多西紫杉醇诱导前列腺癌PC-3细胞自噬的研究

Research of autophagy induced by docetaxel in prostate cancer cell line PC-3

摘要:

目的：观察多西紫杉醇是否诱导前列腺癌PC-3细胞产生自噬，并初步探讨其可能机制。方法：实验应用MTT法测定细胞的增殖率；透射电镜观察细胞超微结构的变化；免疫荧光和Western blot分别定性和定量检测自噬相关蛋白Ⅱ型微管相关蛋白轻链3蛋白（microtubule-associated protein 1 light chain 3 typeⅡ，LC3-Ⅱ）、底物蛋白（sequestosome 1/SQSTM1，P62）表达水平，RT-PCR检测自噬相关基因Beclin-1 mRNA。结果：MTT结果显示多西紫杉醇能有效抑制PC-3细胞增殖（F=58.684，P=0.000）；10-6 mol/L多西紫杉醇处理PC-3细胞24 h，即可发生明显的自噬，透射电镜观察到自噬体双层膜结构的形成。免疫荧光和Western blot结果显示多西紫杉醇处理PC-3细胞后P62表达降低（P=0.003、P=0.000、P=0.000），LC3-Ⅱ表达增强（P=0.002、P=0.000、P=0.000），RT-PCR结果显示多西紫杉醇处理PC-3细胞后可引起细胞Beclin-1 mRNA水平的上调（P=0.000、P=0.000、P=0.000）。结论：多西紫杉醇可以有效抑制前列腺癌PC-3细胞的增殖并诱导自噬，其发生可能与通过上调Beclin-1的mRNA水平有关。

Abstract:

Objective：To investigate the autophagy induced by docetaxel in prostate cancer cell line PC-3 and to explore its possible mechanism. Methods：PC-3 cells cultured in vitro were treated with docetaxel. Proliferation inhibition rate of PC-3 cells was assayed by MTT；ultra-structural cellular changes were observed by transmission electron microscope（TEM）；levels of sequestosome 1/SQSTM1 （P６２） and microtubule-associated protein 1 light chain 3 typeⅡ（LC-3Ⅱ） protein were detected by Immunofluorescence and Western blot，qualitatively and quantitatively；level of mRNA of autophagy gene Beclin-1 was detected by RT-PCR. Results：After docetaxel treatment，proliferation of PC-3 cells was inhibited effectively（F=58.684，P=0.000）. Autophagy could be observed in prostate cancer PC-3 cells treated with 10-6 mol/L docetaxel after 24 h. TEM examination revealed the appearance of autophagosome with double-membrane structure. Immunofluorescence and Western blot showed that expression of LC-3Ⅱ was increased（P=0.002，P=0.000，P=0.000） in PC-3 cells after being treated with docetaxel and expression of P62 was decreased（P=0.003，P=0.000，P=0.000）. RT-PCR showed that levels of mRNA of Beclin-1 were increased by docetaxel（P=0.000，P=0.000，P=0.000）. Conclusion：Docetaxel could inhibit the growth of PC-3 cells and induce the appearance of autophagy in PC-3 cells，and the possible mechanism of autophagy induced by docetaxel is related to the increase of Beclin-1 mRNA.