Integrin β4 and vinculin contained in exosomes are potential markers for progression of prostate cancer associated with taxane-resistance
Integrin β4 and vinculin contained in exosomes are potential markers for progression of prostate cancer associated with taxane-resistance
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DOI：
10.3892/ijo.2015.3011
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发表时间：
2015-07-01
影响因子：
5.2
通讯作者：
Ito, Masafumi
中科院分区：
文献类型：
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作者：
Kawakami, Kyojiro;Fujita, Yasunori;Ito, Masafumi
用紫杉烷治疗去势抵抗的前列腺癌通常会导致耐药性的产生。最近的研究表明，体液中存在的外切体在其来源的细胞中含有蛋白质和RNA，可以作为各种疾病的诊断标记物。在目前的研究中，我们的目标是确定外切体中包含的蛋白质，这些蛋白质可能是前列腺癌进展和紫杉烷耐药的标志。用差速离心法从耐紫杉烷的人前列腺癌PC-3细胞(PC-3R)及其亲本PC-3细胞的培养液中分离到外切体。分离的外切体进行基于iTRAQ的定量蛋白质组学分析。用抗CD9抗体标记的磁珠从培养上清液中分离出外切体。通过siRNA转染下调蛋白质表达，然后分析沉默效果。蛋白质组学分析表明，与PC-3细胞相比，PC-3R细胞外切体中整合素β4(ITGB4)和纽蛋白(VCL)表达上调。抗CD9抗体从PC-3R细胞培养上清液中捕获的外切体中ITGB4和VCL水平明显升高。抑制ITGB4和VCL的表达不影响PC-3R细胞的增殖和紫杉烷耐药性，但ITGB4基因敲除可减弱细胞的迁移和侵袭，VCL基因敲除可减少侵袭。我们的结果表明，外切体中的ITGB4和VCL可能是与紫杉烷耐药相关的前列腺癌进展的有用标记物，为建立基于外切体的诊断系统提供了基础。
Treatment with taxanes for castration-resistant prostate cancer often leads to the development of resistance. It has been recently demonstrated that exosomes present in the body fluids contain proteins and RNAs in the cells from which they are derived and could serve as a diagnostic marker for various diseases. In the present study, we aimed to identify proteins contained in exosomes that could be markers for progression and taxane-resistance of prostate cancer. Exosomes were isolated by differential centrifugation from the culture medium of taxane-resistant human prostate cancer PC-3 cells (PC-3R) and their parental PC-3 cells. Isolated exosomes were subjected to iTRAQ-based quantitative proteomic analysis. Exosomes were also isolated from the culture medium by using anti-CD9 antibody-conjugated magnetic beads. Protein expression was knocked down by siRNA transfection followed by analysis of the silencing effects. Proteomic analysis showed that integrin beta 4 (ITGB4) and vinculin (VCL) were upregulated in exosomes derived from PC-3R cells compared to PC-3 cells. The elevation of ITGB4 and VCL was confirmed in exosomes captured by anti-CD9 antibody from the culture medium of PC-3R cells. Silencing of ITGB4 and VCL expression did not affect proliferation and taxane-resistance of PC-3R cells, but ITGB4 knockdown attenuated both cell migration and invasion and VCL knockdown reduced invasion. Our results suggest that ITGB4 and VCL in exosomes could be useful markers for progression of prostate cancer associated with taxane-resistance, providing the basis for development of an exosome-based diagnostic system.