Liu J R, Fletcher B, Page C, Hu C, Nunez G, Baker V
Division of Gynecologic Oncology, University of Michigan, Ann Arbor, Michigan, 48109, USA.
Gynecol Oncol. 1998 Sep;70(3):398-403. doi: 10.1006/gyno.1998.5125.
To investigate the role of Bcl-xL in resistance to chemotherapy-induced apoptosis in ovarian carcinoma.
Two human ovarian carcinoma cell lines were used in this study: A2780 and SKOV3. A2780 cells were transfected with human Bcl-xL or control plasmid alone. Expression of Bcl-xL in single cell clones was analyzed by flow cytometry and protein expression was confirmed by Western blot. For in vitro chemotherapy-induced death assays, cisplatin and Taxol were used. The percentage of apoptotic cells was determined by nuclear propidium iodide staining followed by flow cytometric analysis. Human ascites samples were used to make tumor lysates which were then analyzed for expression of Bcl-xL protein by Western blot.
A2780 cells express low levels of endogenous Bcl-xL while SKOV3 cells express high amounts as determined by Western blot. In addition, A2780 cells are sensitive to chemotherapy-induced cell death while SKOV3 cells are resistant. To determine if chemoresistance is mediated by expression of Bcl-xL, A2780 cells were transfected with Bcl-xL or control plasmid. Cells were incubated with either cisplatin or Taxol to induce apoptosis. Bcl-xL-expressing cells were highly resistant to cisplatin and Taxol compared with controls (P < 0.05). All samples of malignant ascites analyzed expressed high levels of Bcl-xL on Western blot.
The results of these studies indicate that Bcl-xL is expressed in ovarian carcinoma, and in A2780 cells functions in a manner analogous to Bcl-2 by inhibiting chemotherapy-induced apoptosis. This may have prognostic significance; previous studies have demonstrated that patients with breast cancer that overexpress Bcl-2 have low-grade, hormonally responsive tumors. In ovarian carcinoma, another Bcl-2 family member, Bcl-xL may be responsible for modulating resistance to chemotherapy-induced apoptosis.
探讨Bcl-xL在卵巢癌化疗诱导凋亡抗性中的作用。
本研究使用了两个人卵巢癌细胞系：A2780和SKOV3。A2780细胞分别转染人Bcl-xL或对照质粒。通过流式细胞术分析单细胞克隆中Bcl-xL的表达，并通过蛋白质印迹法确认蛋白质表达。在体外化疗诱导死亡试验中，使用顺铂和紫杉醇。通过碘化丙啶核染色随后进行流式细胞术分析来确定凋亡细胞的百分比。用人腹水样本制备肿瘤裂解物，然后通过蛋白质印迹法分析Bcl-xL蛋白的表达。
蛋白质印迹法测定显示，A2780细胞内源性Bcl-xL表达水平低，而SKOV3细胞表达量高。此外，A2780细胞对化疗诱导的细胞死亡敏感，而SKOV3细胞具有抗性。为了确定化疗抗性是否由Bcl-xL的表达介导，将A2780细胞转染Bcl-xL或对照质粒。细胞与顺铂或紫杉醇孵育以诱导凋亡。与对照相比，表达Bcl-xL的细胞对顺铂和紫杉醇具有高度抗性（P < 0.05）。蛋白质印迹法分析的所有恶性腹水样本均表达高水平的Bcl-xL。
这些研究结果表明，Bcl-xL在卵巢癌中表达，并且在A2780细胞中通过抑制化疗诱导的凋亡发挥类似于Bcl-2的作用。这可能具有预后意义；先前的研究表明，过表达Bcl-2的乳腺癌患者患有低级别、激素反应性肿瘤。在卵巢癌中，另一个Bcl-2家族成员Bcl-xL可能负责调节对化疗诱导凋亡的抗性。
