摘要
目的探讨奥美拉唑(OME)对幽门螺杆菌(Hp)感染的胃黏膜上皮细胞增殖、凋亡、自噬、炎症的影响及其机制。方法人胃黏膜上皮细胞GES-1用Hp感染,不同浓度的OME处理或用si-NC、si-GAS5、si-GAS5与anti-miR-NC、si-GAS5与anti-miR-221-3p转染。采用MTT检测细胞活力,流式细胞术检测细胞凋亡率,ELISA检测IL-1β、IL-6、TNF-α水平,qRT-PCR检测GAS5、miR-221-3p表达量,双荧光素酶报告实验与RIP实验检测GAS5与miR-221-3p的靶向关系,Western blot检测LC3Ⅰ、LC3Ⅱ和Beclin1蛋白表达。结果与对照组比较,Hp组细胞活力、GAS5水平降低(P<0.05),凋亡率、LC3Ⅱ/LC3Ⅰ比值、Beclin1蛋白、IL-1β、IL-6、TNF-α水平、miR-221-3p水平升高(P<0.05),与Hp组比较,OME不同剂量组细胞活力、GAS5水平升高(P<0.05),凋亡率、LC3Ⅱ/LC3Ⅰ比值、Beclin1蛋白、IL-1β、IL-6、TNF-α水平、miR-221-3p水平降低(P<0.05),与Hp+OME+si-NC组比较,Hp+OME+si-GAS5组细胞活力降低(P<0.05),凋亡率、LC3Ⅱ/LC3Ⅰ比值、Beclin1蛋白、IL-1β、IL-6、TNF-α水平升高(P<0.05),与Hp+OME+si-GAS5+anti-miR-NC组比较,Hp+OME+si-GAS5+anti-miR-221-3p组细胞活力升高(P<0.05),凋亡率、LC3Ⅱ/LC3Ⅰ比值、Beclin1蛋白、IL-1β、IL-6和TNF-α水平降低(P<0.05),双荧光素酶报告实验与RIP实验证实GAS5可靶向结合miR-221-3p。结论OME通过调控GAS5/miR-221-3p轴促进Hp感染的胃黏膜上皮细胞增殖,抑制细胞凋亡、自噬及炎症反应。
Objective To investigate the effects of omeprazole(OME)on proliferation,apoptosis,autophagy and inflammation of gastric mucosal epithelial cells infected with Helicobacter pylori(Hp)and its mechanism.Methods Human gastric mucosal epithelial cells GES-1 were infected with Hp and treated with different concentrations of omeprazole or transfected with si-NC,si-GAS5,si-GAS5 and anti-miR-NC,si-GAS5 and anti-miR-221-3 p.MTT was used to detect cell viability.Flow cytometry was used to detect apoptosis rate.ELISA was used to detect the levels of IL-1β,IL-6 and TNF-α.QRT-PCR was used to detect the expressions of GAS5 and miR-221-3 p.Dual luciferase reporter experiment and RIP experiment were used to detect the targeting relationship between GAS5 and miR-221-3 p.Western blot method was used to detect the protein expressions of LC3Ⅰ,LC3Ⅱ,Beclin1.Results Compared with Control group,cell viability and GAS5 level in Hp group were decreased(P<0.05),apoptosis rate,LC3Ⅱ/LC3Ⅰratio,Beclin1 protein,IL-1β,IL-6,TNF-αand miR-221-3 p level were increased(P<0.05).Compared with Hp group,Cell viability and GAS5 levels in OME group were increased(P<0.05),apoptosis rate,LC3Ⅱ/LC3Ⅰratio,Beclin1 protein,IL-1β,IL-6,TNF-αand miR-221-3 p levels were decreased(P<0.05).Compared with Hp+OME+si-NC group,cell viability in Hp+OME+si-GAS5 group was decreased(P<0.05),apoptosis rate,LC3Ⅱ/LC3Ⅰratio,Beclin1 protein,IL-1β,IL-6 and TNF-αlevels were increased(P<0.05).Compared with Hp+OME+si-GAS5+anti-miR-NC group,cell viability in Hp+OME+si-GAS5+anti-miR-221-3 p group was increased(P<0.05),apoptosis rate,LC3Ⅱ/LC3Ⅰratio,Beclin1 protein,IL-1β,IL-6,TNF-αlevels were decreased(P<0.05).Double luciferase reporting assay and RIP assay confirmed that GAS5 could target miR-221-3 p.Conclusion Omeprazole promoted the proliferation of Hp-infected gastric mucosal epithelial cells by regulating the GAS5/miR-221-3 p axis,and could inhibit cell apoptosis,autophagy and inflammation.
作者
房群
王丽宁
FANG Qun;WANG Li-ning(Qilu Medical University,Zibo Shandong 255300,China)
出处
《毒理学杂志》
CAS
CSCD
2022年第3期231-236,242,共7页
Journal of Toxicology
