MLCK通路在重症急性胰腺炎相关肠道屏障损伤中作用及清胰汤的干预

Intestinal mechanical barrier damage in the process of the onset of SIRS and MODS occurred on the early stage for severe acute pancreatitis (SAP) played an action, but its mechanism is unclear. Qingyi decoction is an effective prescription for the clinical treatment of SAP. Dendritic cells (DCs) play a key role in stimulating intestinal mucosal immunity and its mechanism is a hot research topic. Preliminary experiments show that composed of tight junction protein Occludin1, ZO1 closely connection between the intestinal mucosa epithelial cells structure directly involved in intestinal mechanical barrier is damaged, in the intestinal tissue of SAP rats myosin light chain expression increased significantly, and and tight junction protein Occludin1, ZO1 expression change negative correlationand. Based on the above, we get the hypothesis that MLCK-Occludin1/ZO1 pathway involved in the regulation of intestinal mechanical barrier damage. The research project proposed by rat model and invitro cell experiment, application of laser scanning confocal microscope, the liquid phase chips, and other methods, in order to clarify this pathway in the regulation of intestinal mechanical barrier damage mechanism and the intervention mechanism of qingyi decoction and dendritic cells, to explore the pathogenesis and treatment of SAP associated MODS, to provide new way of thinking.

肠道机械屏障损伤在重症急性胰腺炎（SAP）早期诱发SIRS和MODS的发病过程中起到了始动作用，但其机制尚不明确。通里攻下中药清胰汤是临床治疗SAP的有效方剂，树突细胞（DCs）可以吞噬并呈递肠道内外来性抗原，在激发肠道黏膜免疫中起关键作用，其作用机制是研究热点。申请人前期研究发现由紧密连接蛋白Occludin1、ZO1等构成的肠黏膜上皮细胞间紧密连接结构受损直接参与肠道机械屏障损伤,预实验显示在SAP大鼠肠组织中肌球蛋白轻链激酶（MLCK）表达明显升高，且与紧密连接蛋白Occludin1、ZO1表达变化负相关。据此提出MLCK-Occludin1/ ZO1通路参与肠道机械屏障损伤调控的科研假说。本课题拟以大鼠动物模型与体外细胞学实验，应用激光扫描共聚焦显微镜、液相芯片等方法，调控MLCK通路对肠机械屏障损伤的影响，以期阐明该通路调控肠道机械屏障损伤机制及清胰汤和树突细胞干预机制。

通过动物实验观察重症急性胰腺炎（severe acute pancreatitis，SAP）模型大鼠肠组织中紧密连接蛋白Occludin-1、ZO-1和分泌型磷脂酶A2 (secreted phospholipaseA2，sPLA2)表达的变化规律，来探讨重症急性胰腺炎时，因上皮细胞凋亡和紧密连接断裂引起肠道机械屏障损伤的发生发展与分泌型磷脂酶A2、紧密连接蛋白Occludin-1、ZO-1的关系。通过动物实验和细胞实验分析肌球蛋白轻链激酶（myosin light chain kinase，MLCK）对紧密连接蛋白的调控通路在肠黏膜机械屏障中的作用，从而进一步探究重症急性胰腺炎时肠道屏障功能损伤的发病机理，探讨中医通里攻下理论在重症急性胰腺炎时肠道屏障功能损伤发病机制中的作用，同时观察祖国传统中药清胰汤（Qingyitang）、临床常规抗炎药物地塞米松（Dexamethasone）、MLCK抑制剂ML-7和树突细胞（dendritic cells，DC）对肠黏膜上皮细胞凋亡、紧密连接结构变化、炎症因子表达水平的影响，探讨中药清胰汤对重症急性胰腺炎相关肠道机械屏障损伤的保护作用，为临床工作中有效提高重症急性胰腺炎及其肠道损伤的治疗效果提供理论指导和实验依据。