Single-cell genomics of post-pneumonectomy peripheral lung regeneration
Bennett, R., Ysasi, A., Belle, J. et al.
In most mammals, including humans, unilateral pneumonectomy (PNX) results in non-uniform growth of the remaining lung. To identify regions of active regeneration, we used laser microdissection, microfluidics isolation and single-cell microgenomic PCR.
C57BL/6 mice, 8–11 weeks, underwent left PNX. Lungs were harvested on days 1, 3 & 7 after PNX. Lung sections (300 microns) were laser microdissected & enzymatically dissociated. Cells were isolated on an integrated microfluidics chip (Fluidigm). Custom PCR arrays were used to investigate expression of 96 genes implicated in lung regeneration.
Laser microdissection yielded 1–3 x 103 cells/alveolar duct. Single-cell analysis revealed gene clusters corresponding to previously identified cell types (e.g., endothelial, myofibroblasts). Comparison of gene expression between cells isolated on the C1™ chip vs by flow cytometry showed variable expression in some genes post-PNX that was not evident in population-based flow cytometry analysis. Cell-type specific population dynamics were also notable over the first 7 days post-PNX.
Laser microdissection facilitates harvesting of regenerating alveolar ducts. Single-cell transcriptional analysis identified individual cell types involved in lung regeneration & highlighted specific cell-type population dynamics within discreet regenerating areas.
Bennett, R., Ysasi, A., Belle, J. et al. "Single-cell genomics of post-pneumonectomy peripheral lung regeneration" The FASEB Journal (2015): 876.12