幽门螺杆菌代谢产物腺苷通过A2A受体调控炎症因子分泌的机制研究

Mechanism of Helicobacter pylori metabolite adenosine in regulating secretion of inflammatory factors through A2A receptors

目的探讨幽门螺杆菌(H.pylori)代谢产物参与调控炎症因子分泌的潜在机制。方法大肠杆菌(E.Coli)或H.pylori感染后,检测人胃癌细胞AGS中关键炎症因子IL⁃33、TNF⁃α、IFN⁃γ的分泌水平。H.pylori感染AGS细胞24 h后,收取细胞上清进行液相色谱与串联质谱检测H.pylori的代谢物,并通过筛选鉴定关键代谢物。结果E.Coli或H.pylori感染后,AGS细胞中IL⁃33、TNF⁃α、IFN⁃γ的分泌水平上升。相比于感染E.Coli,感染H.pylori的AGS细胞分泌的IL⁃33、TNF⁃α、IFN⁃γ的水平较低。H.pylori代谢物腺苷降低IL⁃33、TNF⁃α、IFN⁃γ的分泌水平存在梯度依赖效应。腺苷合成酶敲除株H.pyloriΔAdA菌株感染后,AGS细胞的腺苷水平无显著改变。H.pyloriΔAdA感染后,AGS细胞中IL⁃33、TNF⁃α、IFN⁃γ的分泌水平均比H.pyloriWT感染的AGS细胞高。敲低A2A受体后,H.pylori感染的AGS细胞IL⁃33、TNF⁃α、IFN⁃γ的分泌水平上升。H.pyloriΔAdA或野生型H.pyloriWT感染小鼠4周后,小鼠血清中的IL⁃33、TNF⁃α、IFN⁃γ的水平均上升。H.pyloriΔAdA感染后小鼠血清中的IL⁃33、TNF⁃α、IFN⁃γ的水平上升更明显。此外,H.pyloriΔAdA感染后小鼠产生的抗幽门螺旋杆菌抗体的滴度上升。结论H.pylori表达的腺苷合成酶能够促进腺苷的产生,随后腺苷通过A2A受体调控炎症因子IL⁃33、TNF⁃α、IFN⁃γ的分泌,并且降低适应性免疫水平。

Objective To explore the potential mechanism of Helicobacter pylori(H.pylori)metabolites in regulating the secretion of inflammatory factors.Methods After infection with E.coli(E.Coli)or H.pylori,the secretion level of key inflammatory factors IL⁃33,TNF⁃αand IFN⁃γin human gastric cancer cells AGS was detected.Twenty⁃four hours after H.pylori infection of AGS cells,the cell supernatant was collected for liquid chromatogra⁃phy and tandem mass spectrometry to detect H.pylori metabolites,and key metabolites were identified by screen⁃ing.Results After E.Coli or H.pylori infection,the secretion level of IL⁃33,TNF⁃αand IFN⁃γin AGS cells increased.Compared with E.coli infection,AGS cells infected with H.pylori secreted lower level of IL⁃33,TNF⁃α,and IFN⁃γ.The H.pylori metabolite adenosine decreased the secretion level of IL⁃33,TNF⁃αand IFN⁃γin a gradi⁃ent⁃dependent manner.Adenosine level of AGS cells were not significantly changed after infection with adenosine synthase knockout strain H.pyloriΔAdA strain.After H.pyloriΔAdA infection,the secretion level of IL⁃33,TNF⁃α,and IFN⁃γin AGS cells was higher than that in H.pyloriWT⁃infected AGS cells.After knockdown of A2A receptor,the secretion level of IL⁃33,TNF⁃α,and IFN⁃γincreased in H.pylori⁃infected AGS cells.Four weeks after H.pyloriΔAdA or wild⁃type H.pyloriWT infection of mice,the level of IL⁃33,TNF⁃α,and IFN⁃γin the serum of mice all increased.The level of IL⁃33,TNF⁃αand IFN⁃γin the serum of mice after H.pyloriΔAdA infection increased significantly.In addition,the titers of anti⁃H.pylori antibodies produced in mice after H.pyloriΔAdA infection increased.Conclusion Adenosine synthase expressed by H.pylori can promote the production of adenosine,and then adenosine regulates the secretion of inflammatory factors of IL⁃33,TNF⁃αand IFN⁃γthrough A2A receptors,and reduces the level of adaptive immunity.