萝卜硫素抑制幽门螺杆菌诱导胃黏膜上皮细胞NLRP3炎症小体活化的研究

Study on Sulforaphane Suppresses NLRP3 Inflammasome Activation in Gastric Mucosa Epithelial Cells Triggered by Helicobacter Pylori

关键词:  萝卜硫素  幽门螺杆菌  胃黏膜上皮细胞  NLRP3炎症小体

Key words:  sulforaphane  Helicobacter pylori  gastric mucosa epithelial cells  NLRP3 inflammasome

摘要:

目的 研究萝卜硫素（sulforaphane，SFP）对幽门螺杆菌感染的胃黏膜上皮GES-1细胞线粒体和氧化应激损伤及NLRP3炎症小体活化的影响。方法 SFP （100 ng·mL^-1）预处理GES-1细胞12 h，然后10⁹CFU·mL^-1的幽门螺杆菌感染细胞6 h；CCK8检测细胞活力，qRT-PCR分析PGC1α、COX-4、NRF1、TFAM、SOD1和HO-1 mRNA表达水平，Western blotting检测NLRP3、CASP1 p20、CASP1 p10、pro-IL-1β和IL-1β蛋白表达水平，并对GES-1细胞线粒体膜电位、氧耗量和ROS水平进行检测。结果 幽门螺杆菌感染可明显诱导NLRP3、CASP1 p20、CASP1 p10、IL-1β蛋白和PGC1α、COX-4、NRF1 mRNA表达，促使氧消耗量和ROS水平增高，而细胞活力、SOD1、HO-1、TFAM mRNA表达和线粒体膜电位降低（P<0.05）；SFP处理后能明显抑制NLRP3、IL-1β蛋白和PGC1α、COX-4、NRF1 mRNA表达，降低氧消耗量和ROS水平，而细胞活力、SOD1、HO-1、TFAM mRNA表达和线粒体膜电位增高（P<0.05）；另外，ROS抑制剂NAC也能显著抑制NLRP3炎症小体和IL-1β表达水平。结论 SFP可通过降低线粒体损伤和ROS水平改善幽门螺杆菌感染诱导GES-1细胞炎症反应。

Abstract:

OBJECTIVE To study the effects of sulforaphane(SFP) on the mitochondrial damage, oxidative stress injury and NLRP3 inflammasome activation in gastric mucosa epithelial cells(GES-1) induced by Helicobacter pylori infection. METHODS GES-1 cells pre-treated with 100 ng·mL^-1 of SFP for 12 h, then cells were infected with 10⁹ CFU·mL^-1 of Helicobacter pylori for 6 h. CCK8 was used to detect the cell viability. qRT-PCR was performed to measure the expression levels of PGC1α, COX-4, NRF1, TFAM, SOD1 and HO-1 mRNA. Western blotting was used to determine the expressions of NLRP3, CASP1 p20, CASP1 p10, pro-IL-1β and IL-1β. In addition, the mitochondrial membrane potential, oxygen consumption and ROS level were detected. RESULTS Helicobacter pylori infection could evidently induce the expression levels of NLRP3, CASP1 p20, CASP1 p10, IL-1β protein and PGC1α, COX-4, NRF1 mRNA, and promote the oxygen consumption and ROS level, whereas significantly inhibit the cell viability, SOD1, HO-1, TFAM mRNA expression and the mitochondrial membrane potential(P<0.05). Furthermore, the expression levels of NLRP3, IL-1β protein and PGC1α, COX-4, NRF1 mRNA, and the oxygen consumption and ROS level were greatly decreased after treatment with SFP in GES-1 cells induced with Helicobacter pylori infection, whereas significantly induced the cell viability, SOD1, HO-1, TFAM mRNA expression and the mitochondrial membrane potential(P<0.05). In addition, ROS inhibitor(NAC) also could suppress the expressions of NLRP3 inflammasome and IL-1β. CONCLUSION SFP can improve the inflammatory response of GES-1 cells triggered by Helicobacter pylori infection by reducing mitochondrial damage and ROS levels.