幽门螺杆菌感染对胃黏膜上皮细胞REDD1表达的影响及调控机制

Effect and mechanism of H.pylori in regulation of REDD1 expression in gastric mucosa

目的探讨DNA损伤应答蛋白1(regulated in development and DNA damage responses-1,REDD1)在幽门螺杆菌(Helicobacter pylori,H.pylori)感染中的表达及调控机制。方法建立H.pylori感染C57小鼠及胃上皮细胞模型,运用实时荧光定量PCR、免疫组织化学染色和Western blot检测REDD1 mRNA和蛋白的表达;并在细胞模型中采用信号通路抑制剂的方法探讨H.pylori感染诱导REDD1上调的机制。结果相对于未感染组,H.pylori感染小鼠胃黏膜中的REDD1水平显著增高;而相对于野生型(Wild Type,WT)全毒株,敲除cagA基因后,H.pylori感染诱导REDD1上调的能力则显著下降(P<0.05);H.pylori感染可诱导胃上皮细胞AGS REDD1表达上调,并具有时间、感染菌量以及cagA依赖性(P<0.05);P38/MAPK信号通路阻断可显著抑制H.pylori感染诱导的REDD1上调表达(P<0.05)。结论H.pylori依赖磷酸化的cagA蛋白激活MAPKp38通路诱导REDD1表达增高。

Objective Regulated in development and DNA damage responses-1(REDD1)is a conserved and ubiquitous protein,and is induced in response to multiple stimuli.However,its regulation,function and clinical relevance in Helicobacter pylori(H.pylori)-associated gastritis are presently unknown.The study aimed to investigate its expression and regulation mechanism in H.pylori infection.Methods C57 mice and gastric epithelial cells(AGS cells)were respectively infected with H.pylori to establish infected models.Then the expression of REDD1 at mRNA and protein levels was detected by quantitative PCR,immunohistochemical staining and Western blotting.The mechanism of REDD1 up-regulation induced by H.pylori infection was investigated by transfection with signaling pathway inhibitor,plasmid or dual-luciferase reporting analysis in cell models.Results The expression of REDD1 was increased in gastric mucosa of the H.pylori-infected mice than the un-infected ones.Compared with the wild-type strain of H.pylori,the strain with cagA knockout(△cagA H.pylori 11637 cells)showed decreased effect on the upregulation of REDD1(P<0.05).H.pylori infection induced up-regulation of REDD1 in AGS cells in a time-,cagA-and pathogen dose-dependent manner(P<0.05).Blocking of P38/MAPKp38 signaling pathway could significantly inhibit the upregulation of REDD1 induced by the infection(P<0.05).Conclusion H.pylori induces the up-regulation of REDD1 through activation of phosphorylated cagA in MAPKp38 pathway in the gastric mucosa of mouse with H.pylori infection.