摘要
目的探究前蛋白转化酶枯草溶菌素9(PCSK9)在幽门螺杆菌(H.pylori)感染导致炎症反应中的作用机制。方法收集2020年5月1日至2021年1月31日初次就诊于十堰市太和医院消化内科的60例胃炎患者(H.pylori阳性和阴性患者各30例)与30例健康体检者,检测其血清PCSK9水平。选取H.pylori标准菌株、4种H.pylori临床分离菌株、正常胃上皮细胞系GES-1与THP-1细胞经诱导形成的巨噬细胞。将H.pylori菌株与胃上皮细胞系GES-1共培养以制备不同上清液培养基,设立磷酸盐缓冲液空培养基即阴性对照培养基(阴性对照组)、H.pylori与GES-1细胞共培养上清液培养基(H.pylori感染GES-1组)、正常GES-1细胞上清液培养基(H.pylori未感染GES-1组)、H.pylori与GES-1细胞共培养上清液+PCSK9中和抗体培养基(抗PCSK9组)、H.pylori与GES-1细胞共培养上清液+人免疫球蛋白G2培养基(同型对照组)。采用Transwell迁移实验、实时荧光定量聚合酶链反应、平板集落形成实验、H.pylori吞噬溶酶体共定位实验等比较H.pylori感染GES-1组、H.pylori未感染GES-1组和阴性对照组,以及抗PCSK9组和同型对照组的巨噬细胞迁移数、CC趋化因子受体(CCR2)mRNA相对表达水平、白细胞介素(IL)-6和细胞坏死因子(TNF)-α释放水平、CD8^(+)T细胞胞膜磷酸化水平、巨噬细胞菌落数,分析PCSK9对H.pylori感染导致炎症反应的调控机制。统计学方法采用独立样本t检验。结果H.pylori阳性胃炎患者的血清PCSK9水平高于H.pylori阴性胃炎患者和健康体检者[(384.00±57.57)μg/L比(208.80±48.89)、(176.10±47.14)μg/L],差异均有统计学意义(t=12.71、15.31,均P<0.001)。与阴性对照培养基相比,H.pylori标准菌株和4种分离菌株均可诱导正常胃上皮细胞系GES-1分泌PCSK9[(1267.00±287.50]g/L比(2717.00±199.20)、(4858.00±302.40)、(3167.00±334.20)、(6075.00±597.30)、(4283.00±331.20)g/L],差异均有统计学意义(t=10.15、21.09、10.56、17.77、16.85,�
Objective To explore the mechanism of proprotein convertase subtilisin/kexin type 9(PCSK9)in the inflammatory response induced by Helicobacter pylori(H.pylori)infection.Methods From May 1,2020 to January 31,2021,60 patients with gastritis(30 H.pylori positive and 30 H.pylori negative)and 30 healthy individuals,who initially visited the Department of Gastroenterology,Shiyan Taihe Hospital were collected,and their serum PCSK9 levels were detected.Normal gastric epithelial cell line GES-1 and macrophages induced from THP-1 cells,and GES-1 infected with H.pylori were selected to prepare different supernatant media.Phosphate buffer saline empty medium(negative control group),normal GES-1 cell supernatant medium(H.pylori-uninfected GES-1 group),H.pylori infected GES-1 cell supernatant medium(H.pylori infected GES-1 group),H.pylori infected GES-1 cell supernatant+PCSK9 neutralizing antibody medium(anti PCSK9 group),H.pylori infected GES-1 cell supernatant+human immunoglobulin G medium(isotype control group)were established.The differences between H.pylori infected GES-1 group and H.pylori-uninfected GES-1 group,negative control group,anti PCSK9 group and isotype control group in number of migrated macrophages,relative expression level of CC chemokine receptor(CCR2),the levels of released interleukin(IL)-6 and cell necrosis factor(TNF)-α,level of CD8^(+)T cell membrane phosphorylation,and the number of macrophage colonies were determined by Transwell assay,real time fluorescence quantitative polymerase chain reaction,plate colony assay,H.pylori and phagocytosis lysosome co-localization assay.The regulating mechanism of PCSK9 in H.pylori infection induced inflammation was analyzed.Independent sample t test was used for statistical analysis.Results The serum level of PCSK9 of patients with H.pylori positive gastritis was higher than that of patients with H.pylori negative gastritis and healthy individuals((384.00±57.57)g/L vs.(208.80±48.89)and(176.10±47.14)g/L),and the differences were statistically significant(t=12.71
作者
朱晓文
袁纯辉
王军
姚莉
蔡秦真
向贇
Zhu Xiaowen;Yuan Chunhui;Wang Jun;Yao Li;Cai Qinzhen;Xiang Yun(Department of Gastroenterology,Shiyan Taihe Hospital,Shiyan 442099,China;Department of Laboratory Medicine,Wuhan Children′s Hospital(Wuhan Maternal and Child Healthcare Hospital),Tongji Medical College,Huazhong University of Science and Technology,Wuhan 430016,China;Department of Pathology,Shiyan Taihe Hospital,Shiyan 442099,China)
出处
《中华消化杂志》
CAS
CSCD
北大核心
2022年第5期304-313,共10页
Chinese Journal of Digestion
基金
湖北省卫生健康委员会科研项目(WJ2021M016)
武汉市科技局"黄鹤英才计划"项目(武人才办[2017]2号)
武汉市卫生计生委科研计划资助项目(WX21Q50)。
