早期人类胎盘生长和分化的失败与严重的妊娠疾病有关，例如先兆子痫和胎儿生长受限。然而，控制其上皮细胞（滋养层）发育的调节机制仍不清楚。利用妊娠早期的滋养层干细胞（TSC）、滋养层类器官（TB-ORG）和原代细胞滋养层（CTB），我们发现自分泌NOTCH3信号传导控制人胎盘扩张和分化。 NOTCH3 受体在增殖性 CTB 祖细胞中特异性表达，其活性形式，即核 NOTCH3 胞内结构域 (NOTCH3-ICD)，与转录共激活因子 Mastermind-like 1 (MAML1) 相互作用。 TSC 系中多西环素诱导的显性失活 MAML1 表达引发了细胞融合，并上调了多核合体滋养层（胎盘中分化的激素产生细胞类型）特异的基因。然而，祖细胞扩增以及滋养层干性和增殖的标志物受到抑制。因此，抑制NOTCH3信号传导会减少TB-ORG的生长，而原代CTB和TSC中NOTCH3-ICD的过度表达则表现出相反的效果。总之，数据表明规范的 NOTCH3 信号在人类胎盘发育中发挥关键作用，促进 CTB 祖细胞的自我更新。© 2023。由 The Company of Biologies Ltd 出版。

Failures in growth and differentiation of the early human placenta are associated with severe pregnancy disorders such as preeclampsia and fetal growth restriction. However, regulatory mechanisms controlling development of its epithelial cells, the trophoblasts, remain poorly elucidated. Using trophoblast stem cells (TSCs), trophoblast organoids (TB-ORGs) and primary cytotrophoblasts (CTBs) of early pregnancy, we herein show that autocrine NOTCH3 signalling controls human placental expansion and differentiation. NOTCH3 receptor was specifically expressed in proliferative CTB progenitors and its active form, the nuclear NOTCH3 intracellular domain (NOTCH3-ICD), interacted with the transcriptional co-activator Mastermind-like 1 (MAML1). Doxycyclin-inducible expression of dominant-negative MAML1 in TSC lines provoked cell fusion and upregulation of genes specific for multinucleated syncytiotrophoblasts, the differentiated hormone-producing cell type of the placenta. However, progenitor expansion and markers of trophoblast stemness and proliferation were suppressed. Accordingly, inhibition of NOTCH3 signalling diminished growth of TB-ORGs whereas overexpression of NOTCH3-ICD in primary CTBs and TSCs showed opposite effects. In conclusion, the data suggest that canonical NOTCH3 signalling plays a key role in human placental development promoting self-renewal of CTB progenitors.© 2023. Published by The Company of Biologists Ltd.