【摘要】 目的:探讨缺氧诱导基因2(HIG2)在喉癌细胞逃逸NK细胞免疫杀伤中的作用机制。方法:免疫组织化学法检测喉癌组织和癌旁组织中HIG2的表达;Western blot检测HIG2、NKG2D配体(MICA、MICB、ULBP1、ULBP2和ULBP3)的蛋白表达;Transwell实验检测Hep-2细胞的侵袭;流式细胞术检测Hep-2细胞的凋亡、CD3~-CD56~+的NK细胞纯度、NKG2D受体表达及Hep-2细胞NKG2D配体的表达;乳酸脱氢酶释放法检测不同效靶比时,NK细胞对Hep-2细胞的杀伤活性。结果:HIG2在喉癌组织中高表达。CD3~-CD56~+的NK细胞纯度大于90%。沉默HIG2抑制Hep-2细胞侵袭,促进细胞凋亡,提高NKG2D配体的表达,增强NK细胞对Hep-2细胞的杀伤敏感性。NKG2D抗体抑制NK细胞NKG2D受体表达,减弱NK细胞对沉默HIG2的Hep-2细胞的杀伤活性。结论:沉默HIG2可抑制喉癌细胞侵袭,促进细胞凋亡,上调NKG2D配体表达,增强NK细胞对喉癌细胞的杀伤敏感性。更多还原

【Abstract】 Objective:To investigate the mechanism of HIG2-mediated immune escape of laryngeal cancer cells from NK cell immune killing.Methods:Immunohistochemistry was used to detect the expression of HIG2 in laryngeal cancer tissues and adjacent tissues.Western blot was used to detect the protein level of HIG2 and NKG2 D ligands(MICA,MICB,ULBP1,ULBP2 and ULBP3).Transwell was used to detect invasion of Hep-2 cells.Flow cytometry was used to detect Hep-2 cell apoptosis, CD3~-CD56~+NK cell purity and NKG2 D receptor expression, and expression of Hep-2 cell NKG2 D ligand.LDH releasing assay was used to detect cytotoxicity of NK cells against Hep-2 cells at effect-to-target cell ratios of 5∶1,10∶1,20∶1.Results:The expression level of HIG2 was up-regulated in laryngeal cancer tissues.The purity of CD3~-CD56~+NK cells is greater than 90%.Silencing HIG2 inhibited Hep-2 cell invasion, promoted apoptosis, increased the expression of NKG2 D ligands(MICA,MICB,ULBP1,ULBP2,and ULBP3),and enhanced the cytotoxicity of NK cells against Hep-2 cells.The NKG2 D monoclonal antibody inhibited the expression of NKG2 D receptors on NK cells and attenuates the cytotoxicity of NK cells against Hep-2 cells that silence HIG2.Conclusion:Silencing HIG2 inhibited laryngeal carcinoma cell invasion, promoted apoptosis, enhanced the killing sensitivity of NK cells to laryngeal carcinoma cells by up-regulating the expression of NKG2 D ligands.更多还原