Transcriptional landscape of pulmonary lymphatic endothelial cells during fetal gestation

• Published in: PloS one 2019-05, Vol.14 (5), p.e0216795-e0216795
• Main Authors: Norman, Jr, Timothy A, Gower, Adam C, Chen, Felicia, Fine, Alan
• Format: Article
• Language: English
• Subjects: Angiogenesis; Animals; Annotations; Antigens; Binding sites; Biochemistry; Biological activity; Biological response modifiers; Biology and Life Sciences; Biosynthesis; Cell adhesion & migration; Cell surface; Cellular signal transduction; Chemokines; Clusters; Coagulation; Criminal investigation; Cytological research; DNA microarrays; EDTA; Endothelial cells; Endothelial Cells - cytology; Endothelial Cells - metabolism; Endothelium; Extracellular matrix; Fetal Development - physiology; Fetus; Fetus - cytology; Fetus - embryology; Fetuses; Gene expression; Gene Expression Profiling; Gene Expression Regulation, Developmental - physiology; Gene set enrichment analysis; Genes; Genetic aspects; Genetic programs; Genomes; Genomics; Gestation; Health aspects; Hybridization; Immune response; Immune system; Innate immunity; Interferon; Lipid metabolism; Lipids; Lungs; Lymphatic system; Maturation; Medicine; Medicine and Health Sciences; Metabolism; Methods; Mice; NF-κB protein; Novels; Pathology; Physiological aspects; Pregnancy; Pregnant women; Research and Analysis Methods; Signal transduction; Signal Transduction - physiology; Signaling; Surfactants; Transcription; Transcription (Genetics); Transcriptome - physiology; Vascular endothelial growth factor receptors; α-Interferon
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0216795

The genetic programs responsible for pulmonary lymphatic maturation prior to birth are not known. To address this gap in knowledge, we developed a novel cell sorting strategy to collect fetal pulmonary lymphatic endothelial cells (PLECs) for global transcriptional profiling. We identified PLECs based on their unique cell surface immunophenotype (CD31+/Vegfr3+/Lyve1+/Pdpn+) and isolated them from murine lungs during late gestation (E16.5, E17.5, E18.5). Gene expression profiling was performed using whole-genome microarrays, and 1,281 genes were significantly differentially expressed with respect to time (FDR q < 0.05) and grouped into six clusters. Two clusters containing a total of 493 genes strongly upregulated at E18.5 were significantly enriched in genes with functional annotations corresponding to innate immune response, positive regulation of angiogenesis, complement & coagulation cascade, ECM/cell-adhesion, and lipid metabolism. Gene Set Enrichment Analysis identified several pathways coordinately upregulated during late gestation, the strongest of which was the type-I IFN-α/β signaling pathway. Upregulation of canonical interferon target genes was confirmed by qRT-PCR and in situ hybridization in E18.5 PLECs. We also identified transcriptional events consistent with a prenatal PLEC maturation program. This PLEC-specific program included individual genes (Ch25h, Itpkc, Pcdhac2 and S1pr3) as well as a set of chemokines and genes containing an NF-κB binding site in their promoter. Overall, this work reveals transcriptional insights into the genes, signaling pathways and biological processes associated with pulmonary lymphatic maturation in the fetal lung.