肠缺血再灌注损伤中的作用及机制研究

Inflammatory cascade reaction is the critical step during the pathogenesis of multiple organ dysfunction syndrome (MODS) which induced by intestinal ischemia/reperfusion (II/R). Tumor necrosis factor-a induced protein-8-like 2 (TIPE2) plays a crucial role in regulating inflammation and apoptosis which may mediate via NFκB. Our previous studies indicated that NFκB is crucial in regulating inflammatory and apoptosis during II/R, while NFκB inhibition is effective to restrict intestinal injury. In addition, studies also showed following results: i) Up-regulation of TIPE2 inhibited NFκB level significantly and reduced the II/R injury; ii) miR-21 exhibits effect likes selective TIPE2 inhibitor/agonist through targeting and combining with TIPE2 coding region directly and stably; iii) miR-21 up-regulated in intestine after II/R, and were negatively correlated with TIPE2 expression. Based on these findings, we firstly hypothesis that regulating TIPE2-NFκB signaling pathway by miR-21 inhibition may be an effective strategy against II/R. To test this hypothesis, II/R models were established using TIPE2 gene knockout mice and the hypoxia/reoxygenation (H/R) models were created using Caco2 cells to imitate II/R in vivo. A variety of advanced molecular biological methods will be used; Following two questions will be clarified: Fistly, the effect and mechanism of TIPE2-NFκB pathway in II/R injury. In addition, we also focus on the therapeutic effect of targeting miR-21 suppression which regulates TIPE2-NFκB pathway against II/R injury.

炎症级联反应是肠缺血再灌注（II/R）引发多器官功能障碍综合征（MODS）的重要环节。肿瘤坏死因子α诱导蛋白8样分子2（TIPE2）是调节炎症反应及细胞凋亡的重要因子，可能通过NFκB发挥作用。我们研究证实，NFκB是II/R中重要的炎症及凋亡调节子。进一步研究发现：1）上调TIPE2可明显抑制NFκB及炎症反应，减轻II/R损伤；2）miR-21与TIPE2编码区直接并稳定结合发挥TIPE2选择性抑制剂/激动剂样作用；3）miR-21在II/R肠组织中下调且与TIPE2负相关。据此，我们假设下调miR-21调控TIPE2-NFκB通路或可有效防治II/R损伤。本研究利用TIPE2基因敲除鼠II/R损伤模型和肠上皮细胞缺氧/复氧模型，采用先进分子生物学方法，探讨TIPE2-NFκB通路在II/R损伤中的作用及机制；研究下调MiR-21靶向TIPE2-NFκB通路在II/R损伤中的防治策略。

炎症级联反应是肠缺血再灌注（I/R）引发多器官功能障碍综合征（MODS）的重要环节。肿瘤坏死因子α诱导蛋白8样分子2（TIPE2）是调节炎症反应及细胞凋亡的重要因子，可能调节炎症级联反应并在肠I/R引发MODS过程中发挥重要作用。本研究采用体内转染的小鼠及小肠上皮IEC-6细胞系及巨噬细胞系，采用多种分子生物学技术，通过体内和体外实验研究了TIPE2及其靶分子在肠I/R中的作用。研究表明：1、TIPE2及相关分子在肠I/R小鼠模型中发挥重要作用。诱导TIPE2表达负性调控NFκB及炎性细胞因子、氧化应激损伤，减少细胞凋亡，最终减轻小肠及肝肺损伤。TIPE2-/-的小鼠中该保护作用明显减弱。2、采用人工合成的 miR-21可以上调TIPE2表达并降低肠I/R损伤，而TIPE2-/-的小鼠中该保护作用明显减弱。同时，TIPE2干扰的小肠上皮细胞给予H/R刺激后，ROS水平、炎症反应与细胞凋亡水平则明显增高。3、天然植物来源的黄芪多糖可以通过上调TIPE2减轻小鼠肠I/R损伤。相同的现象在H/R刺激的巨噬细胞中也被发现。本研究提示：TIPE2可以作为防治肠I/R损伤的重要靶点。APO、Mir-21及黄芪多糖等TIPE2诱导剂可以保护肠IR多器官损伤。本研究进一步阐明了肠I/R多器官损伤的发生机制，并为防治药物的开发提供了依据。