Neutrophil elastase-cleaved corticosteroid-binding globulin is absent in human plasma. Issue 1 (January 2019)

• Record Type: Journal Article
• Authors: Hill, Lesley A; Vassiliadi, Dimitra A; Dimopoulou, Ioanna; Anderson, Anna J; Boyle, Luke D; Kilgour, Alixe H M; Stimson, Roland H; Machado, Yoan; Overall, Christopher M; Walker, Brian R; Lewis, John G; Hammond, Geoffrey L
• Is Part Of: Journal of endocrinology. Volume 240:Issue 1(2019)
• Page Start: 27
• Page End: 39
• Publication Date: 2019-01
• Subjects: cortisol -- monoclonal antibody -- ELISA -- steroid binding; Endocrinology -- Periodicals; 616.4005
• DOI: 10.1530/JOE-18-0479
• Languages: English
• ISSNs: 0022-0795

Abstract

Abstract : Corticosteroid-binding globulin (CBG) transports glucocorticoids in blood and is a serine protease inhibitor family member. Human CBG has a reactive center loop (RCL) which, when cleaved by neutrophil elastase (NE), disrupts its steroid-binding activity. Measurements of CBG levels are typically based on steroid-binding capacity or immunoassays. Discrepancies in ELISAs using monoclonal antibodies that discriminate between intact vs RCL-cleaved CBG have been interpreted as evidence that CBG with a cleaved RCL and low affinity for cortisol exists in the circulation. We examined the biochemical properties of plasma CBG in samples with discordant ELISA measurements and sought to identify RCL-cleaved CBG in human blood samples. Plasma CBG-binding capacity and ELISA values were consistent in arterial and venous blood draining skeletal muscle, liver and brain, as well as from a tissue (adipose) expected to contain activated neutrophils in obese individuals. Moreover, RCL-cleaved CBG was undetectable in plasma from critically ill patients, irrespective of whether their ELISA measurements were concordant or discordant. We found no evidence of RCL-cleaved CBG in plasma using a heat-dependent polymerization assay, and CBG that resists immunoprecipitation with a monoclonal antibody designed to specifically recognize an intact RCL, bound steroids with a high affinity. In addition, mass spectrometry confirmed the absence of NE-cleaved CBG in plasma in which ELISA values were highly discordant. Human CBG with a NE-cleaved RCL and low affinity for steroids is absent in blood samples, and CBG ELISA discrepancies likely reflect structural differences that alter epitopes recognized by specific monoclonal antibodies.