Preparing for the first breath at single cell level [single cell Fluidigm C1]

The respiratory system undergoes remarkable structural, biochemical, and functional changes necessary for adaptation to air breathing at birth. To identify dynamic changes in gene expression in the diverse pulmonary cells at birth, we performed Drop-seq based massive parallel single-cell RNA sequencing. An iterative cell type identification strategy was used to unbiasedly identify the heterogeneity of murine pulmonary cell types on postnatal day 1. Distinct populations of epithelial, endothelial, mesenchymal, and immune cells were identified, each containing distinct subpopulations. Cell type predictions and signature genes identified using Drop-seq were cross-validated using an independent single cell isolation platform. Temporal changes in RNA expression patterns were compared before and after birth to identify signaling pathways selectively activated in specific pulmonary cell types, demonstrating activation of UPR signaling during perinatal adaptation of the lung. Present data provide the first single cell view of the adaptation to air breathing after birth. All data from the present study are freely accessed at https://research.cchmc.org/pbge/lunggens/SCLAB.html. Overall design: Left and right lobes of PND1 mouse lungs were rapidly dissected in ice-cold PBS. Cell concentration was examined with a hemocytometer and adjusted to around 300 cells per microliter for Fluidigm C1.

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Guo M, Du Y, Gokey JJ, Ray S, Bell SM, Adam M, Sudha P, Perl AK, Deshmukh H, Potter SS, Whitsett JA, Xu Y