I

Beta Switch Program

Beta Switch Program

Get Instant Access

Figure 18.3 Use of 11a-3H-cortisol as a tracer to measure 11 ^-HSD2 activity in vivo. 11 ^-dehydrogenase activity is assessed from the rate of accumulation of 3H-HO

Cortisol

Figure 18.3 Use of 11a-3H-cortisol as a tracer to measure 11 ^-HSD2 activity in vivo. 11 ^-dehydrogenase activity is assessed from the rate of accumulation of 3H-HO

Table 18.3 Examples of members of the steroid/thyroid intracellular receptor superfamily and the enzymes which modulate access of ligands

Receptor

Principal ligand

Enzyme modulating ligand concentration

Glucocorticoid receptor Mineralocorticoid receptor

Oestrogen receptor(s)

Androgen receptor Thyroid hormone receptor

Cortisol Aldosterone Cortisol Oestradiol

Dihydrotestosterone Triiodothyronine

Aromatase 5a-reductase type 2 5'-monodeiodinase responsiveness is modulated by a combination of local control of receptor expression, pre-receptor enzyme expression, and availability of key transcription factors (58). Relatively recent research has established that pre-receptor metabolism of glucocorticoids is also an important determinant of tissue-specific responses. There is strong evidence for such a role for 11^-HSD enzymes; the potential for other cortisol metabolizing enzymes to modulate local corticosteroid receptor activation has yet to be addressed.

Mineralocorticoid Receptor Activation

Although corticosteroid receptors were initially classified according to their in vivo binding characteristics as 'mineralocorticoid' (type I) or 'glucocorticoid' (type II) receptors, when the mineralocorticoid receptor was cloned and expressed it was found to have remarkable sequence homology with the glucocorticoid receptor and to bind cortisol and aldosterone with equal affinity (59). Moreover, in vivo the mineralocorticoid receptor was shown to bind glucocorticoids in some tissues (e.g. hippocampus) but not in others (e.g. distal nephron) (60), despite glucocorticoid concentrations in plasma being two to three orders of magnitude higher than concentrations of mineralocorticoids. This paradox has been explained on the basis of the tissue-specific expression of 11£-HSD2 (24,61).

There are circumstances in which activity of 11^-HSD2 is impaired, including congenital mutations in the 11^-HSD2 gene (62), pharmacological inhibition of the enzyme (e.g. with liquorice derivatives such as glycyrrhetinic acid or carbenoxolone) (63,64), or transgenic disruption of the gene in mice

(65). In these circumstances, glucocorticoids gain inappropriate access to mineralocorticoid receptors (24,61), resulting in profound sodium retention and potassium wasting, with hypertension and hy-pokalaemic alkalosis. This occurs despite low concentrations of aldosterone, and has been termed the syndrome of 'apparent' mineralocorticoid excess

(66). Diagnosis of this syndrome is discussed further

Figure 18.4 Plasma Cortisol and cortisone following oral administration of cortisone in humans. Data are from 10 healthy women (open symbols) who received dexamethasone 250 ^g orally at 2300 h then cortisone 25 mg by mouth at 0830 h. Plasma cortisol and cortisone were measured at the intervals shown after cortisone. Note the inconsistent and relatively small rise in plasma cortisone by contrast with the substantial rise in cortisol. This is consistent with avid 11 ^-HSD1 conversion of cortisone to cortisol on first pass metabolism through the liver. Data are also shown for a 36-year-old female patient (solid symbols) with probable congenital 11^-HSD1 deficiency. Note the very poor generation of cortisol and early peak of cortisone. Adapted from Jamieson et al. (93)

Figure 18.4 Plasma Cortisol and cortisone following oral administration of cortisone in humans. Data are from 10 healthy women (open symbols) who received dexamethasone 250 ^g orally at 2300 h then cortisone 25 mg by mouth at 0830 h. Plasma cortisol and cortisone were measured at the intervals shown after cortisone. Note the inconsistent and relatively small rise in plasma cortisone by contrast with the substantial rise in cortisol. This is consistent with avid 11 ^-HSD1 conversion of cortisone to cortisol on first pass metabolism through the liver. Data are also shown for a 36-year-old female patient (solid symbols) with probable congenital 11^-HSD1 deficiency. Note the very poor generation of cortisol and early peak of cortisone. Adapted from Jamieson et al. (93)

Glucocorticoid Receptor Activation

More recent studies have addressed the possibility that pre-receptor reactivation of cortisol from cortisone by 11^-HSDi may also regulate intracellular receptor activation. As above, 11^-HSD1 is expressed in sites which are not classical targets for aldosterone, but which are important sites where glucocorticoids modulate carbohydrate and lipid metabolism, including liver and adipose tissue. Surprisingly, the affinity of glucocorticoid receptors for cortisol is 10-40-fold lower than that of min-eralocorticoid receptors (59). Reactivation of cortisone to cortisol may therefore be an important mechanism for maintaining adequate exposure of glucocorticoid receptors to their endogenous ligand in key target sites, especially during the trough of circulating cortisol levels at night.

In Liver

In support of this hypothesis, pharmacological inhibition of 11^-HSD1 reductase activity with car-benoxolone in humans (67) or oestrogen administration in male rats (68), or transgenic disruption of the 11^-HSD1 gene in mice (69), results in altered carbohydrate metabolism consistent with impaired activation of hepatic glucocorticoid receptors (Figure 18.6). These receptors have numerous interactions with insulin in regulating hepatic glucose metabolism (1), including upregulation of the rate-limiting enzyme in gluconeogenesis, phosphenol-pyruvate carboxykinase (PEPCK). Impaired 11^-HSD1 activity is characterized by impaired PEPCK expression and/or activity, either at baseline or in response to the stimulus of fasting (Figure 18.6). Similarly, induction of another hepatic glucocorticoid-regulated gene product, glucose-6-phosphatase, is also deficient in 11^-HSD1 null mice (69).

below. Thus, 11^-HSD2 activity is important in preventing glucocorticoids from gaining access to mineralocorticoid receptors. This mechanism explains the selectivity of mineralocorticoid receptors for aldosterone in sites where 11^-HSD2 is active, including distal nephron, sweat glands, salivary glands and colonic mucosa. In sites where 11^-HSD2 is not expressed, such as hippocampus, glucocorticoids readily gain access to mineralocor-ticoid receptors in vivo.

In Adipose Tissue

A key question is whether 11^-HSD1 influences glucocorticoid receptor activation in other sites as well as in liver. The 11^-HSD inhibitor car-benoxolone enhances insulin sensitivity in healthy humans (Figure 18.6) (67). This could not be attributed to altered uptake of glucose in skeletal muscle, as measured by forearm arteriovenous glucose uptake, so it must result from enhanced sup-

Figure 18.5 Regulation of gene transcription by corticosteroid receptors ll/?-HSD, 11/Miydroxysteroid dehydrogenases; GR, glucocorticoid receptor; ~~|/1/7zinc fingers, which bind to DNA; 90, 70, 56, and 26 indicate heatshock proteins of these molecular weights, in kDa, which dissociate from the activated receptor; TF, transcription factor, e.g. AP-1; GRE, glucocorticoid response element, typically a palindromic sequence to which the receptors bind; LEM1, a yeast transmembrane transporter; CBG, corticos-teroid-binding globulin

Figure 18.5 Regulation of gene transcription by corticosteroid receptors ll/?-HSD, 11/Miydroxysteroid dehydrogenases; GR, glucocorticoid receptor; ~~|/1/7zinc fingers, which bind to DNA; 90, 70, 56, and 26 indicate heatshock proteins of these molecular weights, in kDa, which dissociate from the activated receptor; TF, transcription factor, e.g. AP-1; GRE, glucocorticoid response element, typically a palindromic sequence to which the receptors bind; LEM1, a yeast transmembrane transporter; CBG, corticos-teroid-binding globulin

Figure 18.6 Evidence that 11 ^-HSD1 influences insulin sensitivity in humans, mice and rats. (a) Comparison of results of euglycaemic hyperinsulinaemic clamps in healthy men receiving placebo or the 11^-HSD inhibitor carbenoxolone (CBX, 100 mg by mouth 8-hourly for 7 days) in a randomized double-blind cross-over study. Carbenoxolone increased insulin sensitivity by 11.3%. Insulin sensitivity is represented by M/I, which is the rate of glucose infusion during hyperglycaemia (M value) divided by prevailing insulin concentration. Adapted from Walker et al. (67).

(b) Comparison of hepatic phosphoenolpyruvate carboxykinase (PEPCK) activity in wild-type mice and 11^-HSD1 knockout mice under fasted and fed conditions. Although basal activity is not different, 11^-HSD1 knockout mice have less induction of gluconeogenic enzymes when fasted. Adapted from Kotelevtsev et al. (69).

(c) Comparison of the effect of oestradiol (E2) administration on hepatic PEPCK mRNA measured by Northern blot in rats with and without corticosterone. In the absence of corticosterone, oestradiol increases expression of gluconeogenic enzymes. However, in the presence of corticosterone, oestradiol (which potently represses 11^-HSD1 expression and activity in these circumstances) lowers PEPCK mRNA levels. ADX, adrenalectomy. Adapted from Jamieson et al. (68)

Figure 18.6 Evidence that 11 ^-HSD1 influences insulin sensitivity in humans, mice and rats. (a) Comparison of results of euglycaemic hyperinsulinaemic clamps in healthy men receiving placebo or the 11^-HSD inhibitor carbenoxolone (CBX, 100 mg by mouth 8-hourly for 7 days) in a randomized double-blind cross-over study. Carbenoxolone increased insulin sensitivity by 11.3%. Insulin sensitivity is represented by M/I, which is the rate of glucose infusion during hyperglycaemia (M value) divided by prevailing insulin concentration. Adapted from Walker et al. (67).

(b) Comparison of hepatic phosphoenolpyruvate carboxykinase (PEPCK) activity in wild-type mice and 11^-HSD1 knockout mice under fasted and fed conditions. Although basal activity is not different, 11^-HSD1 knockout mice have less induction of gluconeogenic enzymes when fasted. Adapted from Kotelevtsev et al. (69).

(c) Comparison of the effect of oestradiol (E2) administration on hepatic PEPCK mRNA measured by Northern blot in rats with and without corticosterone. In the absence of corticosterone, oestradiol increases expression of gluconeogenic enzymes. However, in the presence of corticosterone, oestradiol (which potently represses 11^-HSD1 expression and activity in these circumstances) lowers PEPCK mRNA levels. ADX, adrenalectomy. Adapted from Jamieson et al. (68)

Figure 18.7 110-HSD activity in 3T3 adipocyte cell line. 110-HSD activities were measured in the human adipocyte-derived 3T3 cell line, both in cells with the undifferentiated pre-adipocyte phenotype and in cells differentiated to adipocyte phenotype with insulin/cortisol. In both states, 110-reductase activity (reactivation of cortisone to cortisol) is very much greater than 110-dehydrogenase activity (inactivation of cortisol to cortisone). Adapted from Napolitano et al. (33)

Figure 18.7 110-HSD activity in 3T3 adipocyte cell line. 110-HSD activities were measured in the human adipocyte-derived 3T3 cell line, both in cells with the undifferentiated pre-adipocyte phenotype and in cells differentiated to adipocyte phenotype with insulin/cortisol. In both states, 110-reductase activity (reactivation of cortisone to cortisol) is very much greater than 110-dehydrogenase activity (inactivation of cortisol to cortisone). Adapted from Napolitano et al. (33)

pression of hepatic gluconeogenesis and/or enhanced uptake of glucose in adipose tissue. Experiments to differentiate these possibilities have yet to be reported. The identification of 110-HSD1 conversion of cortisone to cortisol in adipose stromal cells in primary culture (4), and equivalent activity in a cultured adipocyte cell line (Figure 18.7) (33), has received particularly close attention. Inhibition of this enzyme prevents reactivation of cortisone and glucocorticoid regulation of aromatase expression (70). In human tissue culture this activity is higher in cells obtained from omental fat than subcutaneous fat (4). It has been hypothesized that local regeneration of cortisol in adipose tissue explains the characteristic visceral distribution of fat in Cushing's syndrome, and may explain central adiposity in other circumstances, including growth hormone deficiency (71). In rats, there is no such regional difference in 110-HSD1 activity in fat (72), but perhaps this explains why rats do not develop glucocorticoid-dependent visceral obesity.

In Brain and Pituitary: Control of Appetite and Hypothalamic-Pituitary-Adrenal Axis

Glucocorticoid receptors are expressed widely in most neurons and glia (73). In contrast, min-eralocorticoid receptors are expressed at high levels only in neurons of the hippocampus, septum and a few nuclei of the brainstem. At both receptors, glucocorticoids are involved in altering target gene expression, thereby affecting cellular metabolism and electrophysiological properties, and hence mood, memory and neuroendocrine parameters, as well as cell division, maturation, structure and survival. Indeed glucocorticoids have well-documented effects upon their own release pathways, acting in negative feedback control upon the stimulatory hypothalamic-pituitary-adrenal (HPA) axis. Such feedback occurs at both pituitary and brain levels, most notably in the paraventricular nucleus of the hypothalamus (PVN) and in key supra-hy-pothalamic sites including the hippocampus (74). This feedback is mediated by glucocorticoid receptors, but there may also be a contribution, especially at low cortisol concentrations, from mineralocor-ticoid receptors (75).

Adrenal insufficiency and glucocorticoid replacement therapy are associated with alterations in appetite and food intake in humans and in animal models. The mechanisms are rather poorly defined, and may include both glucocorticoid and min-eralocorticoid receptor effects. The critical neurons are probably located primarily in specific regions of the hypothalamus, notably the PVN and arcuate nucleus (76). In particular, glucocorticoids appear essential for the increased appetite associated with food (protein) deprivation (77). Glucocorticoid receptors are perhaps more selectively involved in carbohydrate appetite control, at least in rodents, whereas mineralocorticoid receptors have been more implicated in fat appetite (78,79). However, such mechanistic studies have been weakened by the predominant use of peripherally administered steroids which may exert varied and indirect effects upon the CNS.

110-HSD1 is expressed in neurons of the hippocampus, hypothalamus (including the parvocel-lular part of the PVN where corticotrophin-releas-ing hormone (CRH) is synthesized) and anterior pituitary (37-39). Within hippocampal cells, at least, 110-HSD1 functions as a predominant 110-reductase in vitro (29). Indeed, this glucocorticoid regenerating enzyme appears important in amplifying the deleterious effects of elevated glucocorticoid levels upon neuronal survival in culture. Moreover, 110-HSD1 null mice show elevated basal levels of glucocorticoids, implying reduced effective glucocorticoid feedback control of the HPA axis (69).

That this occurs in the face of modestly elevated basal plasma glucocorticoid levels suggests that in-tracellular regeneration of active 11-hydroxy steroids is an important source of the total glucocor-ticoid signal in the brain as well as in peripheral tissues.

11^-HSD2 has a more restricted distribution than 11^-HSD1 in adult brain. The absence of 11^-HSD2 dehydrogenase activity explains why both glucocorticoid and mineralocorticoid receptors bind cortisol and corticosterone in hippocampus in vivo (80). However, 11^-HSD2 is expressed in a limited population of cells in the ventromedial nucleus of the hypothalamus in adult rats (81,82). The dehydrogenase isozyme is also expressed in the central nucleus of the amygdala (81,82), where it is appropriately located to facilitate the specific central actions of aldosterone, but not corticosterone, upon salt appetite (83,84).

The role, if any, in regulation of central appetite of the 11^-HSD enzymes (85), or of other key players in corticosteroid metabolism, such as 5a-reductase, remains unexplored, but could be addressed with the recent construction of 5a-reductase type 1 (86), 110-HSD1 (69), and 11£-HSD2 (65) knockout mice. Brain-selective, enzyme deficient mice would help to specify the exact locus of any effects seen.

In Other Sites: Further Local and Systemic Modulation of Cortisol Availability

11^-HSDs may also be important in a host of other sites, including in lymphoid tissue where they may modulate cell-mediated immunity (87,88), and in blood vessels where they may modulate glucocor-ticoid-induced vasoconstriction (89-91).

The fact that tissue-specific differences in cortisol metabolism are important in determining local in-tracellular corticosteroid receptor activation does not lessen the importance of regulation of circulating cortisol concentrations by the HPA axis (Figure 18.1). Clearly, circulating levels of cortisol—and cortisone—remain important determinants of in-tracellular cortisol concentrations. There are two ways in which peripheral cortisol metabolism influences feedback regulation of the HPA axis: by influencing local glucocorticoid concentrations in sites where feedback occurs (see above); and by influencing circulating glucocorticoid concentrations.

The metabolic clearance rate for cortisol could play a key role in determining plasma levels of cortisol and resulting feedback regulation of the HPA axis. In circumstances of impaired peripheral inactivation of cortisol (e.g. 11ß-HSD2 deficiency (66)), total cortisol production rate falls in order to maintain normal plasma cortisol levels. In circumstances of enhanced cortisol clearance, e.g. 11ß-HSD1 deficiency with impaired regeneration of cortisol (92,93), plasma cortisol tends to fall but this is corrected by enhanced ACTH-dependent cortisol secretion. A consequence of this effect is increased secretion of other ACTH-dependent steroids, including adrenal androgens, so that these very rare patients present with hirsutism and menstrual irregularity, as described below. Alterations in other metabolic pathways, e.g. A-ring reductases, is predicted to influence cortisol clearance and negative feedback regulation of the hypothalamic-pituitary-adrenal axis but this has not been subject to experimentation.

Regulation of Cortisol Metabolism

Given the importance of peripheral metabolism in determining tissue-specific differences in response to corticosteroids one would expect exquisite regulation of these enzymes. This may in turn give clues to their dysregulation in obesity.

11^-HSD2 appears to be constitutive, required at all times in the adult to protect mineralocorticoid receptors from inappropriate activation by glucocorticoids. However, during embryonic development, this isozyme is expressed much more widely (12,13,94,95). High fetal tissue expression of 11^-HSD2 is dramatically lost in most organs at the end of midgestation in rodents and humans, presumably allowing tissue exposure to glucocorticoids to occur. The signals for these dramatic and acute changes in 11^-HSD2 expression remain to be elucidated.

11^-HSD1 gene expression and/or activity in adulthood is regulated by a number of factors (Table 18.4). These include factors which are potentially important mediators of the insulin resistance and dyslipidaemia of obesity such as growth hormone, insulin, tumour necrosis factor-a (TNFa), and interleukin-6 (IL-6), as well as glucocorticoids themselves. These effects may be mediated in large

Table 18.4 Regulation of 11ß-HSDs

110-HSD1 11ß-HSD2

Glucocorticoids Oestradiol

Thyroxine Retinoic acid

IL-5, IL-6, TNFa, Dehydroepiandrosterone

Insulin ACTH

Oestradiol Progesterone (191) (172,189,202)

Growth hormone Growth hormone

Stress (203,204) Nitric oxide (205)

IL, interleukin; TNFa, tumour necrosis factor a.

part by the C/EBP family of transcription factors, for which there are multiple functional binding sites in the 5' region of the 110-HSD1 gene (96,97).

5^-Reductase activity is lower in female than male rat livers (98) but it is upregulated by oestrogen (99). 5a-Reductase type 1, the principal isozyme in human liver and fat (43), is usually thought to be 'constitutive' and not regulated hormonally (100,101), but there is some evidence that this isozyme is downregulated by androgens (102,103) more so than by oestrogen (104) so that its activity is higher in female liver (105) and adrenal (106). The type 1 isozyme is not regulated by growth hormone (105,107). 5a-Reductase type 2 is expressed mainly in the prostate, but also in liver, where it is up-regulated by androgens (42), oestrogens (108) and growth hormone (107).

In addition to regulation of expression of enzymes metabolizing cortisol, the activity of these

Table 18.5 Inhibitors of 11^-HSDs

Endogenous Exogenous enzymes has been suggested to be influenced by the prevailing concentrations of a large number of endogenous competitive inhibitors, largely other steroids, sterols and their derived bile acids (Table 18.5). Although present at sufficiently high concentrations to be detected in extracts of urine (109), the relevance of these inhibitors in vivo remains uncertain (110-114).

Metabolism of Synthetic Corticosteroids

Modifications of steroid structures has long been recognized to affect their affinity for corticosteroid receptors. In addition, their affinity for metabolizing enzymes may be altered. Thus, 9a-fluoro-cortisol (also known as fludrocortisone) has preserved affinity for mineralocorticoid and glucocor-ticoid receptors, but the steroid is no longer inactivated by 11^-HSD2 in the distal nephron (115) and is therefore a potent mineralocorticoid. By contrast, prednisolone differs from cortisol in having a 1,2 double bond in the A-ring which increases affinity for glucocorticoid receptors but retains metabolism by 11^-HSDs (116). Dexamethasone and be-clomethasone have a combination of these modifications (9a-fluoro, 16a-methyl and 9a-choloro, 16^-methyl, respectively) which increase affinity for glucocorticoid receptors and lower affinity for mineralocorticoid receptors. Interestingly, the 9a-fluoro modification in dexamethasone appears to dictate metabolism by 11^-HSD1 that is exclusively reductive, even with semi-purified enzyme which will act as an effective 11^-dehyd-rogenase for cortisol and corticosterone (117). This interaction between substrate and 11^-HSD1 is unique, since other substrates are metabolized

Unidentified

Upregulation

Downregulation

11-OH-progesterone (206-208) Glycyrrhetinic acid (63,155,209) 'GALFs'(109,110,112,210-213)

3a,5^-Tetrahydroprogesterone (214) Glycyrrhizic acid ACTH-dependent inhibitors (26)

3a,5ß-Tetrahydro-11-deoxy-corticosterone Carbenoxolone (64)

11-Epicortisol CHAPS (215)

Chenodeoxycholic acid Ketoconazole

Cholic acid (216,217) Saiboku-To (218,219)

Gossypol (220,221) Metyrapone (222-224) 11-Epiprednisolone Frusemide (225-227) Bioflavinoids (228,229)

Table 18.6 Biochemical features of deranged 11^-HSD activity

Apparent Apparent mineralocorticoid mineralocorticoid Apparent cortisone Liquorice Carbenoxolone excess type 1 excess type 2 reductase deficiency administration administration

11 fi-dehydrogenase Half life of (11a3H)-cortisol 11 fi-reductase: Conversion of oral cortisone to cortisol

Urinary cortisol metabolites: (5fi- + 5a-THF)/THE ratio 5a-/5fi-THF ratio Cortisol/Cortisone ratio Total cortisol metabolites

No data

Was this article helpful?

0 0
Spiritual Weight Loss Mentality

Spiritual Weight Loss Mentality

Awesome Ways To Get Over Your Mentality That Keeps you Overweight! This Book Is One Of The Most Valuable Resources In The World When It Comes To Results In Your Slim-down and Health Efforts! Day in day out we keep ourselves absorbed with those matters that matter the most to us. A lot of times, it might be just to survive and bring in some money. In doing so we at times disregard or forget about the extra matters that are essential to balance our lives. They’re even more essential to supply real meaning to our world. You have to pay attention to your wellness.

Get My Free Ebook


Post a comment