Impact of myeloid cell specific deletion of EGFR on cardiac Cd11b+ non-myocyte gene expression

Epidermal Growth Factor Receptor (EGFR) is a critical factor regulating numerous aspects of cell biology. Though it is widely recognized as an oncogene, EGFR has been implicated in cardiovascular health, and has been shown to have unique cell specific roles in the heart. Macrophages and other myeloid cells play key roles in normal cardiac physiology and post injury pathological outcomes. Myeloid cell EGFR has been implicated in disease, though its role in the normal or failing heart has yet to be established. Here, we crossed EGFR floxed mice with LysM Cre transgenics to specifically delete EGFR in Cd11b+ myeloid cells. We then isolated non-cardiomyocytes and subject them to Cd11b microbeads and subsequent column concentration to obtain cardiac Cd11b+ non-myocyte myeloid cells. We used samples from floxed EGFR controls, and EGFR specific myeloid knockouts. In comparisson to control, myeloid EGFR knockouts had 763 differential transcripts when consdering p adjusted less than 0.05 and log2foldchange greater than absolute value 1.5. To generate 1 biological sample, roughly 500 mg of heart tissue (5-6 hearts) was digested in 12 mls of RPMI media using 450 u/ml of Collagenase II and 60 u/ml of hyaluronidase. Non-myocyte cells were then collected, washed with PBS and subject to Miltenyi "Debris Removal Kit" to separate cells from Debris. Fresh cells were then washed in PBS and subject to Miltenyi "Cd11b Microbeads" and "MiniMacs Separator" to concentrate Cardiac Cd11b+ non-myocyte myeloid cells. In total, three biological replicates were analyzed for each of the 2 genotypes.