Graduate Program in Medical Sciences, McMaster University, Hamilton, Ontario, L8N 3Z5, Canada.
Department of Pediatrics, McMaster University, Hamilton, Ontario, L8N 3Z5, Canada.
Sci Rep. 2019 Aug 30;9(1):12607. doi: 10.1038/s41598-019-48816-8.
Placental extravillous trophoblast (EVT) invasion is essential in establishing proper blood supply to the fetus during pregnancy. However, traditional 2D in vitro systems do not model the in vivo invasion process in an anatomically-relevant manner. Our objectives were to develop a 3D spheroid model that would allow better emulation of placental invasion in vitro and to characterize the transcriptomic and functional outcomes. HTR8/SVneo EVT cells were self-assembled into 3D spheroids using ultra-low attachment plates. Transcriptomic profiling followed by gene set enrichment and gene ontology analyses revealed major global transcriptomic differences, with significant up-regulations in EVTs cultured as 3D spheroids in canonical pathways and biological processes such as immune response, angiogenesis, response to stimulus, wound healing, and others. These findings were further validated by RT-qPCR, showing significant up-regulations in genes and/or proteins related to epithelial-mesenchymal transition, cell-cell contact, angiogenesis, and invasion/migration. A high-throughput, spheroid invasion assay was applied to reveal the dynamic invasion of EVTs away from the spheroid core into extracellular matrix. Lastly, lipopolysaccharide, dexamethasone, or Δ-tetrahydrocannabinol exposure was found to impact the invasion of EVT spheroids. Altogether, we present a well-characterized, 3D spheroid model of EVT invasion and demonstrate its potential use in drug and toxin screening during pregnancy.
胎盘绒毛外滋养层（EVT）侵入对于妊娠期间为胎儿建立适当的血液供应至关重要。然而，传统的二维体外系统不能以解剖学相关的方式模拟体内侵入过程。我们的目标是开发一种 3D 球体模型，以更好地模拟体外胎盘侵入，并对其转录组和功能结果进行表征。HTR8/SVneo EVT 细胞使用超低附着板自组装成 3D 球体。转录组谱分析后进行基因集富集和基因本体分析，揭示了主要的全局转录组差异，在 3D 球体培养的 EVT 中，经典途径和生物学过程（如免疫反应、血管生成、对刺激的反应、伤口愈合等）中的基因表达显著上调。这些发现通过 RT-qPCR 进一步验证，显示与上皮-间充质转化、细胞-细胞接触、血管生成和侵袭/迁移相关的基因和/或蛋白显著上调。高通量球体侵袭测定法用于揭示 EVT 从球体核心向细胞外基质的动态侵袭。最后，发现脂多糖、地塞米松或Δ-四氢大麻酚暴露会影响 EVT 球体的侵袭。总之，我们提出了一种经过充分表征的 EVT 侵袭 3D 球体模型，并证明了其在妊娠期间药物和毒素筛选中的潜在用途。
