Studies of the release and turnover of a human neutrophil lipocalin

Abstract

A 24-kDa protein was purified from human neutrophil extracts and shown to be the newly discovered neutrophil gelatinase-associated lipocalin (NGAL), based on structural and immunochemical data. A specific enzyme-linked immunosorbent assay (ELISA) was developed for the determination of NGAL in human plasma and tissue fluids. Normal human plasma contains 72 μg 1^‐1 of NGAL (range 40-109 μg 1^‐1) in two main forms, monomer and dimer. ³⁵S-methionine metabolic studies of human neutrophils showed that granulocyte macrophagecolony-stimulating factor (GMCSF) stimulated significant synthesis and secretion of NGAL in a dose- and time-dependent fashion. NGAL was rapidly released as monomer and dimer on incubation of heparinized whole blood with opsonized yeast, reaching a plateau corresponding to about 35% of total cell content after 30 min. Following intravenous injection of ¹²⁵-iodine labelled NGAL there was a more rapid initial clearance of the monomeric than of the dimeric form; t_1/2 10 min vs. 20 min. During the second phase the two forms cleared at similar rates. Severe acute peritonitis was accompanied by a 10-fold increase in NGAL plasma levels and the NGAL level in peritoneal exudates, which reached about 40 mg 1^‐1. There was a good linear correlation between the concentrations of NGAL, leucocyte elastase and NP4 (neutrophil proteinase 4 = P3).

Key Words:

• cytokines
• ELISA
• leucocyte proteinases
• lipocalin
• neutrophil
• peritonitis
• phagocytosis