The Naturally Occurring Glucocorticoids; Cortisol (Hydrocortisone)

THE NATURALLY OCCURRING GLUCOCORTICOIDS; CORTISOL (HYDROCORTISONE)

Pharmacokinetics

Cortisol (also called hydrocortisone, compound F) exerts a wide range of physiologic effects, including regulation of intermediary metabolism, cardiovascular function, growth, and immunity. Its synthesis and secretion are tightly regulated by the central nervous system, which is very sensitive to negative feedback by the circulat-ing cortisol and exogenous (synthetic) glucocorticoids. Cortisol is synthesized from cholesterol (as shown in Figure 39–1).

In the normal adult, in the absence of stress, 10–20 mg of cortisol is secreted daily. The rate of secretion follows a circadian rhythm governed by pulses of ACTH that peak in the early morning hours and after meals, especially after lunch (Figure 39–2). In plasma, cortisol is bound to circulating proteins. Corticosteroid-binding globulin (CBG), an α₂ globulin synthesized by the liver, binds about 90% of the circulating hormone under normal cir-cumstances. The remainder is free (about 5–10%) or loosely bound to albumin (about 5%) and is available to exert its effect on target cells. When plasma cortisol levels exceed 20–30 mcg/dL, CBG is saturated, and the concentration of free cortisol rises rapidly. CBG is increased in pregnancy and with estrogen admin-istration and in hyperthyroidism. It is decreased by hypothyroid-ism, genetic defects in synthesis, and protein deficiency states. Albumin has a large capacity but low affinity for cortisol, and for practical purposes albumin-bound cortisol should be considered free. Synthetic corticosteroids such as dexamethasone are largely bound to albumin rather than CBG.

The half-life of cortisol in the circulation is normally about 60–90 minutes; it may be increased when hydrocortisone (the pharmaceutical preparation of cortisol) is administered in large amounts or when stress, hypothyroidism, or liver disease is present.

Only 1% of cortisol is excreted unchanged in the urine as free cortisol; about 20% of cortisol is converted to cortisone by 11-hydroxysteroid dehydrogenase in the kidney and other tissues with mineralocorticoid receptors  before reaching the liver. Most cortisol is metabolized in the liver. About one third of the cortisol produced daily is excreted in the urine as dihydroxy ketone metabolites and is measured as 17-hydroxysteroids (see Figure 39–3 for carbon numbering). Many cortisol metabolites are conjugated with glucuronic acid or sulfate at the C₃ and C₂₁ hydroxyls, respectively, in the liver; they are then excreted in the urine.

In some species (eg, the rat), corticosterone is the major gluco-corticoid. It is less firmly bound to protein and therefore metabo-lized more rapidly. The pathways of its degradation are similar to those of cortisol.

Pharmacodynamics

A. Mechanism of Action

Most of the known effects of the glucocorticoids are mediated by widely distributed glucocorticoid receptors. These proteins are members of the superfamily of nuclear receptors, which includes steroid, sterol (vitamin D), thyroid, retinoic acid, and many other receptors with unknown or nonexistent ligands (orphan recep-tors). All these receptors interact with the promoters of—and regulate the transcription of—target genes (Figure 39–4). In theabsence of the hormonal ligand, glucocorticoid receptors are pri-marily cytoplasmic, in oligomeric complexes with heat-shock proteins (hsp). The most important of these are two molecules of hsp90, although other proteins are certainly involved. Free hormone from the plasma and interstitial fluid enters the cell and binds to the receptor, inducing conformational changes that allow it to dissociate from the heat shock proteins. The ligand-bound receptor complex then is actively transported into the nucleus, where it interacts with DNA and nuclear proteins. As a homodi-mer, it binds to glucocorticoid receptor elements (GREs) in the promoters of responsive genes. The GRE is composed of two palindromic sequences that bind to the hormone receptor dimer.

In addition to binding to GREs, the ligand-bound receptor also forms complexes with and influences the function of other tran-scription factors, such as AP1 and NF-κB, which act on non-GRE-containing promoters, to contribute to the regulation of transcription of their responsive genes. These transcription factors have broad actions on the regulation of growth factors, proinflammatory cytok-ines, etc, and to a great extent mediate the anti-growth, anti-inflammatory, and immunosuppressive effects of glucocorticoids.

Two genes for the corticoid receptor have been identified: one encoding the classic glucocorticoid receptor (GR) and the other encoding the mineralocorticoid receptor (MR). Alternative splicing of human glucocorticoid receptor pre-mRNA generates two highly homologous isoforms, termed hGR alpha and hGR beta. Human GR alpha is the classic ligand-activated glucocorticoid receptor which, in the hormone-bound state, modulates the expression of glucocorticoid-responsive genes. 

In contrast, hGR beta does not bind glucocorticoids and is transcriptionally inactive. However, hGR beta is able to inhibit the effects of hormone-activated hGR alpha on glucocorticoid-responsive genes, playing the role of a physiologically relevant endogenous inhibitor of glucocorticoid action. It was recently shown that the two hGR alternative tran-scripts have eight distinct translation initiation sites; ie, in a human cell there may be up to 16 GRα and GRβ isoforms, which may form up to 256 homodimers and heterodimers with different tran-scriptional and possibly non-transcriptional activities. This variabil-ity suggests that this important class of steroid receptors has complex stochastic activities.

The prototype glucocorticoid receptor isoform is composed of about 800 amino acids and can be divided into three functional domains (see Figure 2–6). The glucocorticoid-binding domain is located at the carboxyl terminal of the molecule. The DNA-binding domain is located in the middle of the protein and con-tains nine cysteine residues. This region folds into a “two-finger” structure stabilized by zinc ions connected to cysteines to form two tetrahedrons. This part of the molecule binds to the GREs that regulate glucocorticoid action on glucocorticoid-regulated genes. The zinc fingers represent the basic structure by which the DNA-binding domain recognizes specific nucleic acid sequences. The amino-terminal domain is involved in the transactivation activity of the receptor and increases its specificity.

The interaction of glucocorticoid receptors with GREs or other transcription factors is facilitated or inhibited by several families of proteins called steroid receptor coregulators, divided into coactivators and corepressors. The coregulators do this by serving as bridges between the receptors and other nuclear proteins and by expressing enzymatic activities such as histone acetylase or deacetylase, which alter the conformation of nucleosomes and the transcribability of genes.

Between 10% and 20% of expressed genes in a cell are regu-lated by glucocorticoids. The number and affinity of receptors for the hormone, the complement of transcription factors and coregu-lators, and post-transcription events determine the relative speci-ficity of these hormones’ actions in various cells. The effects of glucocorticoids are mainly due to proteins synthesized from mRNA transcribed from their target genes.

Some of the effects of glucocorticoids can be attributed to their binding to mineralocorticoid receptors (MRs). Indeed, MRs bind aldosterone and cortisol with similar affinity. A mineralocorticoid effect of cortisol is avoided in some tissues by expression of 11β-hydroxysteroid dehydrogenase type 2, the enzyme responsible for biotransformation to its 11-keto derivative (cortisone), which has minimal affinity for aldosterone receptors.

Recently, the GR was found to interact with CLOCK/BMAL-1, a transcription factor dimer expressed in all tissues and generating the circadian rhythm of cortisol secretion at the suprachiasmatic nucleus of the hypothalamus. CLOCK is an acetyltransferase that acetylates the hinge region of the GR, neutralizing its transcrip-tional activity and thus rendering target tissues resistant to gluco-corticoids. Interestingly, the glucocorticoid target tissue sensitivity rhythm generated is in reverse phase to that of circulating cortisol concentrations, explaining the increased sensitivity of the organism to evening administration of glucocorticoids.

Prompt effects such as initial feedback suppression of pituitary ACTH occur in minutes and are too rapid to be explained on the basis of gene transcription and protein synthesis. It is not known how these effects are mediated. Among the proposed mechanisms are direct effects on cell membrane receptors for the hormone or nonge-nomic effects of the classic hormone-bound glucocorticoid receptor. The putative membrane receptors might be entirely different from the known intracellular receptors. Recently, all steroid receptors (except the MRs) were shown to have palmitoylation motifs that allow enzymatic addition of palmitate and increased localization of the receptors in the vicinity of plasma membranes. Such receptors are available for direct interactions with and effects on various membrane-associated or cytoplasmic proteins without the need for entry into the nucleus and induction of transcriptional actions.

B. Physiologic Effects

The glucocorticoids have widespread effects because they influ-ence the function of most cells in the body. The major metabolic consequences of glucocorticoid secretion or administration are due to direct actions of these hormones in the cell. However, some important effects are the result of homeostatic responses by insulin and glucagon. Although many of the effects of glucocorticoids are dose-related and become magnified when large amounts are administered for therapeutic purposes, there are also other effects— called permissive effects—without which many normal functions become deficient. For example, the response of vascular and bron-chial smooth muscle to catecholamines is diminished in the absence of cortisol and restored by physiologic amounts of this glucocorticoid. Similarly, the lipolytic responses of fat cells to catecholamines, ACTH, and growth hormone are attenuated in the absence of glucocorticoids.

C. Metabolic Effects

The glucocorticoids have important dose-related effects on carbo-hydrate, protein, and fat metabolism. The same effects are respon-sible for some of the serious adverse effects associated with their use in therapeutic doses. Glucocorticoids stimulate and are required for gluconeogenesis and glycogen synthesis in the fasting state.

They stimulate phosphoenolpyruvate carboxykinase, glucose-6-phosphatase, and glycogen synthase and the release of amino acids in the course of muscle catabolism.

Glucocorticoids increase serum glucose levels and thus stimu-late insulin release and inhibit the uptake of glucose by muscle cells, while they stimulate hormone sensitive lipase and thus lipolysis. The increased insulin secretion stimulates lipogenesis and to a lesser degree inhibits lipolysis, leading to a net increase in fat deposition combined with increased release of fatty acids and glycerol into the circulation.

The net results of these actions are most apparent in the fasting state, when the supply of glucose from gluconeogenesis, the release of amino acids from muscle catabolism, the inhibition of periph-eral glucose uptake, and the stimulation of lipolysis all contribute to maintenance of an adequate glucose supply to the brain.

D. Catabolic and Antianabolic Effects

Although glucocorticoids stimulate RNA and protein synthesis in the liver, they have catabolic and antianabolic effects in lym-phoid and connective tissue, muscle, peripheral fat, and skin. Supraphysiologic amounts of glucocorticoids lead to decreased muscle mass and weakness and thinning of the skin. Catabolic and antianabolic effects on bone are the cause of osteoporosis in Cushing’s syndrome and impose a major limitation in the long-term therapeutic use of glucocorticoids. In children, glucocorti-coids reduce growth. This effect may be partially prevented by administration of growth hormone in high doses, but this use of growth hormone is not recommended.

E. Anti-Inflammatory and Immunosuppressive Effects

Glucocorticoids dramatically reduce the manifestations of inflam-mation. This is due to their profound effects on the concentration, distribution, and function of peripheral leukocytes and to their suppressive effects on the inflammatory cytokines and chemokines and on other mediators of inflammation. Inflammation, regardless of its cause, is characterized by the extravasation and infiltration of leukocytes into the affected tissue. These events are mediated by a complex series of interactions of white cell adhesion molecules with those on endothelial cells and are inhibited by glucocorti-coids. After a single dose of a short-acting glucocorticoid, the concentration of neutrophils in the circulation increases while the lymphocytes (T and B cells), monocytes, eosinophils, and baso-phils decrease. The changes are maximal at 6 hours and are dissi-pated in 24 hours. The increase in neutrophils is due both to the increased influx into the blood from the bone marrow and to the decreased migration from the blood vessels, leading to a reduction in the number of cells at the site of inflammation. The reduction in circulating lymphocytes, monocytes, eosinophils, and basophils is primarily the result of their movement from the vascular bed to lymphoid tissue.

Glucocorticoids also inhibit the functions of tissue mac-rophages and other antigen-presenting cells. The ability of these cells to respond to antigens and mitogens is reduced. The effect on macrophages is particularly marked and limits their ability to phagocytose and kill microorganisms and to produce tumornecrosis factor-α, interleukin-1, metalloproteinases, and plasmi-nogen activator. Both macrophages and lymphocytes produce less interleukin-12 and interferon-γ, important inducers of TH1 cell activity, and cellular immunity.

In addition to their effects on leukocyte function, glucocorti-coids influence the inflammatory response by reducing the pros-taglandin, leukotriene, and platelet-activating factor synthesis that results from activation of phospholipase A₂. Finally, gluco-corticoids reduce expression of cyclooxygenase-2, the inducible form of this enzyme, in inflammatory cells, thus reducing the amount of enzyme available to produce prostaglandins.

Glucocorticoids cause vasoconstriction when applied directly to the skin, possibly by suppressing mast cell degranulation. They also decrease capillary permeability by reducing the amount of histamine released by basophils and mast cells.

The anti-inflammatory and immunosuppressive effects of glu-cocorticoids are largely due to the actions described above. In humans, complement activation is unaltered, but its effects are inhibited. Antibody production can be reduced by large doses of steroids, although it is unaffected by moderate doses (eg, 20 mg/d of prednisone).

The anti-inflammatory and immunosuppressive effects of these agents are widely useful therapeutically but are also respon-sible for some of their most serious adverse effects (see text that follows).

F. Other Effects

Glucocorticoids have important effects on the nervous system. Adrenal insufficiency causes marked slowing of the alpha rhythm of the electroencephalogram and is associated with depression. Increased amounts of glucocorticoids often produce behavioral disturbances in humans: initially insomnia and euphoria and sub-sequently depression. Large doses of glucocorticoids may increase intracranial pressure (pseudotumor cerebri).

Glucocorticoids given chronically suppress the pituitary release of ACTH, growth hormone, thyroid-stimulating hormone, and luteinizing hormone.

Large doses of glucocorticoids have been associated with the development of peptic ulcer, possibly by suppressing the local immune response against Helicobacter pylori. They also promote fat redistribution in the body, with increase of visceral, facial, nuchal, and supraclavicular fat, and they appear to antagonize the effect of vitamin D on calcium absorption. The glucocorticoids also have important effects on the hematopoietic system. In addi-tion to their effects on leukocytes, they increase the number of platelets and red blood cells.

Cortisol deficiency results in impaired renal function (particu-larly glomerular filtration), augmented vasopressin secretion, and diminished ability to excrete a water load.

Glucocorticoids have important effects on the development of the fetal lungs. Indeed, the structural and functional changes in the lungs near term, including the production of pulmonary sur-face-active material required for air breathing (surfactant), are stimulated by glucocorticoids.