Common endothelial progenitor cell assays identify discrete endothelial progenitor cell populations
Thomas J Povsic1, Katherine L Zavodni, Enrikas Vainorius
1Duke University Medical Center, Durham, NC 27710, USA. povsi001@mc.duke.edu
Insights
Assessing endothelial progenitor cells (EPCs) revealed distinct cell populations. Standardization of EPC assays is crucial for cell therapy research and reliable results.
Area of Science:
- Cardiovascular Research
- Cell Biology
- Regenerative Medicine
Background:
- Endothelial progenitor cells (EPCs) are crucial for vascular repair.
- Multiple assays exist for EPC quantification, but their comparability is understudied.
- Understanding assay variability is key for reliable research.
Purpose of the Study:
- To evaluate the reproducibility and correlation of different EPC assay methodologies.
- To compare culture-based assays with cell surface marker analysis.
- To identify distinct EPC populations based on functional and phenotypic characteristics.
Main Methods:
- Simultaneous assessment of EPCs using two culture techniques: endothelial cell outgrowth and colony-forming unit (CFU) assays.
- Analysis of EPCs based on cell surface markers (CD133, CD34, VEGFR-2) and aldehyde dehydrogenase (ALDH) activity.
- Comparison of results from fluorescence-activated cell sorting (FACS) and culture assays.
Main Results:
- Fluorescence-activated cell sorting (FACS) provided more precise EPC enumeration than culture assays.
- Limited correlation was observed between the two common culture-based EPC assays.
- Endothelial CFUs correlated with VEGFR-2 and CD34/VEGFR-2 expressing cells, while CD133/CD34 or ALDH(br) cells correlated with each other but not VEGFR-2(+) cells.
Conclusions:
- EPCs can be broadly categorized into VEGFR-2-expressing cells (forming endothelial CFUs) and CD133/CD34 or ALDH(br) cells.
- Significant differences exist between EPC assay methodologies.
- Improved assay standardization and precise EPC definition are essential for cell therapy research.
Background:
Multiple measures of endothelial progenitor cells (EPCs) have been described, but there has been limited study of the comparability of these assays. We sought to determine the reproducibility of and correlation between alternative EPC assay methodologies.
Methods:
We simultaneously assessed EPC numbers in 140 patients undergoing cardiac catheterization using the 2 most commonly used culture techniques: endothelial cell outgrowth and colony-forming unit (CFU). In the final 77 patients, EPCs were also identified on the basis of cell surface marker expression (CD133, CD34, and vascular endothelial growth factor receptor-2 [VEGFR-2]) and aldehyde dehydrogenase (ALDH) activity.
Results:
Endothelial progenitor cell enumeration based on fluorescence activated cell sorting was more precise than culture assays. There was limited correlation between EPC numbers determined using the 2 common culture-based assays; however, endothelial CFUs correlated with VEGFR-2 and CD34/VEGFR-2-expressing cells. Endothelial progenitor cells defined by expression of CD133, CD34, CD133/CD34, and ALDH activity correlated with each other, but not with VEGFR-2(+) cells.
Conclusions:
Endothelial progenitor cells can be broadly classified into 2 classes: VEGFR-2-expressing cells, which give rise to endothelial CFUs, and CD133/CD34 or ALDH(br) cells. These observations underscore the need for better assay standardization and a more precise definition of EPCs in cell therapy research.
