First trimester trophoblasts forming endothelial-like tubes in vitro emulate a ‘blood vessel development’ gene expression profile

Extravillous cytotrophoblasts isolated from first trimester placenta, and immortalised cell lines derived from them, have the intrinsic ability to form endothelial-like tubes when cultured on Matrigel™ extracellular matrix. This in vitro tube formation may model placental angiogenesis and/or endovascular differentiation by trophoblasts. To interpret the relevance of this phenomenon to placental development, we used a gene expression microarray approach to identify which genes and pathways are associated with the tube-forming phenotype of HTR8/SVneo first trimester trophoblasts (HTR8-M), compared with HTR8/SVneo not forming tubes on plastic culture surface (HTR8-P). Furthermore, we used weighted gene co-expression network analysis (WGCNA) of microarray data to identify modules of co-expressed genes underlying the biological processes. There were 481 genes differentially expressed between HTR8-M and HTR8-P and these were significantly enriched for blood vessel development and related gene ontologies. WGCNA clustered the genes into 9 co-expression modules. One module was significantly associated with HTR8-M (p = 1.15E-05) and contained genes involved in actin cytoskeleton organization, cell migration and blood vessel development, consistent with tube formation on Matrigel. Another module was significantly associated with HTR8-P (p = 1.94E-05) and was enriched for genes involved in mitosis, consistent with proliferation by cells on plastic which do not differentiate. Up-regulation of angiogenesis and vascular development pathways in endovascular trophoblasts in vivo could underpin spiral artery remodelling processes, which are defective in preeclamptic pregnancies.