Cell-type Specific Surface Markers Effectively Isolate Cardiac Cell Subpopulations

Cardiomyocytes comprise only 20-30% of the cells in the heart. To enable downstream applications requiring live cells (including clinical application), identification of surface markers unique to all cardiac subpopulations is needed. Development of such strategies would create a powerful technique to tease apart the multiple cell populations that make up the heart and their role in complex heart diseases. Our hypothesis was that surface markers could be identified in neonatal mice to distinguish smooth muscle cells (SMC), cardiomyocytes (CM), fibroblasts (FB) and endothelial cells (EC), enabling sorting by flow cytometry and analysis of these subpopulations of heart cells. Hearts were surgically removed from one litter of neonatal (Day 0-2) C57BL/6J mice and three litters of transgenic mice with mCherry α-myosin heavy chain promoter (αMHC-mCherry), then digested with trypsin overnight followed by collagenase/dispase and gentle mechanical dissociation. Surface marker staining for Cadherin-2 (Cdh2) showed congruence with αMHC-mCherry positivity. Cells were sorted using fluorescent activated cell sorting with antibodies against the following surface markers: Cdh2 (CM), CD31 (EC), CD90.2 (FB) and CD49a (SMC). Enrichment of these cell populations was then evaluated by qRT-PCR for known intracellular and extracellular markers specific to each cell: EC (CD31 + vWf), CM (TNNT2 + Myh6), FB (CD90) and SMC (Myh10/11 + Tagln). Our panel of surface markers enabled enrichment of four subpopulations of heart cells which will be verified further in coming experiments. In the future, this strategy will enable sorting of cells from each subpopulation for downstream analysis.