目的探讨miR-183对肝癌细胞MHC-Ⅰ类相关蛋白A/B(MICA/B)表达和自然杀伤(NK)细胞杀伤作用影响及其分子机制。方法实验设置miR-con组、miR-183组、anti-miR-con组、anti-miR-183组、pcDNA组、pcDNA-CX3CL1组、antimiR-183+si-con组、anti...目的探讨miR-183对肝癌细胞MHC-Ⅰ类相关蛋白A/B(MICA/B)表达和自然杀伤(NK)细胞杀伤作用影响及其分子机制。方法实验设置miR-con组、miR-183组、anti-miR-con组、anti-miR-183组、pcDNA组、pcDNA-CX3CL1组、antimiR-183+si-con组、anti-miR-183+CX3CL1组、对照(NC)组。实时荧光定量PCR(RT-qPCR)检测miR-183和CX3CL1 m RNA表达水平;蛋白质印迹(Western blot)法检测CX3CL1、MICA和MICB蛋白表达;流式细胞术检测肝癌细胞表面MICA/B的表达;NK细胞与肝癌细胞共培养后,细胞计数试剂盒8(CCK-8)法检测NK细胞对肝癌细胞的杀伤作用;酶联免疫吸附法(ELISA)法检测肝癌细胞与NK细胞共培养后培养液中肿瘤坏死因子-α(TNF-α)和干扰素γ(IFN-γ)水平;荧光素酶报告实验检测miR-183和CX3CL1的靶向关系。结果肝癌组织和肝癌细胞中miR-183高表达,CX3CL1、MICA和MICB低表达。干扰miR-183表达和过表达CX3CL1,肝癌细胞中MICA、MICB阳性表达率显著升高,与NK细胞共培养后TNF-α和IFN-γ水平显著升高,NK细胞对肝癌细胞的杀伤率显著升高(P<0.05)。miR-183靶向调控CX3CL1,沉默CX3CL1逆转了干扰miR-183对肝癌细胞MICA/B表达和NK细胞杀伤作用的影响。结论抑制表达miR-183可增加肝癌细胞MICA/B表达,增加NK细胞对肝癌细胞的杀伤作用,其机制可能与CX3CL1有关。展开更多
MicroRNAs （miRNAs） are a class of small non-coding regulatory RNAs, and changes in miRNAs are involved in tumor origin and progression. Studies have shown that miR-20a is overexpressed in human ovarian cancer tissue...MicroRNAs （miRNAs） are a class of small non-coding regulatory RNAs, and changes in miRNAs are involved in tumor origin and progression. Studies have shown that miR-20a is overexpressed in human ovarian cancer tissues and that this miRNA enhances long-term cellular proliferation and invasion capabilities. In this study, a positive correlation between serum miR-20a expression and ovarian cancer stage was observed. We found that miR-20a binds directly to the 3＇-untranslated region of MICA/B mRNA, resulting in its degradation and reducing its protein levels on the plasma membrane. Reduction of membrane-bound MICA/B proteins, which are ligands of the natural killer group 2 member D （NKG2D） receptor found on natural killer （NK） cells, y＋ T cells and CD8＋ T cells, allows tumor cells to evade immune-mediated killing. Notably, antagonizing miR-20a action enhanced the NKG2D-mediated killing of tumor cells in both in vitro and in vivo models of tumors. Taken together, our data indicate that increased levels of miR-20a in tumor cells may indirectly suppress NK cell cytotoxicity by downregulating MICA/B expression. These data provide a potential link between metastasis capability and immune escape of tumor cells from NK cells.展开更多
文摘目的探讨miR-183对肝癌细胞MHC-Ⅰ类相关蛋白A/B(MICA/B)表达和自然杀伤(NK)细胞杀伤作用影响及其分子机制。方法实验设置miR-con组、miR-183组、anti-miR-con组、anti-miR-183组、pcDNA组、pcDNA-CX3CL1组、antimiR-183+si-con组、anti-miR-183+CX3CL1组、对照(NC)组。实时荧光定量PCR(RT-qPCR)检测miR-183和CX3CL1 m RNA表达水平;蛋白质印迹(Western blot)法检测CX3CL1、MICA和MICB蛋白表达;流式细胞术检测肝癌细胞表面MICA/B的表达;NK细胞与肝癌细胞共培养后,细胞计数试剂盒8(CCK-8)法检测NK细胞对肝癌细胞的杀伤作用;酶联免疫吸附法(ELISA)法检测肝癌细胞与NK细胞共培养后培养液中肿瘤坏死因子-α(TNF-α)和干扰素γ(IFN-γ)水平;荧光素酶报告实验检测miR-183和CX3CL1的靶向关系。结果肝癌组织和肝癌细胞中miR-183高表达,CX3CL1、MICA和MICB低表达。干扰miR-183表达和过表达CX3CL1,肝癌细胞中MICA、MICB阳性表达率显著升高,与NK细胞共培养后TNF-α和IFN-γ水平显著升高,NK细胞对肝癌细胞的杀伤率显著升高(P<0.05)。miR-183靶向调控CX3CL1,沉默CX3CL1逆转了干扰miR-183对肝癌细胞MICA/B表达和NK细胞杀伤作用的影响。结论抑制表达miR-183可增加肝癌细胞MICA/B表达,增加NK细胞对肝癌细胞的杀伤作用,其机制可能与CX3CL1有关。
文摘MicroRNAs （miRNAs） are a class of small non-coding regulatory RNAs, and changes in miRNAs are involved in tumor origin and progression. Studies have shown that miR-20a is overexpressed in human ovarian cancer tissues and that this miRNA enhances long-term cellular proliferation and invasion capabilities. In this study, a positive correlation between serum miR-20a expression and ovarian cancer stage was observed. We found that miR-20a binds directly to the 3＇-untranslated region of MICA/B mRNA, resulting in its degradation and reducing its protein levels on the plasma membrane. Reduction of membrane-bound MICA/B proteins, which are ligands of the natural killer group 2 member D （NKG2D） receptor found on natural killer （NK） cells, y＋ T cells and CD8＋ T cells, allows tumor cells to evade immune-mediated killing. Notably, antagonizing miR-20a action enhanced the NKG2D-mediated killing of tumor cells in both in vitro and in vivo models of tumors. Taken together, our data indicate that increased levels of miR-20a in tumor cells may indirectly suppress NK cell cytotoxicity by downregulating MICA/B expression. These data provide a potential link between metastasis capability and immune escape of tumor cells from NK cells.
