Both bone morphogenetic protein 2 (BMP2) and WNT/β-catenin signaling promote human trophoblast cell invasion. BMP2 has been shown to up-regulate bone morphogenetic protein and activin membrane-bound inhibitor (BAMBI) in granulosa cells. Besides, studies indicate BAMBI is a positive regulator for WNT/β-catenin signaling. However, whether BMP2 can increase BAMBI expression to induce WNT/β-catenin signaling for trophoblast cell invasion is still unknown. To study the roles of BAMBI in BMP2-induced activation of WNT/β-catenin signaling and human trophoblast invasion, we used immortalized human extravillous trophoblast (EVT) cell line (HTR8/SVneo) and primary human EVT cells as study models. Messenger RNA and protein levels of target genes were checked with RT-qPCR and Western blot assay respectively. The function of target proteins was studied via small interfering RNA (siRNA)-mediated knockdown. In addition, trophoblast cell invasiveness was examined by matrigel-coated transwell assays. Our results demonstrate that BMP2 treatment increased BAMBI mRNA levels and the activation of WNT/β-catenin signaling including the raised phosphorylation of GSK3β, the up-regulation of active (non-phosphorylated) β-catenin as well as its downstream target molecule cyclin D1, all of which were totally blocked by the activin receptor-like kinases (ALK) 2/3 inhibitor DMH1 or siRNA-mediated knockdown of BAMBI in HTR8/SVneo cells. Consistently, in primary human EVT cells, BMP2 also induced the up-regulation of BAMBI and the activation of WNT/β-catenin signaling indicated by the increased levels of active β-catenin and cyclin D1, which were completely blocked by BAMBI knockdown. In conclusion, BMP2 promotes the activation of canonical WNT/β-catenin signaling and human trophoblast cell invasion by up-regulating BAMBI.

中文翻译：

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骨形态发生蛋白2通过上调BAMBI诱导WNT /β-catenin信号的激活和人类滋养细胞的侵袭。

骨形态发生蛋白2（BMP2）和WNT /β-catenin信号均促进人类滋养细胞的侵袭。已显示BMP2上调颗粒细胞中的骨形态发生蛋白和激活素膜结合抑制剂（BAMBI）。此外，研究表明，BAMBI是WNT /β-catenin信号转导的正调控因子。然而，BMP2是否可以增加BAMBI表达以诱导WNT /β-catenin信号传导滋养细胞入侵。为了研究BAMBI在BMP2诱导的WNT /β-catenin信号激活和人类滋养层细胞入侵中的作用，我们使用永生化的人类绒毛外滋养层（EVT）细胞系（HTR8 / SVneo）和原代人类EVT细胞作为研究模型。分别用RT-qPCR和Western blot法检查靶基因的信使RNA和蛋白质水平。通过小干扰RNA（siRNA）介导的敲低研究了目标蛋白的功能。另外，通过基质胶包被的transwell测定法检查了滋养层细胞的侵袭性。我们的结果表明，BMP2处理可提高BAMBI mRNA水平和WNT /β-catenin信号的激活，包括提高GSK3β的磷酸化，激活（非磷酸化）β-catenin及其下游靶分子细胞周期蛋白D1的上调，所有这些都被激活素受体样激酶（ALK）2/3抑制剂DMH1或siRNA介导的HTR8 / SVneo细胞中BAMBI的敲低完全阻断。一致地，在人类原发性EVT细胞中，BMP2还诱导BAMBI的上调和WNT /β-catenin信号的激活，这由活性β-catenin和cyclin D1的水平升高所表明，完全被BAMBI的封锁所阻止。总之，BMP2通过上调BAMBI促进经典WNT /β-catenin信号的激活和人类滋养细胞的侵袭。