CAJ | 학술논문

目的探讨顺铂影响胃癌细胞对肿瘤坏死因子相关凋亡诱导配体(TRAIL)敏感性的机制.方法以顺铂和TRAIL单独及联合作用MGC803细胞,采用四甲基偶氮唑蓝(MTT)法测定各组细胞的增殖能力,流式细胞仪检测细胞凋亡,Western blot检测caspase-8和caspase-3的蛋白表达,免疫荧光显微技术观察脂筏和死亡受体4(DR4)的分布.以50 mg/L的制霉菌素预处理MGC803细胞1 h,之后加入顺铂和TRAIL,观察细胞凋亡的变化.结果 100 μg/L TRAIL作用MGC803细胞24 h,增殖抑制率为(8.51±3.45)%,细胞凋亡率为(3.26±0.89)%.8.49 mg/L顺铂作用MGC803细胞24 h,增殖抑制率为(52.58±4.57)%,细胞凋亡率为(23.10±3.41)%.100 μg/L TRAIL和8.49 mg/L顺铂联合作用时,细胞增殖抑制率提高至(76.43±5.35)%,细胞凋亡率提高到(42.56±4.11)%,均高于相同浓度顺铂和TRAIL单独作用组(P＜0.05).同时检测到caspase-8和caspase-3的裂解.TRAIL未引起明显的脂筏和DR4聚集,而顺铂明显促进了DR4在聚集脂筏内的定位.50 mg/L制霉菌素处理MGC803细胞24 h,细胞凋亡率为(3.66±0.52)%.经制霉菌素预处理后,顺铂作用下MGC803细胞的凋亡率为(22.76±2.97)%,与未经制霉菌素预处理组[(25.74±3.28)%]差异无统计学意义(P=0.248);而顺铂和TRAIL联合作用下细胞凋亡率为(31.52±3.99)%,与未经制霉菌素预处理组[(43.16±4.26)%]差异有统计学意义(P＜0.001).结论顺铂通过促进死亡受体在脂筏聚集增强了TRAIL诱导的胃癌MGC803细胞凋亡.
목적 탐토순박영향위암세포대종류배사인자상관조망유도배체(TRAIL)민감성적궤제.방법 이순박화TRAIL단독급연합작용MGC803세포,채용사갑기우담서람(MTT)법측정각조세포적증식능력,류식세포의검측세포조망,Western blot검측caspase-8화caspase-3적단백표체,면역형광현미기술관찰지벌화사망수체4(DR4)적분포.이50 mg/L적제매균소예처리MGC803세포1 h,지후가입순박화TRAIL,관찰세포조망적변화.결과 100 μg/L TRAIL작용MGC803세포24 h,증식억제솔위(8.51±3.45)%,세포조망솔위(3.26±0.89)%.8.49 mg/L순박작용MGC803세포24 h,증식억제솔위(52.58±4.57)%,세포조망솔위(23.10±3.41)%.100 μg/L TRAIL화8.49 mg/L순박연합작용시,세포증식억제솔제고지(76.43±5.35)%,세포조망솔제고도(42.56±4.11)%,균고우상동농도순박화TRAIL단독작용조(P＜0.05).동시검측도caspase-8화caspase-3적렬해.TRAIL미인기명현적지벌화DR4취집,이순박명현촉진료DR4재취집지벌내적정위.50 mg/L제매균소처리MGC803세포24 h,세포조망솔위(3.66±0.52)%.경제매균소예처리후,순박작용하MGC803세포적조망솔위(22.76±2.97)%,여미경제매균소예처리조[(25.74±3.28)%]차이무통계학의의(P=0.248);이순박화TRAIL연합작용하세포조망솔위(31.52±3.99)%,여미경제매균소예처리조[(43.16±4.26)%]차이유통계학의의(P＜0.001).결론 순박통과촉진사망수체재지벌취집증강료TRAIL유도적위암MGC803세포조망.
Objective Gastric cancer cells are insensitive to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). To sensitize gastric cancer cells to TRAIL, we treated gastric cancer MGC803 cells with TRAIL and cisplatin. Methods Cell proliferation was measured using MTT assay. Cell apoptosis was determined by flow cytometry. Expression of proteins was analyzed by Western blot. The distribution of lipid rafts and death receptors was analyzed by immunofluorescence microscopy. MGC803 cells were pretreated with 50 mg/L nystatin for 1 h, and followed by the treatment of cisplatin and TRAIL. Results 100 μg/L TRAIL resulted in (8.51±3.45)% inhibition of cell proliferation and caused (3.26±0.89)% cell apoptosis in MGC803 cells. Compared with the treatment with cisplatin alone, treatment with TRAIL (100 μg/L) and cisplatin (8.49 mg/L, IC50 dose of 24 h) led to a dramatic increase in both inhibition of cell proliferation [(52.58±4.57)% vs. (76.43±5.35)%, P＜0.05]and cell apoptosis [(23. 10±3. 41)% vs. (42.56±4.11)%, P＜0.05]. Moreover, cleavage of caspase-8 and caspase-3 was detected. TRAIL (100 μg/L) did not induce obvious lipid rafts aggregation and death receptor 4 (DR4) clustering, while cisplatin (8.49 mg/L) significantly promoted the localization of DR4 in aggregated lipid rafts. Pretreatment with 50 mg/L nystatin, a cholesterol-sequestering agent, triggered (3.66±0.52)% cell apoptosis after 24 h. Pretreatment with nystatin for 1 h before the addition of 8.49 mg/L cisplatin for 24 h caused a decreased tendency to cell apoptosis [(25.74±3.28)% vs. (22.76±2.97)%]. While, pretreatment with nystatin before the addition of cisplatin and TRAIL, the proportion of apoptotic cells decreased from (43. 16±4.26)% to (31.52 ± 3.99)% (P＜0.05). Conclusion Cisplatin enhances TRAIL-induced apoptosis in gastric cancer MGC803 cells through clustering death receptors into lipid rafts.