PURPOSE Recent studies have shown that 20-hydroxyeicosatetraenoic acid (20-HETE) is a key molecule in sustaining androgen-mediated prostate cancer cell survival. Thus, the aim of this study was to determine whether 20-HETE can affect the metastatic potential of androgen-insensitive prostate cancer cells, and the implication of the newly described 20-HETE receptor, GPR75, in mediating these effects. METHODS The expression of GPR75, protein phosphorylation, actin polymerization and protein distribution were assessed by western blot and/or fluorescence microscopy. Additionally, in vitro assays including epithelial-mesenchymal transition (EMT), metalloproteinase-2 (MMP-2) activity, scratch wound healing, transwell invasion and soft agar colony formation were used to evaluate the effects of 20-HETE agonists/antagonists or GPR75 gene silencing on the aggressive features of PC-3 cells. RESULTS 20-HETE (0.1 nM) promoted the acquisition of a mesenchymal phenotype by increasing EMT, the release of MMP-2, cell migration and invasion, actin stress fiber formation and anchorage-independent growth. Also, 20-HETE augmented the expression of HIC-5, the phosphorylation of EGFR, NF-κB, AKT and p-38 and the intracellular redistribution of p-AKT and PKCα. These effects were impaired by GPR75 antagonism and/or silencing. Accordingly, the inhibition of 20-HETE formation with N-hydroxy-N'-(4-n-butyl-2-methylphenyl) formamidine (HET0016) elicited the opposite effects. CONCLUSIONS The present results show for the first time the involvement of the 20-HETE-GPR75 receptor in the activation of intracellular signaling known to be stimulated in cell malignant transformations leading to the differentiation of PC-3 cells towards a more aggressive phenotype. Targeting the 20-HETE/GPR75 pathway is a promising and novel approach to interfere with prostate tumor cell malignant progression.

中文翻译：

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GPR75受体介导雄激素不敏感的前列腺癌细胞的20-HETE信号和转移特征。

目的最近的研究表明20-羟基二十碳四烯酸（20-HETE）是维持雄激素介导的前列腺癌细胞存活的关键分子。因此，本研究的目的是确定20-HETE是否可以影响雄激素不敏感的前列腺癌细胞的转移潜能，以及新描述的20-HETE受体GPR75在介导这些作用中的意义。方法采用Western blot和/或荧光显微镜观察GPR75的表达，蛋白磷酸化，肌动蛋白聚合和蛋白分布。此外，体外测定包括上皮-间质转化（EMT），金属蛋白酶2（MMP-2）活性，刮擦伤口愈合，使用transwell侵袭和软琼脂集落形成来评估20-HETE激动剂/拮抗剂或GPR75基因沉默对PC-3细胞侵袭性特征的影响。结果20-HETE（0.1 nM）通过增加EMT，释放MMP-2，细胞迁移和侵袭，肌动蛋白应激纤维形成和不依赖锚固的生长促进了间充质表型的获得。同样，20-HETE增强了HIC-5的表达，EGFR，NF-κB，AKT和p-38的磷酸化以及p-AKT和PKCα的细胞内重新分布。GPR75拮抗作用和/或沉默削弱了这些作用。因此，用N-羟基-N'-（4-正丁基-2-甲基苯基）甲idine（HET0016）抑制20-HETE形成引起相反的作用。结论本结果首次显示20-HETE-GPR75受体参与已知在细胞恶性转化中刺激的细胞内信号传导的激活，导致PC-3细胞向更具攻击性的表型分化。靶向20-HETE / GPR75途径是干扰前列腺肿瘤细胞恶性进展的一种有前途的新颖方法。