基于JAK1/STAT6通路探讨安肠愈疡汤对HT-29细胞炎症模型的影响及机制研究

Discussion on the effect and mechanism research of Anchang Yuyang Recipe on HT-29 cell inflammation model based on the JAK1/STAT6 pathway

关键词:  溃疡性结肠炎  安肠愈疡汤  JAK1/STAT6通路  体外实验

Key words:  ulcerative colitis  Anchang Yuyang Recipe  JAK1/STAT6 pathway  in vitro experiment

摘要:

[目的] 观察安肠愈疡汤对脂多糖（LPS）诱导的HT-29细胞炎症模型蛋白酪氨酸激酶1（JAK1）/信号转导和转录活化因子6（STAT6）通路的影响，并探讨其作用机制。[方法] 实验分为6组，空白组、模型组、安肠愈疡汤低、中、高浓度组（0.1、1、10 mg/mL）、美沙拉秦组（1 mg/L），空白组不做任何处理，模型组给予LPS诱导，其余各组在LPS诱导后分别给予相应药物干预。干预24 h后分别应用酶联免疫吸附（ELISA）法、免疫荧光技术检测各组白细胞介素（IL）-13、肠三叶因子3（TFF3）的表达，分别应用实时荧光定量聚合酶链反应（Q-RT-PCR）技术、蛋白免疫印迹（Western Blot）法检测各组IL-13、JAK1、STAT6、TFF3 mRNA及蛋白的表达。[结果] 与空白组比较，模型组IL-8水平上调（P<0.01），IL-13水平下调（P<0.01），提示炎症模型构建成功，模型组JAK1、STAT6、TFF3的水平也不同程度降低（P<0.05或P<0.01）。与模型组比较，各用药组IL-13、TFF3分泌水平增加、荧光强度增强（P<0.05或P<0.01），IL-13、JAK1、STAT6、TFF3 mRNA及蛋白表达不同程度上调（P<0.05或P<0.01）。安肠愈疡汤高浓度组IL-13、TFF3分泌水平、荧光强度以及IL-13、JAK1、STAT6、TFF3 mRNA及蛋白表达不同程度高于中、低浓度组（P<0.05或P<0.01），而与美沙拉秦组比较无统计学差异（P>0.05）。[结论] 安肠愈疡汤可能通过上调IL-13的分泌，以激活JAK1/STAT6通路，促进黏膜修复因子TFF3的分泌，减轻LPS诱导的HT-29细胞的炎症反应，发挥保护作用。

Abstract:

[Objective] To observe the influence of JAK1/STAT6 pathway on LPS induced HT-29 cell with Anchang Yuyang Recipe (AYR) and to explore the mechanism.[Methods] The experiment was divided into 6 groups:control group,model group,low,medium and high concentration group of Anchang Yuyang Recipe (0.1,1,10 mg/mL),mesalazine group (1 mg/L). The control group was done without any treatment. The model group was induced by LPS. The remaining groups were given corresponding drug intervention after LPS induction. After 24 hours of intervention,the expressions of IL-13 and TFF3 in each group were detected by ELISA and immunofluorescence techniques. The expressions of IL-13,JAK1,STAT6,and TFF3 mRNA and protein in each group were detected by Q-RT-PCR and Western Blot.[Results] Compared with the control group,the IL-8 level in the model group was increased (P<0.01) and the IL-13 level was decreased (P<0.01).Itindicated that the inflammation model was successfully constructed. The levels of JAK1,STAT6 and TFF3 in the model group were also reduced (P<0.05 or P<0.01). Compared with the model group,the secretion levels and the fluorescence intensity of IL-13 and TFF3 increased in each medication group (P<0.05 or P<0.01). The expressions of IL-13,JAK1,STAT6,and TFF3 mRNA and protein were up-regulated (P<0.05 or P<0.01). The secretion levels,fluorescence intensity of IL-13,TFF3and the expression of IL-13,JAK1,STAT6and TFF3 mRNA and protein in the high-concentration group of AnchangYuyang Recipe were higher than those of the medium-concentration group and low-concentration group (P<0.05 or P<0.01). But there was no difference compared with the mesalazine group(P>0.05).[Conclusion] The Anchang Yuyang Recipe relieve the inflammatory injury of HT-29 cell and play a protective role probably by up-regulating secretion of IL-13,activating JAK1/STAT6 pathway,promoting the secretion of mucosal repair factor TFF3.