Transcriptional profiling of mouse skin upon antibody-mediated neutrophil depletion

Neutrophils patrol healthy tissues and confer local defense upon perturbations. Recent evidence suggests that they participate in tissue-specific structural programs in addition to their indispensable role in immunity.The goal of this study is to evaluate the effect of neutrophils in healthy mouse skin by comparing the transcriptional landscape of naive skin to neutrophil-depleted skin. Overall design: Neutrophils were depleted with intraperitoneal injection of anti-Ly6G antibody for 2 consecutive days. Back skin was harvested and processed to obtain single-cell suspensions. Samples were barcoded with Oligo-tagged hashtag antibodies (Biolegend) and incubated with an antibody cocktail of Fixable viability dye eFluor780 (eBioscience), CD45-BUV395, CD11b-BV650, Ly6G-PE, EPCAM-APC, mFESK-4-APC, and CD31-APC. Live neutrophils (CD45+CD11b+Ly6G+), other immune cells (CD45+Ly6G-), and structural cells (CD45-Dump+) were isolated by FACS-based sorting.