Department of Surgery , University of Michigan Medical Center , Ann Arbor , Michigan 48109 , United States.
Department of Radiation Oncology , University of Michigan , Ann Arbor , Michigan 48109 , United States.
J Proteome Res. 2018 Apr 6;17(4):1690-1699. doi: 10.1021/acs.jproteome.8b00014. Epub 2018 Mar 9.
Circulating microvesicles are able to mediate long-distance cell-cell communications. It is essential to understand how microvesicles from pancreatic cancer act on other cells in the body. In this work, serum-derived microvesicles were isolated from 10 patients with locally advanced pancreatic cancer and healthy controls. Using Cell Transwell and WST-1 reagents, we found that microvesicles from pancreatic cancer accelerated migration and proliferation of PANC-1 cells. Meanwhile, the proliferation of these cancer-microvesicle-treated cells (CMTCs) was affected less by 10 μM of gemcitabine relative to healthy microvesicle-treated cells (HMTCs). Next, we optimized the filter-aided sample preparation method to increase the recovery of protein samples and then applied it to the quantification of the proteome of CMTCs and HMTCs. The peptides were labeled and analyzed by liquid chromatography-tandem mass spectrometry. In total, 4102 proteins were identified, where 35 proteins were up-regulated with 27 down-regulated in CMTCs. We verified the quantitative results of three key proteins CD44, PPP2R1A, and TP53 by Western blot. The Ingenuity Pathway Analysis revealed pathways that cancer microvesicles might participate in to promote cell migration and proliferation. These findings may provide novel clues of treatment for tumorigenesis and metastasis.
循环微泡能够介导远距离的细胞间通讯。了解胰腺癌来源的微泡如何作用于体内其他细胞是至关重要的。在这项工作中,我们从 10 例局部晚期胰腺癌患者和健康对照者的血清中分离出了血清来源的微泡。通过细胞 Transwell 和 WST-1 试剂,我们发现胰腺癌来源的微泡加速了 PANC-1 细胞的迁移和增殖。同时,与健康微泡处理的细胞(HMTCs)相比,这些经癌症微泡处理的细胞(CMTCs)的增殖对 10μM 的吉西他滨的影响较小。接下来,我们优化了滤过辅助样品制备方法以增加蛋白质样品的回收率,然后将其应用于 CMTCs 和 HMTCs 的蛋白质组定量分析。用液相色谱-串联质谱法对肽进行标记和分析。共鉴定到 4102 种蛋白质,其中 35 种在 CMTCs 中上调,27 种下调。我们通过 Western blot 验证了三个关键蛋白 CD44、PPP2R1A 和 TP53 的定量结果。通路分析揭示了癌症微泡可能参与的促进细胞迁移和增殖的途径。这些发现可能为肿瘤发生和转移的治疗提供新的线索。