Department of Gynecology and Obstetrics, Shanghai First Maternity and Infant Hospital, TongJi University School of Medicine, Shanghai, 201204, People's Republic of China.
Reproductive Medicine, Qingdao Municipal Hospital, Qingdao, 266071, Shandong, People's Republic of China.
J Mol Med (Berl). 2019 Sep;97(9):1359-1373. doi: 10.1007/s00109-019-01819-4. Epub 2019 Jul 16.
NR4A1 (NUR77) is an orphan nuclear receptor that has been implicated in both cell survival and apoptosis. However, the role of NUR77 in trophoblast function during early placenta development has not been fully elucidated. In this study, we showed that NUR77 expression was significantly lower in the villi of the recurrent miscarriage (RM) group compared to that in the healthy controls (HCs) group. We used immunohistochemistry and found that NUR77 was highly expressed in human placental villi during early pregnancy, especially in syncytiotrophoblast (STB), and was expressed at a much lower level in STB from the RM group than in those from HC group. Western blotting data further confirmed that NUR77 was highly expressed in primary human term placental STB and the FSK-induced BeWo cell line. Moreover, antibody array screening and ELISA revealed that NUR77 promoted significant placental growth factor (PGF) expression during trophoblast fusion. Ectopic overexpression and knockdown experiments demonstrated that PGF was a novel downstream target of NUR77, and serum PGF expression correlated positively with trophoblast NUR77 mRNA levels in HCs and RM patients. Importantly, bioinformatics analysis identified two NUR77 binding sites in the PGF promoter region, and chromatin immunoprecipitation (ChIP) coupled with Western blotting analysis further verified that NUR77 bound directly to the PGF promoter region and promoted PGF expression. Furthermore, in a BeWo/HTR-8 co-culture system, FSK-induced BeWo-secreted PGF promoted HTR-8 cell migration and invasion, and an anti-PGF antibody reversed this effect. Collectively, these results indicated that NUR77 may play a key role in regulating trophoblast invasion at early pregnancy. KEY MESSAGES: NUR77 expression was significantly decreased in the syncytiotrophoblast of the recurrent miscarriage group compared to that in the healthy control group. NUR77 promoted PGF expression during trophoblast fusion. ChIP and western blotting experiments verified that NUR77 bound directly to the PGF promoter region and activated PGF expression in trophoblast. Trophoblast-derived PGF promoted HTR-8 cell migration and invasion in a cell co-culture system.
NR4A1（NUR77）是一种孤儿核受体，它与细胞存活和细胞凋亡均有关。然而，NUR77 在早期胎盘发育中滋养细胞功能中的作用尚未完全阐明。在这项研究中，我们发现复发性流产（RM）组绒毛中的 NUR77 表达明显低于健康对照组（HCs）组。我们使用免疫组织化学方法发现，NUR77 在早孕时人胎盘绒毛中高度表达，特别是在合胞滋养层（STB）中，并且在 RM 组的 STB 中表达水平明显低于 HC 组。Western blotting 数据进一步证实，NUR77 在原代人足月胎盘 STB 和 FSK 诱导的 BeWo 细胞系中高度表达。此外，抗体阵列筛选和 ELISA 显示，NUR77 在滋养细胞融合过程中促进显著的胎盘生长因子（PGF）表达。异位过表达和敲低实验表明，PGF 是 NUR77 的新下游靶标，PGF 表达与 HCs 和 RM 患者的绒毛滋养细胞 NUR77 mRNA 水平呈正相关。重要的是，生物信息学分析鉴定了 PGF 启动子区域中的两个 NUR77 结合位点，染色质免疫沉淀（ChIP）结合 Western blot 分析进一步证实 NUR77 直接结合 PGF 启动子区域并促进 PGF 表达。此外，在 BeWo/HTR-8 共培养系统中，FSK 诱导的 BeWo 分泌的 PGF 促进了 HTR-8 细胞的迁移和侵袭，而抗 PGF 抗体逆转了这种作用。总之，这些结果表明 NUR77 可能在调节早孕滋养细胞侵袭中发挥关键作用。
NUR77 在复发性流产组的合胞滋养层中的表达明显低于健康对照组。
NUR77 在滋养细胞融合过程中促进 PGF 表达。
ChIP 和 Western blot 实验验证了 NUR77 直接结合 PGF 启动子区域并激活滋养细胞中 PGF 的表达。
滋养细胞衍生的 PGF 在细胞共培养系统中促进了 HTR-8 细胞的迁移和侵袭。
