Cardiac progenitor cells from the left atrial appendage may originate from a resident non-hematopoietic myeloid progenitor population

Abstract

Purpose: We recently identified three distinct cardiac progenitor cell (CPC) populations from the left atrial appendages (LAAs) of adult murine hearts. These populations have distinct phenotypes and characteristics. Type A CPCs are c-kit+/Sca-1+, Type B CPCs are c-kit+/CD45+, and more mature Type C CPCs can be derived from Type B CPCs in vitro. The purpose of this study was to analyze the embryologic origin and the connection between the different CPC populations. Methods: We performed a whole-genome expression array to the cell sorted Type A, Type B and Type C CPC populations. The LAAs were analyzed using immunohistochemistry. Tissue culture derived cells were analyzed using immunostaining and FACS. Neural crest lineage tracing was performed with Pax3-cre eYFP reporter mice. We investigated the spontaneous cardiac differentiation process from Type B to Type C using live microscopy and FACS. Results: Transcriptome comparison demonstrated a significant up-regulation of the neural crest differentiation –pathway genes in Type A and C CPCs. Type B CPCs expressed a gene profile, which matches that of non-hematopoietic, resident macrophage –like cells found in other tissues. Pax3 lineage tracing demonstrated traced cells in the subepicardial zone of the LAA, co-stained with F4/80 macrophage marker and c-kit. Closely attached were epicardially lined F4/80+ cells with no eYFP signal. In tissue culture, eYFP signal was strongest in the Type B CPCs and lower in the Type A CPCs. Fibroblasts in the culture were eYFP negative. The CD45 expression in the Type B CPCs was stable (∼90% of the cells) up to five passages folllowed by a rapid decline of CD45, c-kit and Flk-1 expression. In live microscopy, formation of giant Sca-1+ and Gata-4+ cells by cell fusion was followed by asymmetric divisions creating Type C CPCs, which expressed cardiac myosin heavy chain and atrial natriuretic factor. The transcriptome was significantly skewed toward a cardiomyocyte phenotype with the spontaneous differentiation, although this differentiation was not complete, as seen by electron microscopy. The partially differentiated Type C CPC population could be grown for >60 passages. Conclusion: Our results suggest that rather than having distinct embryonic origins, the different types of CPCs in the LAA originate from a resident, non-hematopoietic myeloid progenitor cell population. The neural crest differentiation pathway can be activated in these cells during adulthood, possibly in cardiac stress situations, enabling differentiation to cardiac lineage with a novel mechanism consisting of cell fusion and asymmetric division.