Muñoz-Moreno Laura, Bajo Ana M, Prieto Juan C, Carmena María J
Department of Systems Biology, Unit of Biochemistry and Molecular Biology, University of Alcalá, Alcalá de Henares 28871, Spain.
Department of Systems Biology, Unit of Biochemistry and Molecular Biology, University of Alcalá, Alcalá de Henares 28871, Spain.
Mol Cell Endocrinol. 2017 May 5;446:59-69. doi: 10.1016/j.mce.2017.02.011. Epub 2017 Feb 11.
The involvement of growth hormone-releasing hormone (GHRH) in several relevant processes that contribute to prostate cancer progression was analyzed. Firstly, we evaluated GHRH effects on cell proliferation and adhesion in human cancer prostate cell lines, LNCaP and PC3, by using specific assays (BrdU incorporation and collagen adhesion). The expression levels of the main marker molecules of these processes were measured by RT-PCR, Western blotting and zymography assays. GHRH increased both cell proliferation and proliferating cell nuclear antigen (PCNA) levels in LNCaP cells and in PC3 cells; however, such a rise was faster in the PC3 cells that represent the most aggressive stage of prostate cancer. Furthermore, GHRH significantly reduced cell adhesion and E-cadherin levels in LNCaP and PC3 cells and up-regulated the total and nuclear expression of β-catenin in PC3 cells. In addition, we assessed cell cycle, cell migration and VEGF secretion in PC3 cells. GHRH augmented the number of cells in G2/M-phase but diminished that corresponding to G1-phase. Cell-cycle specific markers were evaluated since GHRH effects may be related to their differential expression; we observed a decrease of p53, p21, and Bax/Bcl2 ratio. Furthermore, GHRH increased the expression of CD44, c-myc and cyclin D1, MMP-2 and MMP-9 activity, and VEGF secretion. We also observed that EGFR and/or HER2 transactivation is involved in cell adhesion, cell migration and VEGF secretion produced by GHRH. Consequently, present results define GHRH as a proliferative, anti-apoptotic and migratory agent in prostate cancer.
分析了生长激素释放激素（GHRH）在促成前列腺癌进展的几个相关过程中的作用。首先，我们通过使用特定检测方法（BrdU掺入法和胶原黏附实验）评估了GHRH对人前列腺癌细胞系LNCaP和PC3中细胞增殖和黏附的影响。通过逆转录聚合酶链反应（RT-PCR）、蛋白质免疫印迹法和酶谱分析检测了这些过程中主要标志物分子的表达水平。GHRH增加了LNCaP细胞和PC3细胞中的细胞增殖及增殖细胞核抗原（PCNA）水平；然而，在代表前列腺癌最侵袭性阶段的PC3细胞中，这种升高更快。此外，GHRH显著降低了LNCaP和PC3细胞中的细胞黏附及E-钙黏蛋白水平，并上调了PC3细胞中β-连环蛋白的总表达和核表达。此外，我们评估了PC3细胞中的细胞周期、细胞迁移和血管内皮生长因子（VEGF）分泌情况。GHRH增加了处于G2/M期的细胞数量，但减少了处于G1期的细胞数量。由于GHRH的作用可能与其差异表达有关，因此对细胞周期特异性标志物进行了评估；我们观察到p53、p21以及Bax/Bcl2比值降低。此外，GHRH增加了CD44、c-myc和细胞周期蛋白D1的表达、基质金属蛋白酶-2（MMP-2）和基质金属蛋白酶-9（MMP-9）的活性以及VEGF分泌。我们还观察到表皮生长因子受体（EGFR）和/或人表皮生长因子受体2（HER2）的反式激活参与了GHRH所产生的细胞黏附、细胞迁移和VEGF分泌过程。因此，目前的研究结果将GHRH定义为前列腺癌中的一种增殖、抗凋亡和迁移因子。
