CAJ | 학술논문

目的用5-氟尿嘧啶(5-fluorouracil,5-FU)处理HeLa细胞,检测其NKG2D 配体 MICA 的表达及其对NK92细胞杀伤敏感性的变化.方法不同浓度的 5-FU处理 HeLa细胞,在不同时间点用半定量PCR及流式细胞术检测HeLa细胞表面的NKG2D配体MICA在RNA及蛋白水平的表达变化情况,用MTT法检测NKG2D抗体封闭NK92细胞的NKG2D受体前后, NK92 细胞对HeLa细胞的杀伤作用.结果不同浓度的5-FU作用于 HeLa 细胞后,半定量RT-PCR结果显示MICA表达随5-FU作用浓度增加逐渐增高.而且40 μg/ml 5-FU作用于HeLa细胞后随着作用时间的延长(0、8、16、24 h)MICA表达增加,流式细胞术检测结果表明,MICA表达的增加主要依赖于未凋亡细胞的 MICA表达.在40μg/ml 5-FU作用24 h,效靶比为2.5∶ 1,5∶ 1,10∶ 1,20∶ 1时都检测到NK92细胞对HeLa细胞的杀伤增强,杀伤作用可部分被NKG2D抗体抑制.结论 5-FU 能够上调HeLa细胞表面NKG2D配体MICA的表达,增强HeLa细胞对NK92细胞的敏感性,提示化疗联合NK细胞免疫治疗宫颈癌可产生协同作用,提高治疗效果.
목적 용5-불뇨밀정(5-fluorouracil,5-FU)처리HeLa세포,검측기NKG2D 배체 MICA 적표체급기대NK92세포살상민감성적변화.방법 불동농도적 5-FU처리 HeLa세포,재불동시간점용반정량PCR급류식세포술검측HeLa세포표면적NKG2D배체MICA재RNA급단백수평적표체변화정황,용MTT법검측NKG2D항체봉폐NK92세포적NKG2D수체전후, NK92 세포대HeLa세포적살상작용.결과 불동농도적5-FU작용우 HeLa 세포후,반정량RT-PCR결과현시MICA표체수5-FU작용농도증가축점증고.이차40 μg/ml 5-FU작용우HeLa세포후수착작용시간적연장(0、8、16、24 h)MICA표체증가,류식세포술검측결과표명,MICA표체적증가주요의뢰우미조망세포적 MICA표체.재40μg/ml 5-FU작용24 h,효파비위2.5∶ 1,5∶ 1,10∶ 1,20∶ 1시도검측도NK92세포대HeLa세포적살상증강,살상작용가부분피NKG2D항체억제.결론 5-FU 능구상조HeLa세포표면NKG2D배체MICA적표체,증강HeLa세포대NK92세포적민감성,제시화료연합NK세포면역치료궁경암가산생협동작용,제고치료효과.
Objective To investigate the expression of NKG2D ligand MICA in HeLa cells treated by 5 - fluorouracil (5-FU) and the susceptibility of treated HeLa cells to NK92 cell-mediated killing.Methods HeLa cells were treated with different concentrations of 5-FU. Expression of MICA in RNA and protein level was detected by semi-quantitative PCR and flow cytometry. NK cell-mediated killing of HeLa cells was determined by MTT assay. The cytolytic activity of NK92 cells was compared in the presence or absence of NKG2D antibody.Results The results of semi-quantitative RT-PCR showed that MICA was expressed with the increase of 5-FU dose range and upregulated along with the exposure time course (0h, 8h, 16h, 24h) of HeLa cells to 5-FU. Flow cytometry data demonstrated that the upregulation of MICA expression mainly result from the non-apoptotic cells. The cytolytic effect of NK92 cells on HeLa cells was enhanced by 5-FU treatment for 24h at different effect/target ratio. NK-cell mediated killing of HeLa cells was partially inhibited by NKG2D antibody.Conclusion 5-FU increases cell surface expression of NKG2D ligand MICA on HeLa cells, thus enhancing the sensitivity of Hela cells to lysitic effect of NK92 cells, suggesting that combination of chemotherapy and immunotherapy with NK cells might improve the treatment of cervical cancer patients.