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High throughput quantitative whole transcriptome analysis of individual macrophages 7 days post-pneumonectomy in a B6 CSF1R-GFP mouse

ABSTRACT: Using fluorescence activated cell sorting, we isolated CD45+, CSF1R-GFP+, F4/80+, Ly6G- mouse lung monocytes and macrophages at 7 days after pneumonectomy procedure. We then used microfluidic single cell RNA-sequencing to transcriptional profile unique myeloid subsets. Using the pneumonectomy dataset, we identified 6 cell groups and 4 gene groups that marked several regenerative macrophage subsets including CCR2+, Ly6C+ monocytes and CD206+, Chil3+ M2-like macrophages.

ORGANISM(S): Mus musculus

PROVIDER: GSE96104 | GEO | 2017/06/19

SECONDARY ACCESSION(S): PRJNA378847

REPOSITORIES: GEO

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Project description:The adult human lung has a very limited capacity to regenerate functional alveoli. In contrast, adult mice have a remarkable capacity for neoalveolarization following either lung resection or injury. The molecular basis for this unique capability to regenerate lung tissue in mice is largely unknown. We examined the transcriptomic responses to single lung pneumonectomy in adult mice in order to elucidate prospective molecular signaling used in this species during lung regeneration. Unilateral left pneumonectomy or sham thoracotomy was performed under general anesthesia (n = 8 mice per group for each of the four time points). Total RNA was isolated from the remaining lung tissue at four time points post-surgery (6 hours, 1 day, 3 days, 7 days) and analyzed using microarray technology. The observed transcriptomic patterns revealed mesenchymal cell signaling, including up-regulation of genes previously associated with activated fibroblasts (Tnfrsf12a, Tnc, Eln, Col3A1), as well as modulation of Igf1-mediated signaling. The data set also revealed early down-regulation of pro-inflammatory cytokine transcripts, up-regulation of genes involved in T cell development and function, but few similarities to transcriptomic patterns observed during embryonic or post-natal lung development. Immunohistochemical analysis suggests that early fibroblast but not myofibroblast proliferation is important during lung regeneration and may explain the preponderance of mesenchymal-associated genes that are over-expressed in this model. This appears to differ from embryonic alveologenesis. These data suggest that modulation of mesenchymal cell signaling and proliferation may act in concert with immunomodulation to control inflammation during post-pneumonectomy lung regeneration in adult mice. Experiment Overall Design: For each of the four time points (6 hr, 1 day, 3 day, 7 day), the mice were divided into two groups: (1) pneumonectomy (PNY) and (2) sham operated (SHAM - thoracotomy without lung resection), with eight animals in each group. One microarray was performed on pooled lung tissue from these 8 animals for each time point. Analysis of expression in PNY vs SHAM animals was performed in two ways: using all four microarrays as replicates in a time-independent analysis, as well as analysis of each time point separately (time-dependent) without replicates.

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High throughput quantitative whole transcriptome analysis of individual macrophages 7 days post-pneumonectomy in a B6 CSF1R-GFP mouse

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