牙龈卟啉单胞菌通过GARP促进TGF-β/SMAD轴介导食管鳞状细胞癌细胞的上皮间质转化  

Porphyromonas gingivalis promotes epithelial-mesenchymal transition of esophageal squamous cell carcinoma cells mediated by TGF-β/SMAD signaling via GARP

作　　者：张升华 杨静怡 祁春晖 乔亮[2] 高社干[1] 齐义军 ZHANG Shenghua;YANG Jingyi;QI Chunhui;QIAO Liang;GAO Shegan;QI Yijun(State Key Laboratory of Esophageal Cancer Prevention&Treatment,Henan Key Laboratory of Microbiome and Esophageal Cancer Prevention and Treatment,Henan Key Laboratory of Cancer Epigenetics,Cancer Hospital,the First Affiliated Hospital,College of Clinical Medicine of Henan University of Science and Technology,Luoyang 471003,Henan,China;Department of Pathology,the First Affiliated Hospital of Henan University of Science and Technology,Luoyang 471003,Henan,China)

机构地区：[1]河南科技大学临床医学院,河南科技大学第一附属医院,肿瘤医院,省部共建食管癌防治国家重点实验室,河南省微生态与食管癌防治重点实验室,河南省肿瘤表观遗传重点实验室,河南洛阳471003 [2]河南科技大学第一附属医院病理科,河南洛阳471003

出　　处：《中国肿瘤生物治疗杂志》2024年第8期769-776,共8页Chinese Journal of Cancer Biotherapy

基　　金：国家自然科学基金(No.81872037);河南省医学科技攻关-省部共建重点项目(No.SBGJ202002100)。

摘　　要：目的:阐明牙龈卟啉单胞菌(Pg)诱导食管鳞状细胞癌(ESCC)细胞发生上皮间质转化(EMT)的分子机制。方法:KEGG分析Pg诱导的ESCC差异表达基因富集的生物学通路,WB和/或免疫荧光法检测Pg诱导的ESCC细胞中糖蛋白A重复优势蛋白(GARP)、TGF-β、pSMAD/SMAD、Snail、Oct4和EMT相关分子表达的变化,ELISA检测TGF-β1水平的变化,免疫组织化学法检测ESCC组织中GARP和TGF-β1的表达规律,Transwell实验和动物实验验证Pg对ESCC的促进作用。结果:ESCC细胞感染Pg后,TGF-β、Hippo、PI3K/Akt等信号通路被激活;Pg感染刺激ESCC细胞分泌总TGF-β1和活性TFG-β1的水平升高(均P<0.01),使SMAD2/3磷酸化并发生核转位,诱导N-cadherin、Snail、Oct4等蛋白表达升高、E-cadherin蛋白表达降低,由此促进ESCC细胞的迁移、侵袭和裸鼠皮下移植瘤的生长(均P<0.01)。在ESCC细胞中沉默GARP表达后,逆转了Pg所诱导的上述ESCC细胞表型变化。Pg丰度高的ESCC组织中TGF-β1和GARP蛋白表达高于低丰度的ESCC组织,且Pg丰度与TGF-β1、GARP表达存在正向关联(P=0.0015)。结论:Pg通过GARP激活TGF-β/SMAD轴促进ESCC细胞发生EMT,进而促进ESCC细胞的迁移、侵袭和生长,清除Pg或阻断TGF-β信号转导则可阻断上述Pg对ESCC的促进作用。Objective:To elucidate the molecular mechanism by which Porphyromonas gingivalis(Pg)induces epithelialmesenchymal transition(EMT)in esophageal squamous cell carcinoma(ESCC)cells.Methods:KEGG was used to identify the biological pathways enriched by Pg-induced differentially expressed genes in ESCC.WB and/or immunofluorescence(IF)were used to detect the changes in the expression of glycoprotein-A repetitions predominant protein(GARP),TGF-β,pSMAD/SMAD,Snail,Oct4 and EMT-related molecules in Pg-induced ESCC cells.ELISA was used to measure changes in TGF-β1 level.Immunohistochemistry was used to detect the expression of GARP and TGF-β1 in ESCC tissues.The tumorigenic effect of Pg on ESCC was verified by Transwell assays and animal experiments.Results:Pg activated multiple signaling pathways,such as TGF-β,Hippo,and PI3K/Akt.Pginfection stimulated an increased secretion of total TGF-β1 and active TFG-β1 in ESCC cells(P<0.01),leading to SMAD2/3 phosphorylation and nuclear translocation,which induced increased protein expression of N-cadherin,Snail,and Oct4 and decreased E-cadherin.These alterations collectively promoted the migration and invasion of ESCC cells and the growth of subcutaneous tumor in nude mice(all P<0.01).Silencing GARP expression in ESCC cells reversed the phenotypic changes induced by Pg.The protein levels of TGF-β1 and GARP in ESCC tissues with high Pg abundance were higher than those in ESCC tissues with low Pg abundance.The abundance of Pg was positively correlated with the protein expression of TGF-β1 and GARP(P=0.0015).Conclusion:Pg activates the TGF-β/Smad axis through GARP to promote the occurrence of EMT in ESCC cells,thereby facilitating the migration,invasion and growth of ESCC cells.Pg clearance or TGF-βsignaling blockade can reverse these Pg-induced promotive effects on ESCC.

关 键 词：食管鳞状细胞癌 牙龈卟啉单胞菌 糖蛋白A重复优势蛋白 TGF-Β SMAD 上皮间质转化 

分 类 号：R735.1[医药卫生—肿瘤] R730.2[医药卫生—临床医学]