Molto Luis, Rayman Pat, Paszkiewicz-Kozik Ewa, Thornton Mark, Reese Lisa, Thomas John C, Das Tanya, Kudo Daisuke, Bukowski Ronald, Finke James, Tannenbaum Charles
Department of Immunology, Lerner Research Institute, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, Ohio 44195, USA.
Clin Cancer Res. 2003 Sep 15;9(11):4060-8.
Tumors induce T-cell apoptosis as a mechanism of inhibiting antitumor immunity. Using coculture experiments, it has been shown that tumor lines stimulate T-cell apoptosis by a pathway involving a mitochondrial permeability transition and cytochrome c release. Activated T cells express abundant levels of Bcl-2, an antiapoptotic molecule that would be expected to confer resistance to such tumor-mediated killing. We examined the mechanism by which Bcl-2 is dysregulated in T cells exposed to the renal tumor line SK-RC-45, and we determined whether overexpressing Bcl-2 protects T cells from tumor-mediated apoptosis.
Activated T lymphocytes and Jurkat cells transfected or not transfected with Bcl-2 were exposed to SK-RC-45 for 48-72 h. After coculture, lymphocytes were analyzed for Bcl-2 expression using Western analysis and for tumor-induced apoptosis by terminal deoxynucleotidyl transferase-mediated nick end labeling. The role of SK-RC-45-stimulated caspase activation in degrading T-cell Bcl-2 was assessed using a pan-caspase inhibitor, as well as a specific inhibitor of caspase-9.
The renal cell carcinoma cell line SK-RC-45 sensitizes peripheral blood activated T lymphocytes and Jurkat cells to apoptosis by a mechanism that involves degradation of the antiapoptotic protein Bcl-2. The SK-RC-45-induced modulation of lymphocyte Bcl-2 levels was largely caspase independent because pretreatment of T cells with pan-caspase inhibitor III or an inhibitor of caspase-9 had minimal or no effect on stabilizing the protein, although it did provide protection against apoptosis. Overexpression of Bcl-2 protected Jurkat cells from tumor-mediated killing.
Bcl-2 inhibition is a mechanism by which tumors may render lymphocytes sensitive to other tumor-derived, proapoptotic stimuli.
肿瘤诱导T细胞凋亡是抑制抗肿瘤免疫的一种机制。通过共培养实验表明，肿瘤细胞系通过涉及线粒体通透性转换和细胞色素c释放的途径刺激T细胞凋亡。活化的T细胞表达大量的抗凋亡分子Bcl-2，预期该分子可赋予对这种肿瘤介导杀伤的抗性。我们研究了在暴露于肾肿瘤细胞系SK-RC-45的T细胞中Bcl-2失调的机制，并确定过表达Bcl-2是否能保护T细胞免受肿瘤介导的凋亡。
将转染或未转染Bcl-2的活化T淋巴细胞和Jurkat细胞暴露于SK-RC-45 48 - 72小时。共培养后，使用蛋白质免疫印迹分析淋巴细胞的Bcl-2表达，并通过末端脱氧核苷酸转移酶介导的缺口末端标记分析肿瘤诱导的凋亡。使用泛半胱天冬酶抑制剂以及半胱天冬酶-9的特异性抑制剂评估SK-RC-45刺激的半胱天冬酶活化在降解T细胞Bcl-2中的作用。
肾癌细胞系SK-RC-45通过涉及抗凋亡蛋白Bcl-2降解的机制使外周血活化T淋巴细胞和Jurkat细胞对凋亡敏感。SK-RC-45诱导的淋巴细胞Bcl-2水平调节在很大程度上不依赖于半胱天冬酶，因为用泛半胱天冬酶抑制剂III或半胱天冬酶-9抑制剂预处理T细胞对稳定该蛋白的作用很小或没有作用，尽管它确实提供了对凋亡的保护。Bcl-2的过表达保护Jurkat细胞免受肿瘤介导的杀伤。
Bcl-2抑制是肿瘤使淋巴细胞对其他肿瘤来源的促凋亡刺激敏感的一种机制。
