摘要
目的 探讨桃叶珊瑚苷(Acb)调节T细胞免疫球蛋白结构域和包含粘蛋白结构域分子3(TIM3)/半乳凝素9(Gal9)信号轴对子宫内膜癌上皮间质转化(EMT)和免疫逃逸的影响。方法 HEC-1B细胞分为Acb低剂量组(Acb-L组)、Acb中剂量组(Acb-M组)、Acb高剂量组(Acb-H组)、Acb-H+重组人TIM3蛋白(rh TIM3)组、空白组,5-乙炔基-2’脱氧尿嘧啶核苷(Ed U)染色和CCK-8检测HEC-1B细胞增殖;Transwell检测HEC-1B细胞侵袭及迁移;Western blot检测HEC-1B细胞中N-钙黏蛋白(N-cadherin)、TIM3、Gal9、E-钙黏蛋白(E-cadherin)蛋白。将HEC-1B细胞与CD8~+T细胞共培养,并用低、中、高剂量Acb以及高剂量Acb联合rh TIM3处理共培养细胞,流式细胞术检测CD8~+T细胞凋亡;ELISA检测共培养体系上清液中TNF-α、IFN-γ、粒酶B(granzyme B)水平。结果 与空白组比较,Acb-L组、Acb-M组、Acb-H组Ed U阳性细胞率、A_(450)值、细胞侵袭及迁移数、N-cadherin、TIM3、Gal9蛋白表达降低,E-cadherin蛋白表达升高,呈剂量依赖性(P<0.05);与Acb-H组比较,Acb-H+rh TIM3组Ed U阳性细胞率、OD_(450)值、细胞侵袭及迁移数、N-cadherin、TIM3、Gal9蛋白表达升高,E-cadherin蛋白表达降低(P<0.05)。与共培养组相比,Acb-L+共培养组、Acb-M+共培养组、Acb-H+共培养组CD8+T细胞凋亡率降低,共培养体系上清液中TNF-α、IFN-γ、granzyme B水平升高,呈剂量依赖性(P<0.05);与Acb-H+共培养组相比,Acb-H+rh TIM3+共培养组CD8+T细胞凋亡率升高,共培养体系上清液中TNF-α、IFN-γ、granzyme B水平降低(P<0.05)。结论 Acb抑制HEC-1B细胞EMT及免疫逃逸的机制可能与抑制TIM3/Gal9通路有关。
Objective To investigate the effects of aucubin(Acb)on epithelial mesenchymal transition(EMT)and immune escape in endometrial cancer by regulating the T-cell immunoglobulin domain and mucin do-main-containing molecule-3(TIM3)/galectin-9(Gal9)pathway.Methods HEC-1B cells were separated into low-dose Acb group(Acb-L group),medium-dose Acb group(Acb-M group),high-dose Acb group(Acb-H group),Acb-H+recombinant human TIM3 protein(rh TIM3)group,and blank group.5-ethynyl-2′deoxyuridine nucleoside(EdU)staining and CCK-8 were applied to detect HEC-1B cell proliferation.Transwell was applied to detect HEC-1B cell invasion and migration.Western Blot was applied to detect N-cadherin,TIM3,Gal9,and E-cadherin proteins in HEC-1B cells.HEC-1B cells were co cultured with CD8+T cells,and the co cultured cells were treated with low,medium,and high doses of Acb,and high-dose Acb combined with rh TIM3.Flow cytom-etry was applied to detect CD8+T cell apoptosis.ELISA was applied to detect levels of tumor necrosis factor-α(TNF-α),γ-interferon(IFN-γ),and granzyme B in the supernatant of co culture systems.Results Compared with the blank group,the EdU positive cell rate,OD450 value,cell invasion and migration numbers,the expression of N-cad-herin,TIM3,and Gal9 proteins were reduced in the Acb-L group,Acb-M group,and Acb-H group,the expression of E-cadherin protein was elevated,in a dose-dependent manner(P<0.05).Compared with the Acb-H group,the EdU positive cell rate,OD450 value,cell invasion and migration numbers,the expression of N-cadherin,TIM3,and Gal9 proteins were increased in the Acb-H+rh TIM3 group,the expression of E-cadherin protein was decreased(P<0.05).Compared with the co culture group,the apoptosis rate of CD8+T cells in the Acb-L+co culture group,Acb-M+co culture group,and Acb-H+co culture group decreased,the levels of TNF-α,IFN-γ,and granzyme B in the supernatant of the co culture system increased,in a dose-dependent manner(P<0.05).Compared with the Acb-H+co culture group,the apoptosis rate of CD8+T cells in the Acb-H+rh TIM3+co culture group increased,the levels of TNF-α,IFN-γ,and granzyme B in the supernatant of the co culture system decreased(P<0.05).Conclusion The mechanism of Acb in inhibiting EMT and immune escape of HEC-1B cells may be related to the inhibition of the TIM3/Gal9 pathway.
作者
王晶
张丽
汪鸽
宫亚萍
张杜鹃
李翡
WANG jing;ZHANG Li;WANG Ge;GONG Yaping;ZHANG Dujuan;LI Fei(Gynecological Second Ward,Affiliated Hospital of Shaanxi University of Traditional Chinese Medicine,Xianyang 712000,China)
出处
《解剖学研究》
2025年第4期323-329,共7页
Anatomy Research
基金
陕西省科学技术研究发展计划项目(2020SF-052)
陕西省教育厅专项科研计划项目(20JK0595)
吴阶平基金临床研究课题(320.6750.2022-18-46)。
作者简介
通讯作者:李翡,E-mail:13595024@qq.com。
