内源性obestatin在高甘油三酯血症相关急性胰腺炎中对IKKβ/NF-κB信号通路的作用及其机制研究

For the past few years, the incidence of hypertriglyceridemia-associated pancreatitis(HTGP) had increased. The severity of HTGP is severer than the other causes, and it is more easily to recur. Recently, scientist found a new protein named obestatin involved in the fat metabolism and inflammatory reaction. We measured the level of obestatin in patients with acute pancreatitis. We found that the level of obestatin in patients with HTGP is lower than the other causes and it is negative correlation with the level of proinflammatory cytokine, such as IL-1β and TNF-α. Since the role of obestatin involved in HTGP is unclear, we designed this project for providing new method treating HTGP. Our project included clinical experiment, in vivo and in vitro: ①To collect serum of patients with acute pancreatitis and observe the relationship of obestatin concentration and the severity of patients. ②To establish the HTGP model with rats and inhibit the expression of obestatin with monoclonal antibody. After inhibiting the obestatin, the activity of and pathology injury of HTGP rats was detected. ③To establish the transfection pancreas acinar cell lines(AR42J) of obestatin gene silience and over-expression and investigate the activity of IKKβ/NF-κB pathway with different obestatin expression level in AR42J cell cultured in hypertriglyceridemic medium.

近年来高甘油三酯性胰腺炎（HTGP）发病率增高，有病情重、易复发的临床特点，其发病机制未明。Obestatin是一种新发现与调节脂代谢、抑制炎症反应有关的蛋白质。本课题组前期实验发现obestatin含量在HTGP患者较低，且与促炎细胞因子浓度、AP患者病情的严重程度呈负相关。为探讨obestatin在HTGP中的作用机制，并为治疗HTGP提供新途径，本项目拟通过临床、动物、细胞三方面进行研究：①检测AP病人血清obestatin浓度，了解obestatin在AP患者病程中的动态变化及其与疾病严重程度的关系；②建立HTGP大鼠模型，使用obestatin单克隆抗体抑制内源性obestatin表达，观察对IKKβ/NF-κB信号通路和病理损伤的影响；③用基因转染和siRNA调控胰腺AR42J细胞的obestatin表达，观察对高甘油三酯培养液中AR42J细胞IKKβ/NF-κB信号通路的影响

目前认为，obestatin参与调节脂代谢、抑制炎症反应，可能在高甘油三脂血症相关性急性胰腺炎（HTGP）中发挥作用。本研究发现：（1）obestatin在高甘油三酯血症性急性胰腺炎患者入院第一天的血清中的含量较其他病因的明显降低，但obestatin含量在AP患者和正常健康人之间无明显差异，在不同严重程度的AP中亦无明显差异。（2）利用高脂饲料法建立大鼠高甘油三酯血症模型，后以腹腔注射雨蛙肽法诱发急性胰腺炎，成功模拟人高甘油三酯血症性急性胰腺炎发病过程，采用干预黏膜屏障活性的药物ML-7进行干预，发现大鼠胰腺病理损伤较普通胰腺炎大鼠明显加重，MLCK和钙离子通道蛋白表达增加，JNK信号通路明显活化。（3）在大鼠胰腺肿瘤细胞株AR42J中调节obestatin的表达并置于高甘油三酯培养液中培养后发现，对细胞内IKKβ/NF-κB信号通路的活化受到影响。因此，通过本研究，我们认为：（1）高甘油三酯血症引起的急性胰腺炎患者出现中度以上的AP比率较高，起病早期血清中的obestatin含量可能与高甘油三酯血症性急性胰腺炎发病有关。（2）HTGP大鼠胰腺组织中MLCK的表达可能与胰腺损伤严重程度有关；其中MLCK可能通过调节TJ和钙通道蛋白的表达参与疾病的进展。（3）AR42J细胞在高PA浓度中生存受到一定抑制；obestatin的表达在高PA环境中可能调控IKKβ/NF-κB信号通路的活化。