Cardiac progenitor cells are isolated according to the positive expression of the protein marker c-kit from single cell suspensions of enzymatically digested heart atrial biopsy specimens. The cells are spindle-shaped (resembling cardiac fibroblasts) and can differentiate into endothelial smooth muscle or cardiomyocyte lineages.
Cardiac tissues are minced, digested with collagenase type II (0.2%) and filtrated (0.2 um) to form single cell suspensions. The single cells are incubated at 37°C in a humidity chamber for 4-6 days in CPC expansion medium containing Ham’s F12 medium, supplemented with fetal bovine serum (FBS; 10%), penicillin streptomycin (1X), human erythropoietin (2.5 U), glutathione (0.2 mmol/L), and human recombinant basic fibroblast growth factor (bFGF; 5ug). When reaching 60% -70% confluency, the cells are split (1:10) and when the new cultures (P1 cells) reaches 70%–80% confluency, c-kit-positive cells are isolated using FACS.