Istituto di Patologia Medica, Università Cattolica del Sacro Cuore, 00168 Roma, Italy.
Institute of Cell Biology and Neurobiology, National Research Council, 00168 Rome, Italy.
Int J Mol Sci. 2019 Aug 17;20(16):4012. doi: 10.3390/ijms20164012.
Estrogen and hypoxia promote an aggressive phenotype in prostate cancer (PCa), driving transcription of progression-associated genes. Here, we molecularly dissect the contribution of long non-coding RNA H19 to PCa metastatic potential under combined stimuli, a topic largely uncovered. The effects of estrogen and hypoxia on H19 and cell adhesion molecules' expression were investigated in PCa cells and PCa-derived organotypic slice cultures (OSCs) by qPCR and Western blot. The molecular mechanism was addressed by chromatin immunoprecipitations, overexpression, and silencing assays. PCa cells' metastatic potential was analyzed by in vitro cell-cell adhesion, motility test, and trans-well invasion assay. We found that combined treatment caused a significant H19 down-regulation as compared with hypoxia. In turn, H19 acts as a transcriptional repressor of cell adhesion molecules, as revealed by up-regulation of both β3 and β4 integrins and E-cadherin upon H19 silencing or combined treatment. Importantly, H19 down-regulation and β integrins induction were also observed in treated OSCs. Combined treatment increased both cell motility and invasion of PCa cells. Lastly, reduction of β integrins and invasion was achieved through epigenetic modulation of H19-dependent transcription. Our study revealed that estrogen and hypoxia transcriptionally regulate, via H19, cell adhesion molecules redirecting metastatic dissemination from EMT to a β integrin-mediated invasion.
雌激素和缺氧促进前列腺癌（PCa）的侵袭表型，驱动进展相关基因的转录。在这里，我们从分子水平上剖析了长链非编码 RNA H19 在联合刺激下对 PCa 转移潜能的贡献，这是一个很大程度上尚未被揭示的课题。通过 qPCR 和 Western blot 研究了雌激素和缺氧对 PCa 细胞和 PCa 衍生的器官型切片培养物（OSC）中 H19 和细胞黏附分子表达的影响。通过染色质免疫沉淀、过表达和沉默实验探讨了分子机制。通过体外细胞-细胞黏附、迁移试验和 Transwell 侵袭试验分析了 PCa 细胞的转移潜能。我们发现，与缺氧相比，联合处理导致 H19 显著下调。反过来，H19 作为细胞黏附分子的转录抑制因子发挥作用，这是通过沉默 H19 或联合处理后β3 和β4 整合素和 E-钙黏蛋白的上调来揭示的。重要的是，在处理的 OSC 中也观察到 H19 下调和β整合素诱导。联合处理增加了 PCa 细胞的迁移和侵袭。最后，通过 H19 依赖性转录的表观遗传调控，实现了β整合素的减少和侵袭的降低。我们的研究表明，雌激素和缺氧通过 H19 转录调控细胞黏附分子，将转移扩散从 EMT 重定向到β整合素介导的侵袭。
