Open AccessA Novel Minimally-Invasive Method to Sample Human Endothelial Cells for Molecular Profiling
Author(s) -
Stephen W. Waldo,
Daniel Brenner,
James M. McCabe,
M. D. Soler Cruz,
Brian Long,
Venkata Narla,
Joseph Park,
Ameya Kulkarni,
Elizabeth Sinclair,
Stephen Y. Chan,
Suzaynn F. Schick,
Namita Malik,
Patricia A. Ganz,
Priscilla Y. Hsue
Publication year - 2015
Publication title -
plos one
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.99
H-Index - 332
ISSN - 1932-6203
DOI - 10.1371/journal.pone.0118081
Subject(s) - endothelial stem cell , cell sorting , endothelium , population , cd146 , biology , von willebrand factor , immunology , cell , flow cytometry , cell adhesion molecule , cd34 , endothelial dysfunction , microbiology and biotechnology , medicine , platelet , stem cell , in vitro , biochemistry , endocrinology , environmental health
Objective The endothelium is a key mediator of vascular homeostasis and cardiovascular health. Molecular research on the human endothelium may provide insight into the mechanisms underlying cardiovascular disease. Prior methodology used to isolate human endothelial cells has suffered from poor yields and contamination with other cell types. We thus sought to develop a minimally invasive technique to obtain endothelial cells derived from human subjects with higher yields and purity. Methods Nine healthy volunteers underwent endothelial cell harvesting from antecubital veins using guidewires. Fluorescence-activated cell sorting (FACS) was subsequently used to purify endothelial cells from contaminating cells using endothelial surface markers (CD34 / CD105 / CD146) with the concomitant absence of leukocyte and platelet specific markers (CD11b / CD45). Endothelial lineage in the purified cell population was confirmed by expression of endothelial specific genes and microRNA using quantitative polymerase chain reaction (PCR). Results A median of 4,212 (IQR: 2161 – 6583) endothelial cells were isolated from each subject. Quantitative PCR demonstrated higher expression of von Willebrand Factor (vWF, P<0.001), nitric oxide synthase 3 (NOS3, P<0.001) and vascular cell adhesion molecule 1 (VCAM-1, P<0.003) in the endothelial population compared to similarly isolated leukocytes. Similarly, the level of endothelial specific microRNA-126 was higher in the purified endothelial cells (P<0.001). Conclusion This state-of-the-art technique isolates human endothelial cells for molecular analysis in higher purity and greater numbers than previously possible. This approach will expedite research on the molecular mechanisms of human cardiovascular disease, elucidating its pathophysiology and potential therapeutic targets.