Assisted Reproduction Unit, Department of Obstetrics and Gynecology, Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Key Laboratory of Reproductive Dysfunction Management of Zhejiang Province, Hangzhou, China.
Mol Hum Reprod. 2020 May 15;26(5):327-339. doi: 10.1093/molehr/gaaa021.
The study explores the role of neddylation in early trophoblast development and its alteration during the pathogenesis of recurrent spontaneous abortion (RSA). Immunofluorescence and western blot were conducted to evaluate the expression pattern of NEDD8 protein in the first-trimester placentas of healthy control and RSA patients. Neddylated-cullins, especially neddylated-cullin1, were downregulated and their substrate, p21, was accumulated in RSA samples. NEDD8 cytoplasmic recruitment was observed in extravillous trophoblast (EVT) progenitors of RSA placentas. Consistent with the results of clinical samples, neddylation inhibition using MLN4924 in trophoblast cell lines caused obvious p21 accumulation and free NEDD8 cytoplasmic recruitment. Further in vitro study demonstrated neddylation inhibition attenuated proliferation of Jeg-3 cells via p21 accumulation. Moreover, when trophoblast stem (TS) cells derived from first-trimester placentas were cultured for differentiation analyses. MLN4924 impaired the differentiation of TS cells towards EVTs by downregulating HLA-G and GATA3. p21 knockdown could partly rescue MLN4924-suppressed HLA-G and GATA3 expression. In conclusion, cullin1 neddylation-mediated p21 degradation is required for trophoblast proliferation and can affect trophoblast plasticity by affecting HLA-G and GATA3 expression. The results provide insights into the pathological mechanism of RSA and the biological regulation of trophoblast development.
这项研究探讨了 neddylation 在早期滋养层发育中的作用及其在复发性自然流产（RSA）发病机制中的改变。免疫荧光和 Western blot 用于评估健康对照组和 RSA 患者的早期胎盘 NEDD8 蛋白的表达模式。Neddylated-cullins，特别是 neddylated-cullin1，在 RSA 样本中下调，其底物 p21 积累。在 RSA 胎盘的绒毛外滋养层（EVT）祖细胞中观察到 NEDD8 细胞质募集。与临床样本的结果一致，使用 MLN4924 在滋养层细胞系中抑制 neddylation 导致明显的 p21 积累和游离 NEDD8 细胞质募集。进一步的体外研究表明，neddylation 抑制通过 p21 积累抑制 Jeg-3 细胞的增殖。此外，当从早期胎盘培养的滋养层干细胞（TS）细胞进行分化分析时。MLN4924 通过下调 HLA-G 和 GATA3 来损害 TS 细胞向 EVT 的分化。p21 敲低可部分挽救 MLN4924 抑制的 HLA-G 和 GATA3 表达。总之，cullin1 neddylation 介导的 p21 降解对于滋养层增殖是必需的，并且可以通过影响 HLA-G 和 GATA3 表达来影响滋养层的可塑性。这些结果为 RSA 的病理机制和滋养层发育的生物学调节提供了新的见解。
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