5α-Reduced glucocorticoids: a story of natural selection

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   Figure 1

   Glucocorticoid (GC) effects on inflammatory signalling. GCs act through several mechanisms to exert anti-inflammatory effects: 1) non-genomic pathways involve GC receptor (GR)-mediated direct interactions with second messenger proteins, including the MAPK protein JNK, inhibiting the activation of this signalling pathway. 2) GR-mediated transactivation of key anti-inflammatory genes involves direct DNA binding of both GR dimers and monomers/multimers to GC-response elements (GRE) in the promoter region of target gene. 3) Transrepression of pro-inflammatory genes does not require direct DNA binding of GR, but rather ‘tethering’ of GR monomers to DNA-bound pro-inflammatory transcription factors.
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   Figure 2

   Metabolism of cortisol and corticosterone to generate 5α-reduced glucocorticoids. Generic scheme of reaction catalysed by 5α-reductases (5αRs) followed by 3α-hydroxysteroid dehydrogenases (3αHSDs). Specific reactions relating to cortisol, corticosterone and testosterone are shown.
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   Figure 3

   5α-Reduced glucocorticoid binds and selectively activates GR. (A) 5α-Tetrahydrocorticosterone (5αTHB) and 5α-dihydrocorticosterone (5αDHB) (overnight incubation with concentration range from 10⁻⁵ to 10⁻⁹ M) displaced tritiated dexamethasone (100 nM) from glucocorticoid receptors in rat hepatic cytosol. The K_ds for binding of 5αTHB (268 nM) and 5αDHB (336 nM) were similar to that of corticosterone (153 nM) and higher than dexamethasone (38 nM; n=6/treatment). (B) 5αTHB, but not 5αDHB, induced transcription of tyrosine aminotransferase (TAT) to a lesser extent than corticosterone (B) when incubated for 24 h in H4iiE cells (rat hepatoma cells). Abundance of transcript was quantified by northern blot and normalised for that of U1, n=6/treatment. *P<0.05 vs vehicle (veh), ^#P<0.05 vs B. Following chronic infusion of steroids (50 μg/day, 2 weeks) to mice (n=10–12/group): (C) corticosterone, but not 5αTHB, impaired glucose tolerance, as demonstrated by increased insulin concentrations (P<0.001 vs vehicle) after a glucose tolerance test (2 g/kg body weight i.p.) performed in mice following a 6 h fast. Insulin was quantified by immunoassay. (D) 5αTHB caused immune suppression to a similar extent to corticosterone, assessed by a significant reduction (P<0.01) in the ability of lipopolysaccharide (LPS; 0.01–100 ng/ml)) to induce release of interleukin 6 (IL6) following incubation (24 h) from cells in whole blood harvested at cull. IL6 was quantified by immunoassay. Data are mean±s.e.m. Data presented in A and B are adapted from those originally published in the McInnes KJ, Kenyon CJ, Chapman KE, Livingstone DEW, Macdonald LJ, Walker BR & Andrew R 2004 5α-Reduced glucocorticoid metabolites, novel endogenous activators of glucocorticoid receptors (GR). Journal of Biological Chemistry 279 22908–22912 © the American Society for Biochemistry and Molecular Biology. Data presented in C and D were originally published in the Yang CA, Nixon M, Kenyon CJ, Livingstone DEW, Duffin R, Rossi AG, Walker BR & Andrew R 2011 5α-Reduced glucocorticoids exhibit dissociated anti-inflammatory and metabolic effects. British Journal of Pharmacology. (In press) (doi:10.1111/j.1476-5381.2011.01465.x).
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   Figure 4

   Binding of testosterone and 5α-dihydrotestosterone (5αDHT) in the androgen receptor ligand-binding domain (AR-LBD). When bound with AR, testosterone (A) and 5αDHT (B) are positioned near the side chain of Arg-752 (R752), a helix 5 residue required for ligand binding. The bond between the 3-keto-O on the steroid and R752 is influenced by the 4–5 double bond in testosterone, which imparts a more planar structure to testosterone than that seen in 5αDHT (C, dashed arrow), allowing more favourable H-bonding. The presence of structural water HOH1 permits a H-bond network between the steroid and key residues in the AF2 transactivation domain of the receptor. In particular, a bridged H-bond with Met-745 (M745), a residue that lies directly above the A-ring of the steroid, is formed. The structure of this bond is believed to be important as M745 projects towards Leu712 (L712), a proximal residue in the AF2 domain crucial in mediating contacts with residues within accessory molecules. Alterations in the structural integrity of L712 as a result of this projection (C, Block arrow) are believed to directly affect the binding of key cofactors essential for gene transcription. 5αDHT has been suggested to impart greater structural integrity to L712, stabilising cofactor binding and thus enhancing AR activity (Askew et al. 2007). Images were generated with ICM-Pro Software (MolSoft, San Diego, CA, USA) for testosterone with AR-LBD (Protein Data Bank Code 2Q7I) and 5αDHT with AR-LBD (Protein Data Bank Code 1T63).

• © 2012 Society for Endocrinology