Diversity in mesenchymal lineages during early human lung development

Abstract

The mesenchyme gives rise to multiple distinct cell lineages in the mature respiratory system, including smooth muscle cells of the airway and vasculature, vascular endothelial cells, and parenchymal fibroblasts. However, a thorough understanding of the specification and the inter-relationships among the diverse mesenchymally derived cells in the human lung is lacking. We used single cell RNAseq of human fetal lung cells, to characterize cellular phenotypes in the pseudoglandular (11.5 weeks) and early canalicular (18.5 weeks) stage human lung. Confirmed live cells obtained from protease-dispersed tissue were captured and sequenced using the Chromium 10X system. Expression analysis was performed using Seurat 2.0 and functional analysis was performed using ToppFun. At these early stages, we were able to identify molecularly distinct cell populations representing fetal human lung endothelial cells, pericytes and smooth muscle cells. Early endothelial lineages expressed “classic” endothelial cell markers (PECAM, CDH5, VWF) as well as CD34, KIT and CLDN5. Cells with pericyte characteristics were evident early in development, and defined by expression of PDGFRB, THY1, notch signaling, and broad expression of basement membrane molecules (COL4, laminin, proteoglycans). Smooth muscle cells, defined by expression of the canonical lineage marker ACTA2, demonstrated evidence for Hedgehog signaling, with high expression of HHIP and PTCH1. At both 11 and 18 weeks, we identified a large population of undefined mesenchymal cells characterized by expression of COL1, PDGFRA and elastin fiber genes (ELN, MFAP4, FBLN1, LOXL1). Our data suggest that early specification of distinct mesenchymal lineages occurs in the human lung.