Decidual natural killer cells promote extravillus trophoblast developmental pathways: evidence from trophoblast organoid co-cultures

The placenta an essential extra-embryonic organ that supports the fetus throughout gestation. The interactions between the placenta and the maternal immune system during the first trimester have not been completely characterized, despite their close physical association and hem-allogeneic relationship to each other. The most abundant type of immune cell found in the uterus in the first trimester is the decidual natural killer cell (dNK). Despite their name, dNKs play supportive roles during pregnancy by remodelling uterine spiral arteries. We present evidence suggesting that the matrix metalloproteinases (MMPs) that dNKs secrete to promote this remodelling also drive placental development. This study was performed using a novel co-culture system of dNKs and trophoblast organoids, which are mini-organs that represent two to three different cell types of the human placenta. We found that co-cultures for one week led to significant (p=0.020) increases in organoid area. Through bulk RNA sequencing and immunohistochemical examinations we also observed significant decreases in trophoblast stemness markers, and upregulation of gene sets associated with extravillous trophoblast (EVT) development. These changes were accompanied by significant (p<0.001) increases in collagen subunit gene expression in the organoids, with simultaneous significant decreases (p<0.001) in the proportion of organoid area occupied by collagen as determined through Masson’s Trichrome. Cultures containing dNKs also contained significantly higher levels of MMP1, 3, 9, and 10 in their culture media, each of which can break down collagen. Collectively, these findings demonstrate that dNKs promote changes concordant with trophoblast differentiation towards EVTs and villous branching morphogenesis.