Department of Biology, York University, Toronto, ON, Canada.
Centre for Research on Biomolecular Interactions, York University, Toronto, ON, Canada.
Front Endocrinol (Lausanne). 2022 Mar 1;13:842587. doi: 10.3389/fendo.2022.842587. eCollection 2022.
The acquisition of an endovascular trophoblast (enEVT) phenotype is essential for normal placental development and healthy pregnancy. MicroRNAs (miRNAs) are small noncoding RNAs that play critical roles in regulating gene expression. We have recently reported that miR-218-5p promotes enEVT differentiation and spiral artery remodeling in part by targeting transforming growth factor β2 (TGFβ2). We also identified , which encodes interleukin 1β (IL1β), as one of the most highly upregulated genes by miR-218-5p. In this study, we investigated how miR-218-5p regulates expression and IL1β secretion and the potential role of IL1β in enEVT differentiation. Using two cell lines derived from extravillous trophoblasts (EVTs), HTR-8/SVneo and Swan 71, we found that stable overexpression of miR-218-5p precursor, mir-218-1, or transient transfection of miR-218-5p mimic, significantly increased mRNA and IL1β protein levels in cells and conditioned media. We also showed that miR-218-5p directly interacted with SMAD2 3'UTR and reduced SMAD2 at mRNA and protein levels. Knockdown of SMAD2 induced expression and attenuated the inhibitory effect of TGFβ2 on expression. On the other hand, overexpression of SMAD2 reduced IL1β levels and blocked the stimulatory effects of miR-218-5p on expression, trophoblast migration and endothelial-like network formation. In addition, treatment of trophoblasts with IL1β induced the formation of endothelial-like networks and the expression of enEVT markers in a dose-dependent manner. These results suggest that miR-218-5p inhibits the TGFβ/SMAD2 pathway to induce IL1β and enEVT differentiation. Finally, low doses of IL1β also inhibited the expression of miR-218-5p, suggesting the existence of a negative feedback regulatory loop. Taken together, our findings suggest a novel interactive miR-218-5p/TGFβ/SMAD2/IL1β signaling nexus that regulates enEVT differentiation.
滋养层细胞向血管内皮细胞的转化（enEVT）表型的获得对于正常胎盘发育和妊娠健康至关重要。微小 RNA（miRNA）是一种小的非编码 RNA，在调节基因表达中发挥关键作用。我们最近报道，miR-218-5p 通过靶向转化生长因子β2（TGFβ2）促进 enEVT 分化和螺旋动脉重塑。我们还鉴定出编码白细胞介素 1β（IL1β）的 是 miR-218-5p 上调最显著的基因之一。在这项研究中，我们研究了 miR-218-5p 如何调节 的表达和 IL1β 的分泌，以及 IL1β 在 enEVT 分化中的潜在作用。使用源自绒毛外滋养层细胞（EVTs）的两个细胞系 HTR-8/SVneo 和 Swan 71，我们发现稳定过表达 miR-218-5p 前体 mir-218-1 或瞬时转染 miR-218-5p 模拟物显著增加了细胞和条件培养基中的 mRNA 和 IL1β 蛋白水平。我们还表明，miR-218-5p 直接与 SMAD2 3'UTR 相互作用，降低了 SMAD2 的 mRNA 和蛋白水平。SMAD2 的敲低诱导了 的表达，并减弱了 TGFβ2 对 的表达的抑制作用。另一方面，SMAD2 的过表达降低了 IL1β 水平并阻断了 miR-218-5p 对 的表达、滋养层迁移和内皮样网络形成的刺激作用。此外，IL1β 处理滋养层细胞以剂量依赖性方式诱导内皮样网络的形成和 enEVT 标志物的表达。这些结果表明，miR-218-5p 通过抑制 TGFβ/SMAD2 途径诱导 IL1β 和 enEVT 分化。最后，低剂量的 IL1β 也抑制了 miR-218-5p 的表达，表明存在负反馈调节环。总之，我们的研究结果表明，miR-218-5p/TGFβ/SMAD2/IL1β 信号轴在调节 enEVT 分化中存在一种新的相互作用。
