CAJ | 학술논문

目的探讨荷载白细胞介素(IL)-24的条件增殖腺病毒ZD55-IL-24诱导黑色素瘤细胞凋亡的分子机制.方法 ZD55-IL-24感染人黑色素瘤A375细胞.Western blot和免疫沉淀技术检测细胞的IL-24、B淋巴细胞瘤/白血病-2(bcl-2)亚硝基化、bcl-2泛素化、bcl-2蛋白表达水平及半胱氨酰天冬氨酸特异性蛋白酶(Caspase)蛋白活性;加入不同浓度的一氧化氮(NO)调节剂改变bcl-2亚硝基化水平,检测其对bcl-2泛素化水平和Caspase蛋白活性及黑色素瘤细胞凋亡的影响.结果 ZD55-IL-24导致A375细胞bcl-2亚硝基化水平下调,泛素化水平上调,bcl-2蛋白水平下降,Caspase 信号通路激活,黑色素瘤细胞凋亡,在作用72 h后细胞凋亡达高峰,细胞凋亡率由对照组的11％上升到72％;NO供体SNP可促进bcl-2亚硝基化、抑制bcl-2泛素化水平从而上调bcl-2蛋白水平,细胞凋亡率由68％下降到19％;SNP拮抗剂DTT则加速ZD55-IL-24介导的肿瘤细胞凋亡,细胞凋亡率上升到93％;NO清除剂PTIO也同样促进黑色素瘤细胞的凋亡,细胞凋亡率上升到85％.结论 IL-24通过抑制bcl-2亚硝基化促进其泛素化降解,导致黑色素瘤细胞凋亡.
목적 탐토하재백세포개소(IL)-24적조건증식선병독ZD55-IL-24유도흑색소류세포조망적분자궤제.방법 ZD55-IL-24감염인흑색소류A375세포.Western blot화면역침정기술검측세포적IL-24、B림파세포류/백혈병-2(bcl-2)아초기화、bcl-2범소화、bcl-2단백표체수평급반광안선천동안산특이성단백매(Caspase)단백활성;가입불동농도적일양화담(NO)조절제개변bcl-2아초기화수평,검측기대bcl-2범소화수평화Caspase단백활성급흑색소류세포조망적영향.결과 ZD55-IL-24도치A375세포bcl-2아초기화수평하조,범소화수평상조,bcl-2단백수평하강,Caspase 신호통로격활,흑색소류세포조망,재작용72 h후세포조망체고봉,세포조망솔유대조조적11％상승도72％;NO공체SNP가촉진bcl-2아초기화、억제bcl-2범소화수평종이상조bcl-2단백수평,세포조망솔유68％하강도19％;SNP길항제DTT칙가속ZD55-IL-24개도적종류세포조망,세포조망솔상승도93％;NO청제제PTIO야동양촉진흑색소류세포적조망,세포조망솔상승도85％.결론 IL-24통과억제bcl-2아초기화촉진기범소화강해,도치흑색소류세포조망.
Objective To investigate tumor-selective replicating adenovirus ZD55-interleukin (IL)-24 induces B-cell lymphoma/leukemia-2 (bcl-2) S-nitrosylation decline and further facilitates bcl-2 ubiquitin degradation,leading to melanoma cell apoptosis.Methods ZD55-IL-24 was transfected into melanoma A375 cells,bcl-2 S-nitrosylation,bcl-2 ubiquitination,bcl-2 protein level and downstream Caspase protein were detected by Western blotting and Co-IP.Furthermore,nitric oxide (NO) regulator single nucleotide polymorphism (SNP),PTIO and DTT were used to coffirm the relationship between bcl-2 S-nitrosylation and its ubiquitination.Results ZD55-IL-24 induced the bcl-2 protein level decline in time-dependent manner.Moreover,ZD55-IL-24 decreased bcl-2 S-nitrosylation and increased bcl-2 ubiquitination,which initiated the Caspase signal pathway activation and promoted melanoma cell apoptosis from 11％ to 72％ ; NO donor SNP promoted bcl-2 S-nitrosylation but restrained bcl-2 ubiquitination,which restored bcl-2 protein level.Moreover,melanoma cell apoptosis decreased from 68％ to 19％.In contrast,SNP antagonist DTT reversed the above results by enhancing cell apoptosis to 93 ％.NO inhibitor PTIO similarly increased melanoma cell apoptosis to 85％.Conclusion ZD55-IL-24 reduced bcl-2 S-nitrosylation and facilitated bcl-2 ubiquitin proteasome degradation,thereby resulting in melanoma cell apoptosis.