Enzyme-linked immunoabsorbent assay detection of a soluble form of urokinase plasminogen activator receptor in vivo.
The receptor for urokinase plasminogen activator (uPA-R, CD87) is a glycosylphosphatidylinositol (GPI)-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, Blood 84:1268, 1994; Mizukami et al., Clin Res 42:115A, 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 ultracentrifugation 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.
Mizukami
I F
IF
Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, USA.
Faulkner
N E
NE
Gyetko
M R
MR
Sitrin
R G
RG
Todd
R F
RF
D000328:Adult; D000818:Animals; D001202:Ascitic Fluid / Q000737:chemistry* / Q000166:cytology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D007249:Inflammation / Q000378:metabolism; D016403:Lymphoma, Large B-Cell, Diffuse / Q000473:pathology; D008297:Male; D008875:Middle Aged; D009000:Monocytes / Q000378:metabolism; D009369:Neoplasms / Q000378:metabolism; D010996:Pleural Effusion / Q000737:chemistry* / Q000166:cytology; D011817:Rabbits; D011956:Receptors, Cell Surface / Q000032:analysis* / Q000276:immunology; D055293:Receptors, Urokinase Plasminogen Activator; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured
C524898:PLAUR protein, human; D011956:Receptors, Cell Surface; D055293:Receptors, Urokinase Plasminogen Activator