Knockdown of PSCA induces EMT and decreases metastatic potentials of the human prostate cancer DU145 cells.
Knockdown of PSCA induces EMT and decreases metastatic potentials of the human prostate cancer DU145 cells.
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PSCA 的敲除可诱导 EMT 并降低人前列腺癌 DU145 细胞的转移潜力。
DOI：
10.1186/s12935-016-0295-4
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发表时间：
2016
影响因子：
5.8
通讯作者：
Zhao Z
中科院分区：
文献类型：
--
作者：
Kang R;Zhao S;Liu L;Li F;Li E;Luo L;Xu L;Wan S;Zhao Z
前列腺干细胞抗原（PSCA）的表达已被证实与前列腺癌发生及前列腺癌（PCa）进展相关。目前，这些过程背后的机制尚不清楚。上皮 - 间充质转化（EMT）与前列腺癌的侵袭性和远处转移有关。在本研究中，我们在体外和体内研究了敲低PSCA对前列腺癌细胞系DU145的细胞迁移、侵袭能力以及EMT的影响。
设计了针对PSCA的4条小发夹RNA靶序列，筛选出敲低效果最佳的序列shRNA#1，构建稳定转染的DU145细胞系（DU145 shRNA#1），同时设计乱序序列构建稳定转染的DU145细胞系（DU145 scramble）。采用Transwell实验研究细胞迁移和侵袭能力。通过实时荧光定量逆转录聚合酶链反应（qRT - PCR）、蛋白质免疫印迹法（WB）对DU145、DU145 scramble、DU145 shRNA#1在体外和体内的PSCA、E - 钙黏蛋白、β - 连环蛋白、波形蛋白、纤连蛋白的表达进行定量分析。采用RT - PCR、免疫荧光染色对体外PSCA、E - 钙黏蛋白和波形蛋白的表达进行定量分析。通过qRT - PCR对体外EMT相关基因Snail、Slug和Twist进行定量分析。
构建的在DU145细胞中稳定敲低PSCA的细胞系具有高达90.5%的沉默效果。DU145 shRNA#1细胞从紧密排列的集落中分散开来。这与细胞迁移和侵袭能力下降有关。与体外和体内的DU145及DU145 scramble相比，在mRNA和蛋白质水平上，波形蛋白和纤连蛋白表达增加，E - 钙黏蛋白和β - 连环蛋白表达受到抑制。此外，除Snail外，EMT相关的Slug和Twist基因表达上调。
我们的数据表明，敲低PSCA可诱导DU145细胞发生EMT并降低其转移潜能，这表明PSCA在前列腺癌发生和进展中发挥着重要作用。
Prostate stem cell antigen (PSCA) expression has been shown to correlate with prostatic carcinogenesis and prostate cancer (PCa) progression. The underlying mechanisms for these processes are currently unknown. Epithelial to mesenchymal transition (EMT) has been associated with the invasiveness and the distant metastasis of PCa. In this study, we investigated the effects of knocking down the PSCA on the cell migration, the invasiveness, and the EMT of the PCa cell line DU145 in vitro and in vivo. Four target sequences of the small hairpin RNA for PSCA were designed, and the best effect knockdown sequence shRNA#1 was screened to construct the stable transfected DU145 cell line (DU145 shRNA#1), the scramble sequence was also designed to construct the stable transfected DU145 cell line(DU145 scramble). Cell migration and invasion were studied using Transwell assay. Quantitative RT-PCR, Western blot (WB) were used to quantify PSCA, E-cadherin, β-catenin, Vimentin, Fibronectin expression in DU145, DU145 scramble, DU145 shRNA#1 in vitro and in vivo. RT-PCR, immunofluorescent staining were used to quantify PSCA, E-cadherin, and Vimentin expression in vitro. EMT-related genes Snail, Slug, and Twist, were quantified by quantitative RT-PCR in vitro. The constructed stable knockdown of the PSCA in the DU145 cell had a silencing effect up to 90.5 %. DU145 shRNA#1 became scattered from the tightly packed colonies. It was associated with decreased cell migration and invasion. There was also an increased Vimentin and Fibronectin expression, an inhibited E-cadherin and β-catenin expression at both the mRNA and the protein levels when compared to the DU145 and the DU145 scramble in vitro and vivo. Furthermore, with the exception of the Snail, the expression of EMT-related Slug and Twist genes were upregulated. Our data indicated that knockdown of PSCA induced EMT and reduced metastatic potentials of the DU145 cells, suggesting that PSCA played an important role in prostatic carcinogenesis and progression.