Shi Xiaoyu, Peng Yanfei, Du Xiaoling, Liu Haitao, Klocker Helmut, Lin Qimei, Shi Jiandang, Zhang Ju
Bioactive Materials Key Lab of Ministry of Education, Department of Biochemistry and Molecular Biology, College of Life Sciences, Nankai University, Tianjin, 300071, China.
School of Integrative Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, 300193, China.
Prostate. 2017 Oct;77(14):1424-1437. doi: 10.1002/pros.23404. Epub 2017 Aug 29.
Epithelial-to-mesenchymal transition (EMT) is involved in pathogenesis of human benign prostatic hyperplasia (BPH). Estrogenic signaling pathways may stimulate the induction of EMT. However, the details of estradiol (E2) and estrogen receptors (ERs) effects on EMT, as well as E2-induced modulation of benign prostatic epithelial cell phenotype in vitro have not been completely clarified.
The effects of E2 on EMT markers and cytokeratins (CKs) expression were evaluated in benign epithelial cell lines BPH-1 and RWPE-1, which were cultured both in two-dimensional (2D) culture and three-dimensional (3D) culture model using hanging drop technique or 3D Matrigel model. ER antagonist, ICI182,780, was used to confirm the regulatory effects of E2 on EMT and phenotypic modulation. In 3D culture, immunohistochemical stainings were performed to detect the specific phenotype of cells that underwent EMT in acinar-like spheroids formed by RWPE-1. To illustrate the exact function of ERs in E2-induced EMT and phenotypic modulation, specific short interfering RNAs (siRNAs), and agonists were used to knockdown or activate individual ERs, respectively.
E2-induced EMT was observed both in 2D and 3D culture, with related regulation of EMT markers expression at both mRNA and protein level. In addition, E2 down-regulated luminal cell type markers CK18 and CK8 and up-regulated basal cell type markers CK5 and CK14. E2 also increased intermediate type markers CK15 and CK17, while it attenuated CK19 in 3D culture. ICI182,780 blocked E2-induced EMT and cell phenotypic switching. In 3D Matrigel culture, Vimentin was co-expressed with ERα and CK17, as well as with SMemb, which is related to cell status switching and proliferation. Knockdown of ERα but not GPR30 inhibited EMT, while ERβ knockdown facilitated EMT process. Knockdown of ERα blocked E2-induced EMT both in RWPE-1 and BPH-1. MRNA expression of EMT markers was stimulated by ERα-specific agonist PPT and inhibited by ERβ-specific agonist DPN.
Estrogenic effect mediated by ERα can promote EMT. E2 is also an inductive factor of cell phenotypic switching. Cell type modulation is associated with E2-induced EMT in benign prostatic epithelial cells. Taken together the results support a contribution of estrogens to the pathogenesis of BPH in elderly men.
上皮-间质转化（EMT）参与人类良性前列腺增生（BPH）的发病机制。雌激素信号通路可能刺激EMT的诱导。然而，雌二醇（E2）和雌激素受体（ERs）对EMT的影响细节，以及E2在体外诱导良性前列腺上皮细胞表型的调节作用尚未完全阐明。
在良性上皮细胞系BPH-1和RWPE-1中评估E2对EMT标志物和细胞角蛋白（CKs）表达的影响，这些细胞系在二维（2D）培养和使用悬滴技术或3D基质胶模型的三维（3D）培养模型中培养。使用ER拮抗剂ICI182,780来确认E2对EMT和表型调节的作用。在3D培养中，进行免疫组织化学染色以检测RWPE-1形成的腺泡样球体中发生EMT的细胞的特定表型。为了阐明ERs在E2诱导的EMT和表型调节中的具体功能，分别使用特异性短发夹RNA（siRNAs）和激动剂来敲低或激活单个ERs。
在2D和3D培养中均观察到E2诱导的EMT，在mRNA和蛋白质水平上均有相关的EMT标志物表达调节。此外，E2下调腔面细胞类型标志物CK18和CK8，并上调基底细胞类型标志物CK5和CK14。E2还增加中间类型标志物CK15和CK17，而在3D培养中减弱CK19。ICI182,780阻断E2诱导的EMT和细胞表型转换。在3D基质胶培养中，波形蛋白与ERα和CK17共表达，也与SMemb共表达，后者与细胞状态转换和增殖有关。敲低ERα而非GPR30可抑制EMT，而敲低ERβ则促进EMT进程。敲低ERα可在RWPE-1和BPH-1中均阻断E2诱导的EMT。EMT标志物的mRNA表达受到ERα特异性激动剂PPT的刺激，并受到ERβ特异性激动剂DPN的抑制。
由ERα介导的雌激素效应可促进EMT。E2也是细胞表型转换的诱导因子。细胞类型调节与E2诱导的良性前列腺上皮细胞EMT相关。综合这些结果支持雌激素对老年男性BPH发病机制的作用。
