Variability of PECAM-1 Expression on Blood Cells.

PECAM-1 is a 130 kDa member of the Immunoglobulin gene superfamily that is expressed on circulating platelets, neutrophils, monocytes, and certain lymphocyte subpopulations. Deficiency of PECAM-1 in mice results in hyper-reactive platelets, B cells, and mast cells, compromised vascular integrity, and endothelial cells that are more susceptible to radiation-induced apoptosis in vivo. Whether or not PECAM-1 expression levels vary in the human population, and whether variable PECAM-1 expression renders one more or less susceptible to cardiovascular disease, autoimmune disorders, or sepsis has not been investigated to date. As a first step in examining these questions, we have developed a quantitative flow cytometry-based assay to determine the number of PECAM-1 molecules expressed on the surface of blood and vascular cells. Whole blood samples obtained from anonymous normal volunteer blood donors were stained with saturating amounts of fluorochrome-conjugated anti-PECAM-1 antibody, and relative fluorescence intensities were determined for platelets, neutrophils, monocytes, and lymphocytes by flow cytometry. Polystyrene microbeads coated with calibrated numbers of antibody binding sites were saturated with the same fluorochrome-conjugated anti-PECAM-1 antibody and used to create a standard curve that allowed us to relate relative fluorescence intensities to known numbers of antibody binding sites. By plotting the fluorescence intensity of anti-PECAM-1 antibody binding to each cell population against this standard curve, we were able to quantify the number of PECAM-1 molecules on the surface of each cell population. We found that PECAM-1 expression levels most often ranged between 5,000–12,000 molecules/platelet, 6,000–10,000/lymphocyte, 30,000–40,000/neutrophil, and 50,000–80,000/monocyte. Some individuals, however, expressed 2–5 times more PECAM-1 on a given blood cell subtype. Interestingly, individuals who expressed high levels of platelet PECAM-1 did not necessarily also express high levels of leukocyte PECAM-1, and vice versa. Since a number of naturally-occurring alleles of the PECAM-1 gene have recently been associated with the progression of atherosclerosis, myocardial infarction, and/or coronary artery disease, correlation of genetic variation within the PECAM-1 locus with its variable expression on the surface of blood and vascular cells may permit addition of PECAM-1 to the growing list of cell adhesion and signaling receptors whose expression is linked to bleeding and clotting disorders in humans.