Plasma free cortisol fraction reflects levels of functioning corticosteroid-binding globulin
Introduction
Human plasma corticosteroid-binding globulin (CBG) is a 52 kDa glycoprotein, which is reported, based on theoretical considerations of albumin and CBG binding affinities, to bind 80–90% of circulating cortisol with high affinity [1], [2], [3]. Albumin bound cortisol is reported to represent 14% and free cortisol 6% of total plasma cortisol under basal conditions [4]. We recently found a patient with a genetically determined complete deficiency of circulating CBG, providing a unique opportunity to examine the distribution of cortisol in plasma [5]. Our aim was to determine the true quantitative role of CBG in binding circulating cortisol and to this end we studied healthy subjects' plasma with ligand competition with and without heat treatment to inactivate CBG. We also measured circadian total and free cortisol levels in a CBG-deficient subject and a control individual. We found that CBG binds more than 80% of circulating cortisol in normal intact plasma. However when CBG is inactivated by heat or absent then the majority of cortisol becomes associated with albumin (75%) with the percentage free cortisol rising markedly to 25% or more.
Section snippets
Measurements of free and total cortisol
Plasma was collected from normal subjects in EDTA-containing tubes between 0800 and 1000 h, unless otherwise stated, and from subjects undergoing an ACTH 1–24 (cosyntropin 250 μg i.m.) with cortisol levels at 0 and 30 min post-dose. Plasma was stored frozen until required. Free cortisol was measured by an ultrafiltration/ligand binding method [6] validated in our laboratory. Briefly, plasma (0.5 mL) was equilibrated with [3H]-cortisol (0.1 μCi, Amersham TRK 407) for 30 min at 37 °C followed by
Results
Free cortisol fractions in plasma and albumin, expressed as a percentage (free cortisol/total cortisol × 100) are shown in Table 1. Untreated normal plasma showed expected free cortisol fractions of less than 6% which is similar to normal subjects (Table 2). Heat treatment of plasma led to undetectable CBG levels by ELISA and unchanged total cortisol levels. Either heat-treatment of plasma or adding an excess of unlabelled cortisol or both led to free cortisol fractions of 20–26%. Human albumin
Discussion
Our data indicate that while most cortisol is bound to CBG under normal conditions and less than 6% is free this increases markedly to between 25% and 35% in CBG-deficient individuals and when plasma CBG is heat inactivated. This is also in contrast to patients with abnormally high total basal cortisol levels (1500–2000 nmol/L, data not shown) or following cosyntropin where the free cortisol normally rises but only to a maximum of 14% (Table 3) when CBG would be expected to be saturated. The
References (12)
- et al.
The serum transport of steroid hormones
Recent Prog Horm Res
(1982) - et al.
An enzyme-linked immunosorbent assay for corticosteroid-binding globulin using monoclonal and polyclonal antibodies: decline following synthetic ACTH
Clin Chim Acta
(2003) - et al.
Production of a monoclonal antibody to cortisol: application to a direct enzyme-linked immunosorbent assay of plasma
Steroids
(1992) - et al.
Dialyzable free cortisol after stimulation with synacthen
Clin Biochem
(2002) - et al.
Clinical use of unbound plasma cortisol as calculated from total cortisol and corticosteroid-binding globulin
J Steroid Biochem
(1987) Corticosteroid-binding globulin (CBG)
Cited by (162)
Corticosteroid-binding globulin (CBG): spatiotemporal distribution of cortisol in sepsis
2023, Trends in Endocrinology and MetabolismCortisol: Analytical and clinical determinants
2023, Advances in Clinical ChemistryEffects of genetic selection on activity of corticotropic and thyrotropic axes in modern broiler chickens
2022, Domestic Animal EndocrinologyFree and bound cortisol, corticosterone, and metabolic adaptations during the early inflammatory response to an intramammary lipopolysaccharide challenge in dairy cows
2021, Domestic Animal EndocrinologyCitation Excerpt :Only the small remaining fraction of free cortisol (FC) is considered to be biologically active after binding to intracellular receptors [6,7]. More than 80% of corticosteroids are considered to be transported by CBG, whereas 10%–15% is bound by serum albumin, and around 5% remain free [8]. However, the proportion of FC relative to TC is not constant as changes in both corticosteroid binding capacity and affinity alter the amount of circulating FC.