Chen Fanghong, Zhang Guanjian, Cao Yanli, Payne Ryan, See William A
Department of Urology, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
J Urol. 2007 Nov;178(5):2166-70. doi: 10.1016/j.juro.2007.06.040. Epub 2007 Sep 17.
By inducing cell cycle arrest at the G1/S transition bacillus Calmette-Guerin has been shown to have a direct antiproliferative effect on urothelial carcinoma cell lines. In other systems cell cycle arrest has been shown to confer a relative state of apoptotic resistance. We assessed the effect of bacillus Calmette-Guerin on the susceptibility of urothelial carcinoma cells to apoptotic stimuli.
The human UC cell lines T24 and 253J (American Type Cell Culture, Rockville, Maryland) were used to evaluate the effect of bacillus Calmette-Guerin or antibody mediated alpha5beta1cross-linking on apoptosis and apoptotic sensitivity. Following treatment baseline apoptosis and the response to the apoptotic inducing agent camptothecin was evaluated using assays for caspase 3 activation and DNA fragmentation. Pharmacological blockade of signaling pathways known to be activated in response to bacillus Calmette-Guerin/alpha5beta1 cross-linking was used to assess the role of these pathways in the bacillus Calmette-Guerin apoptotic response. A final series of experiments used the MTT assay to study the impact of bacillus Calmette-Guerin pretreatment on the cytotoxicity of the antineoplastic agent mitomycin C.
Treatment with bacillus Calmette-Guerin failed to induce apoptosis, as measured by caspase 3 activation or DNA laddering. Bacillus Calmette-Guerin pretreatment significantly inhibited the induction of apoptosis in response to camptothecin. These effects were reproduced by antibody mediated cross-linking of alpha5beta1 integrin. Pharmacological inhibition of nuclear factor kappaB and/or AP1 signaling pathways reversed the anti-apoptotic effect of bacillus Calmette-Guerin. Mitomycin C cytotoxicity was significantly decreased by bacillus Calmette-Guerin pretreatment.
Bacillus Calmette-Guerin exerts a direct anti-apoptotic effect on human urothelial carcinoma cell lines. The ability of antibody mediated cross-linking to reproduce the effect and the ability of signal transduction inhibitors to block it are consistent with a mechanism involving integrin mediated signaling. Apoptotic resistance represents a therapeutic target for modulating the response to bacillus Calmette-Guerin and it may have clinical implications in the sequencing of intravesical therapies.
通过诱导细胞周期在G1/S期转换时停滞，卡介苗已被证明对膀胱癌细胞系具有直接的抗增殖作用。在其他系统中，细胞周期停滞已被证明可赋予相对的抗凋亡状态。我们评估了卡介苗对膀胱癌细胞对凋亡刺激敏感性的影响。
使用人膀胱癌细胞系T24和253J（美国典型培养物保藏中心，马里兰州罗克维尔）来评估卡介苗或抗体介导的α5β1交联对凋亡及凋亡敏感性的影响。处理后，使用半胱天冬酶3激活和DNA片段化检测来评估基线凋亡及对凋亡诱导剂喜树碱的反应。对已知在卡介苗/α5β1交联反应中被激活的信号通路进行药理学阻断，以评估这些通路在卡介苗凋亡反应中的作用。最后一系列实验使用MTT检测来研究卡介苗预处理对抗肿瘤药物丝裂霉素C细胞毒性的影响。
通过半胱天冬酶3激活或DNA梯状条带检测，卡介苗处理未能诱导凋亡。卡介苗预处理显著抑制了对喜树碱诱导凋亡的反应。这些效应通过抗体介导的α5β1整合素交联得以重现。对核因子κB和/或AP1信号通路的药理学抑制逆转了卡介苗的抗凋亡作用。卡介苗预处理显著降低了丝裂霉素C的细胞毒性。
卡介苗对人膀胱癌细胞系发挥直接的抗凋亡作用。抗体介导的交联能够重现该效应以及信号转导抑制剂能够阻断该效应，这与涉及整合素介导信号传导的机制相一致。抗凋亡作用代表了调节对卡介苗反应的一个治疗靶点，并且它可能在膀胱内治疗的顺序安排中具有临床意义。
