The Cell Phenotyping and Mouse Core

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Project Summary The Cell Phenotyping and Mouse Core, designated as Core C, is a centralized facility that will provide investigators with a well-established and reproducible model of influenza A–induced pneumonia. Core C will generate stocks of influenza (A/WSN/33) virus, titrate and maintain viral stocks, and perform infection of the experimental animals as outlined in Projects 1, 2, 3 and 4. Quantitative tools such as Flexivent measurements of lung mechanics, quantitative histological assessment, flow cytometric phenotyping and sorting of the immune cellular populations, and multiplex measures of pro-inflammatory cytokines will be conducted by Core C according to the research plan outlined in Projects 1, 2, 3 and 4. Core C will use advanced flow cytometry capabilities to quantify, phenotype, and sort specific inflammatory and parenchymal cell populations (such as tissue-resident and recruited alveolar macrophages, perivascular and peribronchial interstitial macrophages, monocyte subsets, dendritic cell subsets, T, B and NK cell subsets, alveolar epithelial type II cells, fibroblast subsets, endothelial cells) from the murine lung and lymphoid organs. The sorting and isolation capabilities of the Core will allow cell-type-specific assessment of transcriptomic response via RNA-sequencing, including single-cell RNA-sequencing. To this matter, Core will provide automated DNA/RNA isolation from sorted cells, followed by quality assessment and RNA-sequencing library construction and sequencing. Core C will breed mice to generate the cell-type- or tissue-specific Cre recombinase lines to induce tissue-specific knockout mice. Core C will perform the genotyping of all the murine strains proposed by Project Leaders. Core C will maintain a uniform environment in which wild-type and genetically engineered mice will be maintained, subjected to lung injury and allowed to recover. Collectively, these tools provide a unique resource for Project Investigators, which would be difficult to reproduce without the support of this PPG.