Unbiased profiling of the origin and function of cardiac interstitial cells

The mammalian heart is an organ that has traded regenerative capacity for complexity, meaning )) that human hearts must replace dead tissue from injuries with fibrotic scar tissue. Fibrosis, thickening of heart tissue caused by a buildup of scar tissue, has a detrimental effect on the mechanical ability of the heart. The interstitial cells found between cardiomyocytes have a major role in scar formation in the heart but very little is understood about the composition of the interstitium. These include immune cells, endothelial cells, smooth muscle cells, pericytes, fibroblasts and various putative stem cell populations. A better understanding of the relation between all these different populations and the dynamic changes occurring after injury can provide important cues on how to modulate cardiac repair. We\u27ll be using a conditional reporter mouse line (WtICre;RosaZsgreerf+) labelling epicardial-derived cells to obtain information about the developmental origin of different cardiac subpopulation. In order to characterize our Wtl xZsGreen mouse model, immunohistochemistry was performed on the heart at homeostasis and at different time points after MI. Additionally, we created an optimal protocol for cell isolation in order to perform single cell RNA sequencing to create an unbiased profile of the cardiac intersitium at baseline and after injury. It was found that there is a significant increase in Wtl positive cells in the ventricles, but not in the atria. Additionally, we empirically determined the best protocol to get the highest yield of single live nucleated cells from a dissociated adult murine heart. This optimization is an essential prelude to the droplet-based single-cell RNA sequencing experiments