布雷菲德菌素A通过线粒体凋亡途径增强顺铂抗肺癌GLC-82细胞的作用

本研究证明了线粒体凋亡途径在布雷菲德菌素A（brefeldin A，BFA）联合顺铂（cis-dichlorodiamine platinum，CDDP）抗非小细胞肺癌（non-small cell lung cancer，NSCLC）中的作用。MTT结果显示，BFA对肺癌GLC-82和NCI-H1299细胞的半数有效抑制浓度（half maximal inhibitory concentration，IC50）分别是100 ng/mL和400 ng/mL，CDDP对GLC-82和NCI-H1299细胞的IC50分别是4 μg/mL和15 μg/mL；而分别采用半量的BFA和CDDP联合处理GLC-82或NCI-H1299细胞后，抑制作用均进一步加强。DAPI染色结果进一步证明了二者的协同作用——与单独用药组相比，细胞核染色质固缩加剧，核裂解碎片增多，乃至形成凋亡小体，表明细胞凋亡的发生。与单药组比较，联合用药导致肺癌GLC-82细胞线粒体膜电位显著下降；q-RT-PCR及Western印迹结果显示，在联合用药早期（24 h），GLC-82细胞可能通过提高Bcl2表达以促进存活；而在联合用药晚期（48 h），细胞已发生不可逆转的凋亡，Bcl2表达受抑制，同时二者通过促进Bax表达来诱导细胞色素C释放，使胱天蛋白酶 3发生剪切激活，最终诱导细胞凋亡发生。提示线粒体凋亡途径可能是BFA协同CDDP抗非小细胞肺癌的分子机制之一，为肺癌的临床治疗方案提供了更多的理论依据。

Enhancement of Cis-dichlorodiamine Platinum Against Lung Cancer Cells Through Mitochondrial Apoptosis Pathway by Brefeldin A

In this study，we investigated the involvement of mitochondrial apoptosis pathway in the synergistic anti-cancer effect of brefeldin A (BFA) and cis-dichlorodiamine platinum (CDDP) on the non-small cell lung cancer (NSCLC). MTT assays revealed that BFA alone inhibited the proliferation of GLC-82 cells or NCI-H1299 cells with an IC50 of 100 ng/mL or 400 ng/mL respectively, and CDDP alone inhibited the proliferation of GLC-82 cells or NCI-H1299 cells with an IC50 of 4 μg/mL or 15 μg/mL respectively. The inhibitory effect was further strengthened with half doses of BFA and CDDP in combination. With combined exposure of drugs, more abnormal cells with chromatin condensation were observed, together with increased numbers of apoptotic bodies with nucleus and cytoplasm fragmentation. Compared to single drug exposures, the mitochondrial membrane potential of GLC-82 cells decreased significantly. qRT-PCR and Western blotting results showed that cells may improve survival by increasing the expression level of Bcl2 at the early stage (24 h), and an irreversible apoptosis at the late stage (48 h) when the expression level of Bcl2 was inhibited. At the same time, BFA and CDDP induced the release of Cyt C by promoting the expression level of Bax, which induced the shear activation of Caspase 3 and eventually induced apoptosis. These data suggested that the mitochondrial apoptosis pathway may be one of the molecular mechanisms of BFA in combination with CDDP against human lung cancer cells. Our work thus provide a more theoretical basis for the clinical treatment of NSCLC.