A novel minimally-invasive method to sample human endothelial cells for molecular profiling.

Autor: Waldo SW; Division of Cardiology, Department of Medicine, Massachusetts General Hospital, Boston, MA, United States of America., Brenner DA; Division of Cardiovascular Medicine, Stanford University Medical Center, Stanford, CA, United States of America., McCabe JM; Currently, Division of Cardiology, Department of Medicine, University of Washington, Seattle, WA, United States of America., Dela Cruz M; Division of Cardiology and the Center of Excellence in Vascular Research, San Francisco General Hospital, University of California, San Francisco, CA, United States of America., Long B; Division of Experimental Medicine, University of California, San Francisco, CA, United States of America., Narla VA; Division of Cardiology and the Center of Excellence in Vascular Research, San Francisco General Hospital, University of California, San Francisco, CA, United States of America., Park J; Division of Cardiovascular Medicine, Brigham and Women’s Hospital, Boston, MA, United States of America., Kulkarni A; Division of Cardiology, Mid-Atlantic Permanente Medical Group., Sinclair E; Division of Experimental Medicine, University of California, San Francisco, CA, United States of America., Chan SY; Division of Cardiovascular Medicine, Brigham and Women’s Hospital, Boston, MA, United States of America., Schick SF; Division of Occupational and Environmental Medicine, University of California, San Francisco, CA, United States of America., Malik N; Division of Occupational and Environmental Medicine, University of California, San Francisco, CA, United States of America., Ganz P; Division of Cardiology and the Center of Excellence in Vascular Research, San Francisco General Hospital, University of California, San Francisco, CA, United States of America., Hsue PY; Division of Cardiology and the Center of Excellence in Vascular Research, San Francisco General Hospital, University of California, San Francisco, CA, United States of America.
Jazyk: angličtina
Zdroj: PloS one [PLoS One] 2015 Feb 13; Vol. 10 (2), pp. e0118081. Date of Electronic Publication: 2015 Feb 13 (Print Publication: 2015).
DOI: 10.1371/journal.pone.0118081
Abstrakt: 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.
Databáze: MEDLINE
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