Cell trajectory modeling identifies a primitive trophoblast state defined by BCAM enrichment

Abstract

In early placental development, progenitor cytotrophoblasts (CTB) differentiate along one of two cellular trajectories: the villous or extravillous pathways. CTB committed to the villous pathway fuse with neighboring CTB to form the outer multinucleated syncytiotrophoblast (SCT), whereas CTB committed to the extravillous pathway differentiate into invasive extravillous trophoblasts (EVT). Unfortunately, little is known about the processes controlling human CTB progenitor maintenance and differentiation. To address this, we established a single cell RNA sequencing (scRNA-seq) dataset from first trimester placentas to identify cell states important in trophoblast progenitor establishment, renewal and differentiation. Multiple distinct trophoblast states were identified, representing progenitor CTB, column CTB, SCT precursors and EVT. Lineage trajectory analysis identified a progenitor origin that was reproduced in human trophoblast stem cell organoids. Heightened expression of basal cell adhesion molecule (BCAM) defined this primitive state, where BCAM enrichment or gene silencing resulted in enhanced or diminished organoid growth, respectively. Together, this work describes at high-resolution trophoblast heterogeneity within the first trimester, resolves gene networks within human CTB progenitors and identifies BCAM as a primitive progenitor marker and possible regulator.

Transcriptomic mapping of the metzincin landscape in human trophoblasts

“…To examine the kinetics of CTB-associated protease expression more closely, we applied RNA velocity (scVelo) [35], an algorithm that measures relative abundances of unspliced versus spliced mRNA transcripts to predict cellular state progression. In line with scVelo modeling performed previously on this dataset [26], a predicted origin overlapping with the CTB2 state was identified with multiple paths extending from this origin towards the CTB3, SCTp, and EVT states (Fig. 4A).…”

“…Quality control, cell cycle scoring, scaling, normalization, removal of doublets and integration was done as previously described in Shannon et al . [26].…”

“…Using these published datasets, we set out to comprehensively examine the metzincin protease transcriptomic landscape in human trophoblasts. Implementing our previously described single cell dataset filtering and analysis pipeline [26],…”

“…Recapitulating our previous findings [26], eight cell states were resolved, and by referencing known trophoblast marker genes, four CTB (CTB1-4; TEAD + , EGFR + ), one syncytiotrophoblast precursor (SCTp;…”

“…Advances in single cell transcriptomics allow for unprecedented examination of cell heterogeneity, cellular hierarchy, and gene pathways central to stem cell maintenance and the control of differentiation. In this work, we use published single cell RNA sequencing (scRNA-seq) datasets [25,26] to define the metzincin protease landscape of human trophoblasts in early pregnancy. We show that human trophoblasts express 24 metzincin proteases.…”