1. Enzyme-Linked Immunoabsorbent Assay Detection of a Soluble Form of Urokinase Plasminogen Activator Receptor In Vivo
- Author
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Mizukami, Ikuko F., Faulkner, Neil E., Gyetko, Margaret R., Sitrin, Robert G., and III, Robert F.Todd
- Abstract
The receptor for urokinase plasminogen activator (uPA-R, CD87) is a glycosylphosphatidylinositol (GPD-anchored 50 to 65 kD glycoprotein that, by regulating membrane-associated plasmin activity, may facilitate the invasion of inflammatory and malignant cells. Certain other GPI-anchored glycoproteins are shed from the cell membrane and exist as soluble products in vitro and in vivo. To determine if uPA-R undergoes a similar phenomenon, we have developed a sensitive enzyme-linked immunoabsorbent assay (ELISA) (using a rabbit antiserum as both capture and detection reagents) to measure the quantity of soluble uPA-R (suPA-R) in tissue culture supernatants and biologic fluids. Using this ELISA, we have detected suPA-R in the culture supernatants of U-937 cells and human monocytes stimulated in vitro by certain soluble inflammatory mediators (Sitrin et al. Blood84:1268, 1994; Mizukami et al., Clin Res42:115A, 1994). To determine if suPA-R exists in vivo, we have screened the plasma of 20 normal volunteers (mean ± SD, 3 ± 3 ng/mL; median, 2 ng/mL; range, 1 to 11 ng/mL [serum values slightly higher]); the plasma of 13 ICU patients with clinical sepsis syndrome (mean ± SD, 30 ± 11 ng/mL; median, 11 ng/mL; range, 4 to 221 ng/mL); and the extravascular fluids (pleural, pericardial, and peritoneal) of 84 individuals with presumed inflammatory or malignant conditions (mean ± SD, 21 ± 39 ng/mL; median, 10 ng/mL; range, 2 to 253 ng/ mL). Among the latter specimens, most were inflammatory exudates (only six were malignant by positive cytology) with the highest quantities of suPA-R associated with neutrophilic exudates. The solubility of suPA-R contained within these fluids was confirmed by reanalysis after ultracentrifu-gation to remove particulate material. When tested in a uPA ligand capture ELISA, representative specimens of extravascular body fluids and sepsis plasma contained suPA-R capable of binding uPA ligand (generally representing a small fraction of the immunoreactive material). We conclude from these data that suPA-R is immunologically detectable in vitro and in vivo with high concentrations of receptor found under conditions of inflammatory stimulation. The possibility of suPA-R's biologic activity is suggested by its partial retention of ligand binding capacity.
- Published
- 1995
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