Beck T P, Kirsh E J, Chmura S J, Kovar D A, Chung T, Rinker-Schaeffer C W, Stadler W M
Department of Surgery, University of Chicago Pritzker School of Medicine, Illinois 60637, USA.
Urology. 1999 Sep;54(3):573-7. doi: 10.1016/s0090-4295(99)00210-1.
Calphostin C, a highly specific protein kinase C inhibitor, induces apoptosis in the presence of visible light. We report the photoactivatable cytotoxicity of calphostin C in a series of well-characterized human bladder cancer cell lines: RT4, UM-UC-3, and 5637.
The human bladder cancer cell lines RT4, UM-UC-3, and 5637 were chosen on the basis of their p53, pRb and 9p21 deletion status. Using standard tissue culture techniques, the cytotoxicity of 10 to 100 nM calphostin C in combination with increasing exposures of visible light was examined. Controls consisted of cells treated with calphostin C without visible light and cells exposed to visible light without calphostin C treatment. Cell viability was determined by MTT assay. The induction of apoptosis by activated calphostin C was determined by 4,6-diamidino-2-phenylindole (DAPI) staining/fluorescence microscopy of nuclei.
In the absence of light, calphostin C did not demonstrate a cytotoxic effect on any of the cell lines tested. Increasing the duration of light exposure resulted in a concomitant decrease in cell viability. Significant cell death was seen with calphostin C concentrations as low as 10 nM. These studies also demonstrated that calphostin C induced apoptosis by a mechanism independent of p53 and pRb status and the presence or absence of 9p21 deletions.
We demonstrated the ability of activated calphostin C to induce apoptosis in a light-dependent and concentration-dependent fashion in a bladder cancer model system. Activated calphostin C cytotoxicity is independent of tumor genetic background and the status of p53 and pRb. Further development of calphostin C as a photosensitizer for photodynamic therapy of superficial bladder cancer may be warranted.
钙泊三醇C是一种高度特异性的蛋白激酶C抑制剂，在可见光存在的情况下可诱导细胞凋亡。我们报告了钙泊三醇C在一系列特征明确的人膀胱癌细胞系（RT4、UM-UC-3和5637）中的光激活细胞毒性。
根据人膀胱癌细胞系RT4、UM-UC-3和5637的p53、pRb和9p21缺失状态进行选择。采用标准组织培养技术，检测了10至100 nM钙泊三醇C与增加的可见光照射相结合时的细胞毒性。对照组包括未接受可见光处理的钙泊三醇C处理细胞和未接受钙泊三醇C处理的可见光照射细胞。通过MTT法测定细胞活力。通过细胞核的4,6-二脒基-2-苯基吲哚（DAPI）染色/荧光显微镜检查活化的钙泊三醇C诱导的细胞凋亡。
在无光条件下，钙泊三醇C对任何测试细胞系均未表现出细胞毒性作用。增加光照时间导致细胞活力随之降低。钙泊三醇C浓度低至10 nM时可见明显的细胞死亡。这些研究还表明，钙泊三醇C通过一种独立于p53和pRb状态以及9p21缺失与否的机制诱导细胞凋亡。
我们证明了在膀胱癌模型系统中，活化的钙泊三醇C能够以光依赖和浓度依赖的方式诱导细胞凋亡。活化的钙泊三醇C细胞毒性独立于肿瘤遗传背景以及p53和pRb状态。钙泊三醇C作为浅表性膀胱癌光动力治疗的光敏剂可能值得进一步开发。
