Department of Biological Sciences, Clark Atlanta University, Atlanta, Georgia 30314, USA.
Prostate. 2010 Jun 15;70(9):982-92. doi: 10.1002/pros.21132.
Snail transcription factor induces epithelial-mesenchymal transition (EMT) via decreased cell adhesion-associated molecules like E-cadherin, and increased mesenchymal markers like vimentin. We previously established Snail-mediated EMT model utilizing androgen-dependent LNCaP cells. These cells express increased vimentin protein and relocalization of E-cadherin from the cell membrane to the cytosol. Interestingly, Snail transfection in LNCaP cells resulted in cells acquiring a neuroendocrine (NE)-like morphology with long neurite-like processes.
We tested for expression of NE markers neuron-specific enolase (NSE) and chromogranin A (CgA) by Western blot analysis, and performed proliferation assays to test for paracrine cell proliferation.
LNCaP cells transfected with Snail displayed increase in the NE markers, NSE and CgA as well as translocation of androgen receptor (AR) to the nucleus. LNCaP C-33 cells that have been previously published as a neuroendocrine differentiation (NED) model exhibited increased expression levels of Snail protein as compared to LNCaP parental cells. Functionally, conditioned medium from the LNCaP-Snail transfected cells increased proliferation of parental LNCaP and PC-3 cells, which could be abrogated by NSE/CgA siRNA. Additionally, NED in LNCaP-C33 cells or that induced in parental LNCaP cells by serum starvation could be inhibited by knockdown of Snail with siRNA.
Overall our data provide evidence that Snail transcription factor may promote tumor aggressiveness in the LNCaP cells through multiple processes; induction of EMT may be required to promote migration, while NED may promote tumor proliferation by a paracrine mechanism. Therefore, therapeutic targeting of Snail may prove beneficial in not only abrogating EMT but also NED.
蜗牛转录因子通过降低细胞黏附相关分子如 E-钙黏蛋白，增加间充质标志物如波形蛋白，诱导上皮-间充质转化（EMT）。我们之前利用雄激素依赖性 LNCaP 细胞建立了蜗牛介导的 EMT 模型。这些细胞表达增加的波形蛋白蛋白和 E-钙黏蛋白从细胞膜到细胞质的重新定位。有趣的是，Snail 转染 LNCaP 细胞导致细胞获得具有长神经突样过程的神经内分泌（NE）样形态。
我们通过 Western blot 分析测试了神经内分泌标志物神经元特异性烯醇化酶（NSE）和嗜铬粒蛋白 A（CgA）的表达，并进行了增殖实验以测试旁分泌细胞增殖。
转染 Snail 的 LNCaP 细胞显示出 NE 标志物 NSE 和 CgA 的增加以及雄激素受体（AR）向核内易位。先前已发表为神经内分泌分化（NED）模型的 LNCaP C-33 细胞与 LNCaP 亲本细胞相比，Snail 蛋白表达水平增加。功能上，来自转染 Snail 的 LNCaP 细胞的条件培养基增加了亲本 LNCaP 和 PC-3 细胞的增殖，这可以通过 NSE/CgA siRNA 来阻断。此外，通过 siRNA 敲低 Snail 可以抑制 LNCaP-C33 细胞中的 NED 或通过血清饥饿诱导的亲本 LNCaP 细胞中的 NED。
总的来说，我们的数据提供了证据，表明蜗牛转录因子可能通过多种途径促进 LNCaP 细胞的肿瘤侵袭性；EMT 的诱导可能需要促进迁移，而 NED 可能通过旁分泌机制促进肿瘤增殖。因此，Snail 的治疗靶向不仅可能有利于 EMT 的消除，而且可能有利于 NED 的消除。
