Abstract 1054: Widely Used Endothelial Progenitor Cell Assays Define Different Classes of Circulating Progenitor Cells
Introduction: Interest in the assessment of circulating endogenous progenitor cells (EPCs) is growing as evidence of their role in vascular repair mounts. Multiple measures of EPCs have been described, but there has been limited study of the comparability of these assays.
Methods: We sought to determine the correlation between and reproducibility of alternative EPC assay methodologies. The precision of each EPC assay was determined by comparing EPC numbers in duplicate samples. Temporal stability of EPCs was assessed in samples obtained 24 hours apart from healthy volunteers. We then assessed the correlation of EPCs identified using the two most commonly used culture techniques: endothelial cell outgrowth and colony forming unit outgrowth (CFUs) (n=110), as well as EPCs based on cell surface marker expression (CD133, CD34, and VEGFR-2) and aldehyde dehydrogenase (ALDH) activity (n=77) in patients with suspected coronary disease.
Results: EPC enumeration based on flow cytometry (FACS) based techniques was more precise (r2 = 0.84 vs. 0.70) and temporally stable (r2 = 0.82 vs. 0.21) than culture assays. There was limited correlation in EPC numbers determined using the two common culture based assays, either with each other (p=NS) or with EPCs enumerated based on CD34/CD133 expression (p=NS); however, endothelial CFUs correlated with VEGFR-2 (p<0.0001, Spearman r = 0.60) and CD34/VEGFR-2 (p<0.05, Spearman r = 0.31) expressing cells. EPCs defined by expression of CD133, CD34 and CD133/CD34 correlated with each other (CD133+ vs. CD34+ cells, p<0.05, r = 0.27; CD34+ vs. CD133+CD34+ cells, p<0.0001, r2 = 0.52) and with ALDHbr cells (ALDHbr vs. CD133+CD34+ cells, p<0.05, r2 = 0.27), but not with VEGFR-2+ cells (p=NS).
Conclusion: Identification of EPCs using flow cytometry can be performed more precisely and reliably than using culture techniques. EPCs can be broadly grouped into two classes:
VEGFR-2 expressing cells which are correlated with the numbers of endothelial CFUs, and
cells identified based on expression of CD133/CD34 or ALDH activity.
These observations underscore the need for better assay standardization and a more precise definition of EPCs in cellular therapy research.