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β-cyclodextrins derivatives (CDs) have applied in steroids biotransformation industry because of their unique properties. However, the effect of β-CDs on the growth rate, activity, conversion, and characters of whole cells has not been concerned. In this study, the growth rate and cellular morphology of Arthrobacter simplex (ASP) pretreated by six kinds of β-CDs were measured. The results showed that most β-CDs inhibited the growth of ASP, among which randomly methylated-β-CDs has the most serve inhibition; however, sulfonic acid-β-CD promoted the growth of cells. The morphology size and the surface of all β-CDs-pretreated cells were changed compared with the control group. Besides, the conversion of cortisone acetate (CA) increased in β-CDs-pretreatment system and β-CDs-containing system, which reached 97.98% in hydroxypropyl-β-cyclodextrin (HP-β-CD) containing-system and 78.69% in HP-β-CD-pretreatment-system, but the dehydrogenase activity of all β-CDs-pretreated cells decreased. β-CDs with higher K value have stronger inclusion ability with CA, and along with the membrane permeability of β-CDs-pretreated cells increased more, but they also have more serve damage on the ASP cells, which is negative to increase the conversion of CA. This study improved our understanding of the effect of β-CDs when they were used in the steroids biotransformation by ASP whole cells, and provided data basis for the selection of suitable CDs for application.

Background LC-MS/MS methods offer high selectivity in cortisol determinations. However, endogenous steroid metabolites may still interfere and compromise the results, for example in the diagnosis of Cushing's syndrome. Erroneously elevated cortisol may, in particular, be misleading at the low concentrations found in salivary samples obtained at late night and after dexamethasone suppression. Methods Interferences in our LC-MS/MS method used for determination of cortisol in saliva and urine were identified by comparing their retention times and mass spectra with those of pure candidate substances. The chromatographic conditions used in our LC-MS/MS method, including column and mobile phase gradient, were varied in order to separate the target compound from the interferences. Results Two interferences, which were co-eluting or eluting close to cortisol in our original method, were successfully separated from cortisol by adjustment of the chromatographic conditions. These interferences were found in both urine and saliva and were identified as the two endogenous cortisol isomers 20α- and 20β-dihydrocortisone. The isomers share molecular mass and mass spectrometric fragmentation pattern with cortisol using electrospray ionization in the positive-ion mode. Both give rise to the transitions m/z 363.1>121.1, 363.1>115.1 and 363.1>97.1. In our original LC-MS/MS setup, the 20β-dihydrocortisone co-eluted with cortisol in the chromatography step resulting in false high determinations. Conclusions Cortisol determination by LC-MS/MS may suffer from erroneously elevated results unless 20α- and 20β-dihydrocortisone are chromatographically separated from cortisol.

DHRS7 (SDR34C1) has been associated with potential tumor suppressor effects in prostate cancer; however, its function remains largely unknown. Recent experiments using purified recombinant human DHRS7 suggested several potential substrates, including the steroids cortisone and Δ4-androstene-3,17-dione (androstenedione). However, the substrate and cofactor concentrations used in these experiments were very high and the physiological relevance of these observations needed to be further investigated. In the present study, recombinant human DHRS7 was expressed in intact HEK-293 cells in order to investigate whether glucocorticoids and androgens serve as substrates at sub-micromolar concentrations and at physiological cofactor concentrations. Furthermore, the membrane topology of DHRS7 was revisited using redox-sensitive green-fluorescent protein fusions in living cells. The results revealed that (1) cortisone is a substrate of DHRS7; however, it is not reduced to cortisol but to 20β-dihydrocortisone, (2) androstenedione is not a relevant substrate of DHRS7, (3) DHRS7 catalyzes the oxoreduction of 5α-dihydrotestosterone (5αDHT) to 3α-androstanediol (3αAdiol), with a suppressive effect on androgen receptor (AR) transcriptional activity, and (4) DHRS7 is anchored in the endoplasmic reticulum membrane with a cytoplasmic orientation. Together, the results show that DHRS7 is a cytoplasmic oriented enzyme exhibiting 3α/20β-hydroxysteroid dehydrogenase activity, with a possible role in the modulation of AR function. Further research needs to address the physiological relevance of DHRS7 in the inactivation of 5αDHT and AR regulation., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Evogliptin is a novel potent and selective dipeptidyl peptidase-4 (DPP-4) inhibitor. The aim of the present study was to evaluate the pharmacokinetic (PK) and pharmacodynamic (PD) characteristics of evogliptin in participants with renal impairment (RI). An open-label, parallel-group clinical study was conducted in participants with mild, moderate and severe RI and in matched participants with normal renal function (NRF). A single oral 5-mg dose of evogliptin was administered and serial blood and urine samples were obtained to assess the PK and PD characteristics of evogliptin. Baseline urine samples were collected to evaluate endogenous CYP3A metabolic markers. The plasma exposure to evogliptin and degree of DPP-4 activity inhibition increased with decreasing renal function. The mean areas under the concentration-time curves from 0 to 120 hours were increased 1.2-, 1.8- and 1.98-fold in the mild, moderate and severe RI groups, respectively, compared with the NRF group. The levels of CYP3A metabolic markers were lower in the RI group than in the NRF group. The increase in the plasma concentration of evogliptin is unlikely to result in changes in its efficacy or safety, considering the results of previous clinical studies., (© 2016 John Wiley & Sons Ltd.)

Cyclodextrins (CDs) can improve the productivity of steroid biotransformation by enhancing substrate solubility. CDs can be recycled by grafting them with appropriate carriers. Loofah fiber is an excellent grafting material for CDs, and can be applied to the biotransformation and recycling of β-cyclodextrin (β-CD). In this work, a technique for recycling β-CD in cortisone acetate (CA) biotransformation by Arthrobacter simplex CPCC 140451 was studied. Loofah fiber-grafted β-CD (LF-β-CD) was prepared using epichlorohydrin, which is a cross-linking agent. The grafting yield of β-CD was 74.8 mg g -1 dried fibers. LF-β-CD could increase the solubility of CA and enhance biotransformation. The initial conversion rate of CA was 1.5-fold higher than that of the blank group. LF-β-CD was also used in biocatalytic reactions for eight cycles, and it maintained the conversion ratio of CA at approximately 90%. Given the above positive results, LF-β-CD can be utilized in biotechnological recycling applications. This method can also be applied to CD derivatives and hydrophobic compounds.

In this work, the ultrahigh-performance liquid chromatography quadrupole orbitrap high resolution mass spectrometry (UHPLC-Q-Orbitrap HRMS) was applied to the rapid screening, identification and quantification of the illegal adulterated glucocorticoids in herbal medicines. The mass spectrometer was operated in positive ion mode and Full MS/dd-MS 2 (data-dependent MS 2 ) mode, where selected ions were subjected to a dd-MS 2 scan with given fragmentation energy following a Full MS scan. The application of 70 000 FWHM mass resolution and narrow mass windows (5ppm) effectively improve the selectivity of the method, and a single injection was sufficient to perform the simultaneous screening and identification/quantification of 14 glucocorticoids in 15min. The method validation including selectivity, sensitivity, calibration curve, accuracy, precision, recovery, matrix effect and stability were evaluated. The results of all analytes showed excellent linear relationship while all coefficient of determination (r 2 ) were>0.9990 over wide concentration ranges (e.g., 5-1000ng/mL for hydrocortisone butyrate, r 2 =1.0000). The recoveries were in the range of 86.1-102.7%, while the matrix effects ranged from 95.8%-105.8%. Accuracies and precisions were performed. The intra- and inter-day accuracies ranged from 90.6% to 108.9%, while the intra- and inter-day precisions were in the range of 0.5% to 8.5%. Finally, the established method was employed to detect illegal adulterated glucocorticoids in herbal medicines. It will provide more reliable technical basis for the drug quality supervision department and ensure public health., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Background: Inflammation and oedema of the facial nerve are implicated in causing Bell's palsy. Corticosteroids have a potent anti-inflammatory action that should minimise nerve damage. This is an update of a review first published in 2002 and last updated in 2010., Objectives: To determine the effectiveness and safety of corticosteroid therapy in people with Bell's palsy., Search Methods: On 4 March 2016, we searched the Cochrane Neuromuscular Specialised Register, the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, EMBASE and LILACS. We reviewed the bibliographies of the randomised trials and contacted known experts in the field to identify additional published or unpublished trials. We also searched clinical trials registries for ongoing trials., Selection Criteria: Randomised trials and quasi-randomised trials comparing different routes of administration and dosage schemes of corticosteroid or adrenocorticotrophic hormone therapy versus a control group receiving no therapy considered effective for this condition, unless the same therapy was given in a similar way to the experimental group., Data Collection and Analysis: We used standard Cochrane methodology. The main outcome of interest was incomplete recovery of facial motor function (i.e. residual facial weakness). Secondary outcomes were cosmetically disabling persistent sequelae, development of motor synkinesis or autonomic dysfunction (i.e. hemifacial spasm, crocodile tears) and adverse effects of corticosteroid therapy manifested during follow-up., Main Results: We identified seven trials, with 895 evaluable participants for this review. All provided data suitable for the primary outcome meta-analysis. One of the trials was new since the last version of this Cochrane systematic review. Risk of bias in the older, smaller studies included some unclear- or high-risk assessments, whereas we deemed the larger studies at low risk of bias. Overall, 79/452 (17%) participants allocated to corticosteroids had incomplete recovery of facial motor function six months or more after randomisation; significantly fewer than the 125/447 (28%) in the control group (risk ratio (RR) 0.63, 95% confidence interval (CI) 0.50 to 0.80, seven trials, n = 895). The number of people who need to be treated with corticosteroids to avoid one incomplete recovery was 10 (95% CI 6 to 20). The reduction in the proportion of participants with cosmetically disabling sequelae six months after randomisation was very similar in the corticosteroid and placebo groups (RR 0.96, 95% CI 0.40 to 2.29, two trials, n = 75, low-quality evidence). However, there was a significant reduction in motor synkinesis during follow-up in participants receiving corticosteroids (RR 0.64, 95% CI 0.45 to 0.91, three trials, n = 485, moderate-quality evidence). Three studies explicitly recorded the absence of adverse effects attributable to corticosteroids. One trial reported that three participants receiving prednisolone had temporary sleep disturbances and two trials gave a detailed account of adverse effects occurring in 93 participants, all non-serious; the combined analysis of data from these three trials found no significant difference in adverse effect rates between people receiving corticosteroids and people receiving placebo (RR 1.04, 95% CI 0.71 to 1.51, n = 715)., Authors' Conclusions: The available moderate- to high-quality evidence from randomised controlled trials showed significant benefit from treating Bell's palsy with corticosteroids.

Aim: To examine how the endogenous CYP3A4 phenotype and CYP3A5(*)3 genotype of Chinese renal transplant recipients influenced the dose-corrected trough concentration (C0/D) and weight-corrected daily dose (D/W) of tacrolimus., Methods: A total of 101 medically stable kidney transplant recipients were enrolled, and their blood and urine samples were gathered. The endogenous CYP3A4 phenotype was assessed by the ratio of 6β-hydroxycortisol and 6β-hydroxycortisone to cortisol and cortisone in urine. CYP3A5(*)3 genotype was determined using PCR-RELP., Results: In overall renal transplant recipients, a multiple regression analysis including the endogenous CYP3A4 phenotype, CYP3A5(*)3 genotype and post-operative period accounted for 60.1% of the variability in C0/D ratio; a regression equation consisting of the endogenous CYP3A4 phenotype, post-operative period, body mass index, CYP3A5(*)3 genotype, gender, total bilirubin and age explained 61.0% of the variability in D/W ratio. In CYP3A5(*)3/(*)3 subjects, a combination of the endogenous CYP3A4 phenotype, post-operative period and age was responsible for 65.3% of the variability in C0/D ratio; a predictive equation including the endogenous CYP3A4 phenotype, post-operative period, body mass index, gender and age explained 61.2% of the variability in the D/W ratio. Base on desired target range of tacrolimus trough concentrations, individual daily dosage regimen was calculated, and all the observed daily doses were within the predicted range., Conclusion: This study provides the equations to predict tacrolimus metabolism and dosage requirements based on the endogenous CYP3A4 phenotype, CYP3A5(*)3 genotype and other non-genetic variables.

Objective: To establish a method for the recovery and reutilization of hydroxypropyl-β-cyclodextrin (HP-β-CD) to lower the cost of its industrial application in cortisone acetate bioconversion., Results: HP-β-CD is not degraded by Arthrobacter simplex CPCC140451 (ASP) resting cells and 96.4 % HP-β-CD could be recovered by isobutyl acetate extraction. Moreover, the inclusion ability of recovered HP-β-CD barely decreased. The saccharide metabolic and catalytic activities of ASP were greater in the aqueous phase after extracting with isobutyl acetate than other organic solvents. Cyclic utilization tests showed that cortisone acetate conversion ratio was 91.0 % after eight cycles and reached 95.7 % with 0.2-0.6 mM HP-β-CD. Furthermore, >90 % conversion ratio was reached per cycle through a co-cyclic-utilization method with HP-β-CD and immobilized ASP., Conclusion: Cortisone acetate conversion ratio in the HP-β-CD cyclic-utilization method is promising for industrial applications. The method can also be expanded to other CDs and other hydrophobic compounds bioconversion.

Glucocorticoids are included in the S9 section of the World Anti-doping Agency (WADA) prohibited list international standard. Some among them are pseudo-endogenous steroids, like cortisol and cortisone, which present the same chemical structure as endogenously produced steroids. We are proposing an analytical method based on gas chromatography coupled to isotope ratio mass spectrometry (GC-C-IRMS) which allows discrimination between endogenous and synthetic origin of the urinary metabolites of the pseudo-endogenous glucocorticoids. A preliminary purification treatment by high-performance liquid chromatography (HPLC) of the target compounds (TC) (i.e., cortisol, tetrahydrocortisone (THE) 5α-tetrahydrocortisone (aTHE), tetrahydrocortisol (THF), and 5α-tetrahydrocortisol (aTHF)) allows collection of extracts with adequate purity for the subsequent analysis by IRMS. A population of 40 urine samples was analyzed for the TC and for the endogenous reference compounds (ERC: i.e., 11-desoxy-tetrahydrocortisol (THS) or pregnanediol). For each sample, the difference between the delta values of the ERCs and TCs (Δδ values) were calculated and based on that, some decision limits for atypical findings are proposed. The limits are below 3% units except for cortisol. The fit to purpose of the method has been confirmed by the analysis of urine samples collected in two patients under treatment with 25 mg of cortisone acetate (p.o). The samples showed Δδ values higher than 3 for at least 24 h following administration depending on the TC considered. The method can easily be integrated into existing procedures already used for the HPLC purification and IRMS analysis of pseudo-endogenous steroids with androgenic/anabolic activity., (Copyright © 2015 John Wiley & Sons, Ltd.)

The combined clearance of endogenous 6β-hydroxycortisol and 6β-hydroxycortisone is suggested biomarker for in vivo cytochrome P450 3A (CYP3A) activity. We aimed to determine whether the combined clearance of these two markers together with information of biopharmaceutics classification system (BCS) of drugs could be used to predict CYP3A-mediated metabolism of immunosuppressants. The BCS of drug formulations were determined based on the solubility and permeability. Sixty-seven healthy subjects were divided into three groups and group 1 (n = 23), 2 (n = 22), and 3 (n = 22) received oral single dose of cyclosporine, tacrolimus, and sirolimus, respectively. Blood and urine samples were gathered at various times. The combined clearance of 6β-hydroxycortisol and 6β-hydroxycortisone correlated significantly with cyclosporine pharmacokinetics (p < 0.001) after oral dose of a BCS 1 formulation, whereas no relationships were seen after administration of tacrolimus and sirolimus formulations, both of which belonged to BCS 2. Regarding the biopharmaceutical characteristics, the endogenous CYP3A biomarker explains 74.5% of variability in oral cyclosporine clearance between individuals., (© 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.)

The aim of this study was to examine the incidence of adrenal crises (AC) and the prescription of short-acting glucocorticoids (GC) in different geographic areas. To do this we conducted a descriptive study of AC hospitalisations and prescriptions for two GCs (hydrocortisone (HC) and cortisone acetate (CA)), and fludrocortisone acetate (FA), in different geographic areas of Australia between 1999/2000 and 2011/2012, using government databases.There were 2,584 hospital admissions for AC in Australia between 1999/00 and 2011/12 and the corresponding admission rates increased significantly from 7.4 to 11.1/10(6)/year (p<0.001). AC admission rates increased in 5 out of 6 geographic areas. Prescription rates for the combined GCs (HC/CA) increased at an annual rate of between 0.2-2.0% in all areas. All areas had significant (p<0.01) increases in HC prescription rates (4.5% to 13.7% annually) and CA prescription rates decreased in 5 out of the 6 regions (3.5% annual decrease to a 0.5% annual increase). When the geographic areas were combined, there was a significant correlation between the AC admission rates and HC/CA prescription rates (r=0.30, p<0.01). Admissions for AC and GC prescriptions increased significantly in Australia after 1999 and these varied significantly by geographic area. These results suggest that modern recommendations for lower dose, short-acting GC replacement may be of concern and further investigation is warranted., (© Georg Thieme Verlag KG Stuttgart · New York.)

A specific and sensitive method based on high-performance liquid chromatography with ultraviolet absorbance detection (HPLC-UV) was developed for the simultaneous determination of urinary cortisol (F), cortisone (E), 6β-hydroxycortisol (6β-OHF) and 6β-hydroxycortisone (6β-OHE) using dexamethasone as the internal standard. The method involved solid-phase extraction of the five compounds from urine using Oasis HLB Waters cartridges with an elution solvent of ethyl acetate-diethyl ether (5 mL; 4:1, v/v), followed by 1 mol/L of NaOH (1 mL) and 1.0% acetic acid (1 mL). Separation of the five analytes was achieved within 31 min by using a reversed-phase C18 analytical column (200 × 4.6 mm, 5 µm, Agilent). A UV detector operated at 245 nm was used. According to the method validation, inter-run and intra-run precision was below 9.45% and accuracy ranged from 98.16 to 115.50%. The lower limits of quantitation were 5 ng/mL for four analytes. This is the first HPLC method that can simultaneously determine F, E, 6β-OHF and 6β-OHE in human urine. The assay was applied to research the ratio of (6β-OHF + 6β-OHE)/(F + E) as a non-invasive biomarker for the metabolism of tacrolimus., (© The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

In Aspergillus nidulans, the AcuK and AcuM transcription factors form a complex that regulates gluconeogenesis. In Aspergillus fumigatus, AcuM governs gluconeogenesis and iron acquisition in vitro and virulence in immunosuppressed mice. However, the function of AcuK was previously unknown. Through in vitro studies, we found that A. fumigatus ΔacuK single and ΔacuK ΔacuM double mutants had impaired gluconeogenesis and iron acquisition, similar to the ΔacuM mutant. Also, the ΔacuK, ΔacuM, and ΔacuK ΔacuM mutants had similar virulence defects in mice. However, the ΔacuK mutant had a milder defect in extracellular siderophore activity and induction of epithelial cell damage in vitro than did the ΔacuM mutant. Moreover, overexpression of acuM in the ΔacuK mutant altered expression of 3 genes and partially restored growth under iron-limited conditions, suggesting that AcuM can govern some genes independently of AcuK. Although the ΔacuK and ΔacuM mutants had very similar transcriptional profiles in vitro, their transcriptional profiles during murine pulmonary infection differed both from their in vitro profiles and from each other. While AcuK and AcuM governed the expression of only a few iron-responsive genes in vivo, they influenced the expression of other virulence-related genes, such as hexA and dvrA. Therefore, in A. fumigatus, while AcuK and AcuM likely function as part of the same complex, they can also function independently of each other. Furthermore, AcuK and AcuM have different target genes in vivo than in vitro, suggesting that in vivo infection stimulates unique transcriptional regulatory pathways in A. fumigatus., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

The photocatalytic decomposition of cortisone 21-acetate (CA), a model compound for the commonly used steroid, cortisone, was studied. CA was photocatalytically decomposed in a slurry reactor with the initial rates between 0.11 and 0.46 mg L(-1)min(-1) at 10 mg L(-1) concentration, using the following heterogeneous photocatalysts in decreasing order of their catalytic activity: ZnO>Evonik TiO2 P25>Hombikat TiO2>WO3. Due to the lack of ZnO stability in aqueous solutions, TiO2 P25 was chosen for further experiments. The decomposition reaction was found to be pseudo-first order and the rate constant decreased as a function of increasing initial CA concentration. Changing the initial pH of the CA solution did not affect the reaction rate significantly. The decomposition reaction in the presence of the oxidizing sacrificial agent sodium persulfate showed an observed decomposition rate constant of 0.004 min(-1), lower than that obtained for TiO2 P25 (0.040 min(-1)). The highest photocatalytic degradation rate constant was obtained combining both TiO2 P25 and S2O8(2-) (0.071 min(-1)) showing a synergistic effect. No reactive intermediates were detected using LC-MS showing fast photocatalytic decomposition kinetics of CA., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Resting cells of Arthrobacter simplex with 1-en-dehydrogenation ability were prepared and treated by ethanol at subinhibitory concentrations (4%-15%, v/v), then added into the ethanol-free system containing low concentration of cortisone acetate (1 g L(-1)) to produce prednisone acetate by C1,2 dehydrogenation reaction. Results showed that, within the range of ethanol concentration, the initial conversion rate was varied significantly with the concentration of ethanol and the maximum was obtained at 8% (v/v) ethanol, which was increased by 32.6% compared with the control. A series of cell features closely relevant to biotransformation efficiency were further analyzed. It indicated that ethanol acting on cell wall and membrane could be used as a mediator to enhance cell permeability, which facilitated the penetration of substrate across cell barrier within a short time, resulting in the elevated initial conversation rate. The observation of fatty acids composition suggested that the increased unsaturated fatty acids, especially cis-isomers, in the presence of ethanol led to the disorganization of the native arrangement of lipids and thus increased cell permeability. Our findings demonstrated that another facilitation of ethanol was to promote substrate transport into cells by permeabilization, which would provide the guidance in the practical application of organic solvents in steroid biotransformation., (© 2014 International Union of Biochemistry and Molecular Biology, Inc.)

Primary adrenal insufficiency (PAI), or Addison's disease, is a rare, potentially deadly, but treatable disease. Most cases of PAI are caused by autoimmune destruction of the adrenal cortex. Consequently, patients with PAI are at higher risk of developing other autoimmune diseases. The diagnosis of PAI is often delayed by many months, and most patients present with symptoms of acute adrenal insufficiency. Because PAI is rare, even medical specialists in this therapeutic area rarely manage more than a few patients. Currently, the procedures for diagnosis, treatment and follow-up of this rare disease vary greatly within Europe. The common autoimmune form of PAI is characterized by the presence of 21-hydroxylase autoantibodies; other causes should be sought if no autoantibodies are detected. Acute adrenal crisis is a life-threatening condition that requires immediate treatment. Standard replacement therapy consists of multiple daily doses of hydrocortisone or cortisone acetate combined with fludrocortisone. Annual follow-up by an endocrinologist is recommended with the focus on optimization of replacement therapy and detection of new autoimmune diseases. Patient education to enable self-adjustment of dosages of replacement therapy and crisis prevention is particularly important in this disease. The authors of this document have collaborated within an EU project (Euadrenal) to study the pathogenesis, describe the natural course and improve the treatment for Addison's disease. Based on a synthesis of this research, the available literature, and the views and experiences of the consortium's investigators and key experts, we now attempt to provide a European Expert Consensus Statement for diagnosis, treatment and follow-up., (© 2013 The Association for the Publication of the Journal of Internal Medicine.)

This study aimed to evaluate endogenous metabolic markers of hepatic cytochrome P450 (CYP)3A activity in healthy subjects using a metabolomics approach. Twenty-four subjects received the following medication during the following three study periods: 1 mg of i.v. midazolam alone (control phase), 1 mg of i.v. midazolam after 4 days of pretreatment with 400 mg of ketoconazole once daily (CYP3A-inhibited phase), and 2.5 mg of i.v. midazolam after 10 days of pretreatment with 600 mg of rifampicin once daily (CYP3A-induced phase). During each study period, 24 h before and after the administration of midazolam, urine samples were collected at 12-h intervals for metabolomic analyses. We derived an equation to predict midazolam clearance (CL) based on several of these markers. We demonstrated that a combination of the concentrations and ratios of several endogenous metabolites and the CYP3A5*3 genotype is a reliable predictive marker of hepatic CYP3A activity as assessed by i.v. administration of midazolam.

Arthrobacter simplex 156 is a microorganism that is used for steroid drug biotransformation of cortisone acetate (CA) to prednisone acetate (PA). The enzyme 3-ketosteroid-△(1)-dehydrogenase encoded by the ksdD gene plays an important role in the bioconversion process. To further improve the biotransformation efficiencies of the industrial strain, a genetic manipulation system for A. simplex 156 was developed. Additional copies of the ksdD gene under the control of the cat promoter (from pXMJ19) were transferred into the strain A. simplex 156 and integrated into the 16S rDNA sites, yielding a series of recombinant strains. One of these recombinant strains, designated A. simplex M158, exhibited superior properties for CA biotransformation. At the substrate concentration of 83.6 g/l, the highest PA production of the recombinant strain reached 66.7 g/l, which is approximately 32.9 % higher than that of wild-type strains, and the incubation time for CA to PA bioconversion was reduced by 20 h. Southern blotting analysis of the recombinant strain indicated two copies of deregulated ksdD genes were integrated into the 16S rDNA sites, which means two of five 16S rRNA operons were insertionally disrupted in the recombinant strain. However, the disruption of the two 16S rRNA operons did not affect the growth rate of the recombinant strain, which survived and thrived under desired conditions. In addition, the new strain was genetically stable for more than 100 generations without the use of antibiotics for selection. These superior characteristics make the new strain more suitable than the wild-type strain for PA production.

At weaning, the intestinal mucosa surface glycans change from predominantly sialylated to fucosylated. Intestinal adaptation from milk to solid food is regulated by intrinsic and extrinsic factors. The contribution by glucocorticoid, an intrinsic factor, and colonization by microbiota, an extrinsic factor, was measured as the induction of α1,2/3-fucosyltransferase and sucrase-isomaltase (SI) activity and gene expression in conventionally raised, germ-free, and bacteria-depleted mice. In conventionally raised mice, cortisone acetate (CA) precociously accelerated SI gene expression up to 3 weeks and fut2 to 4 weeks of age. In germ-free mice, CA treatment induces only SI expression but not fucosyltransferase. In post-weaning bacteria-deficient (germ-free and bacteria-depleted) mice, fut2 expression remains at low suckling levels. In microbiota deficient mice, intestinal fut2 (but not fut1, fut4 or fut7) was induced only by adult microbiota, but not immature microbiota or CA. Fut2 induction could also be restored by colonization by Bacteroides fragilis, but not by a B. fragilis mutant unable to utilize fucose. Restoration of fut2 expression (by either microbiota or B. fragilis) in bacteria-depleted mice is necessary for recovery from dextran sulfate sodium-induced mucosal injury. Thus, glucocorticoids and microbes regulate distinct aspects of gut ontogeny: CA precociously accelerates SI expression and, only in colonized mice, fut2 early expression. The adult microbiota is required for the fut2 induction responsible for the highly fucosylated adult gut phenotype and is necessary for recovery from intestinal injury. Fut2-dependent recovery from inflammation may explain the high incidence of inflammatory disease (Crohn's and necrotizing enterocolitis) in populations with mutant FUT2 polymorphic alleles.

Mucormycosis is a life-threatening fungal infection almost uniformly affecting diabetics in ketoacidosis or other forms of acidosis and/or immunocompromised patients. Inhalation of Mucorales spores provides the most common natural route of entry into the host. In this study, we developed an intratracheal instillation model of pulmonary mucormycosis that hematogenously disseminates into other organs using diabetic ketoacidotic (DKA) or cyclophosphamide-cortisone acetate-treated mice. Various degrees of lethality were achieved for the DKA or cyclophosphamide-cortisone acetate-treated mice when infected with different clinical isolates of Mucorales. In both DKA and cyclophosphamide-cortisone acetate models, liposomal amphotericin B (LAmB) or posaconazole (POS) treatments were effective in improving survival, reducing lungs and brain fungal burdens, and histologically resolving the infection compared with placebo. These models can be used to study mechanisms of infection, develop immunotherapeutic strategies, and evaluate drug efficacies against life-threatening Mucorales infections.

Background/aims: Glucocorticoid (GC) therapy is known to predispose to an adverse metabolic profile. Therefore, we investigated the prevalence of obesity and metabolic syndrome (MetS) in young patients with congenital adrenal hyperplasia (CAH) and to correlate this prevalence with GC treatment and family history., Methods: The study population consisted of 33 young CAH patients who received cortisone acetate during their growth periods; those who were salt wasters also received fludrocortisone. Obesity was defined by a body mass index (BMI) >95th percentile and MetS by the National Cholesterol Education Program Third Adult Treatment Panel modified criteria. Each patient's familial history of MetS components was assessed. The impact of GC therapy on the metabolic profile was analyzed by comparing CAH patients with BMI z-score-matched controls., Results: MetS and obesity were observed in 12.1 and 30.3% of the CAH patients, respectively, both of which were higher than in the reference population. A positive family history of MetS was found to be more prevalent in the obese patients compared with the nonobese CAH patients, and similar findings were observed for the controls. The metabolic profile did not differ between the CAH patients and matched subjects., Conclusion: CAH patients presented a higher prevalence of obesity and MetS, which were not correlated with the GC treatment. This study suggests that obesity and familial predisposition are significant determining factors for an adverse metabolic profile in CAH patients., (Copyright © 2013 S. Karger AG, Basel.)

Objective: The protocols for glucocorticoid replacement in children with salt wasting 21-hydroxylase deficiency are well established; however, the current recommendation for mineralocorticoid replacement is general and suggests individualized dose adjustments. This study aims to retrospectively review the 9-α-fludrocortisone dose regimen in salt wasting 21-hydroxylase deficient children who have been adequately treated during infancy., Methods: Twenty-three salt wasting 21-hydroxylase deficient patients with good anthropometric and hormonal control were followed in our center since diagnosis. The assessments of cortisone acetate and 9-α-fludrocortisone doses, anthropometric parameters, and biochemical and hormonal levels were rigorously evaluated in pre-determined intervals from diagnosis to two years of age., Results: The 9-α-fludrocortisone doses decreased over time during the first and second years of life; the median fludrocortisone doses were 200 µg at 0-6 months, 150 µg at 7-18 months and 125 µg at 19-24 months. The cortisone acetate dose per square meter was stable during follow-up (median = 16.8 mg/m²/day). The serum sodium, potassium and plasma rennin activity levels during treatment were normal, except in the first month of life, when periodic 9-α-fludrocortisone dose adjustments were made., Conclusions: The mineralocorticoid needs of salt wasting 21-hydroxylase deficient patients are greater during early infancy and progressively decrease during the first two years of life, which confirms that a partial aldosterone resistance exists during this time. Our study proposes a safety regiment for mineralocorticoid replacement during this critical developmental period.

Objective: Patients with Addison's disease require lifelong treatment with glucocorticoids. At present, no glucocorticoid replacement therapy (GRT) can exactly mimic normal physiology. As a consequence, under- and especially overtreatment can occur. Suboptimal GRT may lead to various side effects. The aim of this study was to investigate the use of salivary cortisol day curves (SCDC) in the individual adjustment of GRT in order to approach normal cortisol levels as closely as possible, reduce over- and underreplacement and study the short-term effects on quality of life (QoL)., Design and Methods: Twenty patients with Addison's disease were included in this prospective study. A SCDC was obtained and compared to normal controls; general and disease specific QoL-questionnaires were completed. Based on SCDC assessment of over- and undertreatment (calculated as duration (h) × magnitude (nmol/L) at different time points, glucocorticoid dose and regime were adjusted. After 4 weeks SCDC and QoL assessment were repeated and the effect of adjusting GRT was analysed., Results: At baseline, underreplacement was present in 3 and overreplacement in 18 patients; total calculated overreplacement was 32.8 h.nmol/L. Overreplacement decreased significantly to 13.3 h. nmol/L (p =0.005) after adjustment of GRT. Overreplacement was found particularly in the afternoon and evening. After reducing overreplacement in the evening, complaints about sleep disturbances significantly decreased., Conclusions: Individual adjustment of GRT based on SCDC to approach normal cortisol concentrations during the day can reduce overreplacement, especially in the evening. This can lead to a reduction of sleep disturbances and fatigue in patients with Addison's disease. A SCDC is a simple and patient-friendly tool for adjusting GRT and can be useful in the follow-up of patients with Addison's disease.

Raoul Dufy (1877-1953) and Niki de Saint Phalle (1930-2002) were 2 famous artists who suffered from rheumatoid arthritis (RA). Both artists represent an additional outstanding example of successful coping with RA in former times when, for the first time, corticosteroids were available, but nevertheless treatment was very limited in the pre-biological era. Dufy was one of the earliest patients with RA who received corticosteroids and regained his creativity to paint for a few additional years, but finally he died of massive intestinal hemorrhages, the adverse event of the combination of corticosteroid plus aspirin. Niki de Saint Phalle, a self-taught French painter and sculptor, was one of the most significant and unconventional female artists of the 20th century. Her eventful life was full of emotional burdens and lifelong lung disease in addition to RA. Niki de Saint Phalle came out from each physical and emotional crisis with new forces and new artistic ideas. Interestingly, it has been suggested that the occupational exposure to colors contributed to the development of RA in artists, which used significantly more bright and clear colors based on toxic heavy metals such as Renoir and Dufy. Moreover, these 2 were cigarette smokers, a recently described risk factor for developing RA and increasing the severity once it does develop. Niki de Saint Phalle produced her sculptures made of plastic material without protection while she assumed that exposition to polyester and toxic fumes of polystyrene caused severe damage to her lungs, resulting in recurrent health problems.

Glucocorticoids are listed on the World Anti-Doping Agency (WADA) Prohibited List of substances. The detection of the administration of hydrocortisone and cortisone is complicated by the fact that the human body also produces these steroids naturally. Gas chromatography-combustion-isotope ratio mass spectrometry can be utilized to determine the use of endogenous glucocorticoids by measuring the carbon isotope ratio (CIR) of their resulting metabolites in human urine samples. A comprehensive sample preparation protocol for the analysis of endogenous glucocorticoid urinary metabolites was developed and validated, incorporating the use of high performance liquid chromatography (HPLC) for purification and chemical oxidation for derivatisation. Target compounds were tetrahydrocortisol and tetrahydrocortisone, and 11β-hydroxyetiocholanolone, 11-oxoetiocholanolone and 11β-hydroxyandrosterone, while pregnanediol functioned as the endogenous reference compound. Urine samples from a population of 50 volunteers were analyzed to determine CIR reference limits. Excretion studies of the endogenous glucocorticoid preparation cortisone acetate (25 mg oral) and the dietary supplement adrenosterone (75 mg oral) were conducted with six male individuals. Variable changes in steroid metabolite isotopic composition were found across subjects after administration. The study also revealed that CIR analysis of the major glucocorticoid metabolites tetrahydrocortisol and tetrahydrocortisone is necessary to unambiguously distinguish administration of cortisone and adrenosterone, the former officially restricted to out-of-competition use by athletes, the latter not being restricted at the current time. Moreover, this study reaffirms that CIR methods for the doping control of endogenous steroids should not rely upon a single ERC, as the administration of an appropriate precursor to that ERC could cause complications during analysis., (Copyright © 2012 John Wiley & Sons, Ltd.)

In liver cirrhosis, hepatoadrenal syndrome has been described recently as a progressive impairment in the adrenocortical reserve, with deficient production or action of glucocorticoids resulting in adrenal insufficiency. Data on the treatment of this syndrome are scarce. We report a case of a 60-year-old male patient referred to our hospital because of rectal bleeding and bilateral leg swelling. He complained of marked weakness, bilateral leg swelling, and shortness of breath with exertion for the last 2 months. Biochemistry and imaging indicated liver cirrhosis. Because of the weakness and persistent hypotension, we performed a low-dose synacthen test, which indicated adrenal insufficiency (baseline cortisol level 1.8 μg/dl, increasing to 3.5 and 3.7 μg/dl at 20 and 30 min, respectively). The patient's general condition improved promptly after corticosteroid supplementation.

The Kv1 family voltage-dependent K(+) channels are essential for termination of action potentials in neurons and myocytes. These channels form a stable complex with their beta subunits (Kvβ), some of which inhibit channel activity. Cortisone potentiates Kv1 channel by binding to Kvβ and promoting its dissociation from the channel, but its half-maximum effective concentration is ∼46 μM. To identify corticosteroids that are more efficient than cortisone, we examined 25 cortisone analogues and found that fluticasone propionate potentiates channel current with a half-maximum effective concentration (EC(50)) of 37 ± 1.1 nM. Further studies showed that fluticasone propionate potentiates channel current by inducing dissociation of Kvβ, and docking of fluticasone propionate into the cortisone binding site reveals potential interactions that enhance the EC(50) value. Thus, fluticasone propionate provides a starting point for rational design of more efficient small-molecule compounds that increase Kv1 activity and affect the integrity of the Kv1-Kvβ complex.

Glucocorticoids (GC) are implicated in the development of metabolic syndrome, and patients with GC excess share many clinical features, such as central obesity and glucose intolerance. In patients with obesity or type 2 diabetes, systemic GC concentrations seem to be invariably normal. Tissue GC concentrations determined by the hypothalamic-pituitary-adrenal (HPA) axis and local cortisol (corticosterone in mice) regeneration from cortisone (11-dehydrocorticosterone in mice) by the 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) enzyme, principally expressed in the liver. Transgenic mice have demonstrated the importance of 11β-HSD1 in mediating aspects of the metabolic syndrome, as well as HPA axis control. In order to address the primacy of hepatic 11β-HSD1 in regulating metabolism and the HPA axis, we have generated liver-specific 11β-HSD1 knockout (LKO) mice, assessed biomarkers of GC metabolism, and examined responses to high-fat feeding. LKO mice were able to regenerate cortisol from cortisone to 40% of control and had no discernible difference in a urinary metabolite marker of 11β-HSD1 activity. Although circulating corticosterone was unaltered, adrenal size was increased, indicative of chronic HPA stimulation. There was a mild improvement in glucose tolerance but with insulin sensitivity largely unaffected. Adiposity and body weight were unaffected as were aspects of hepatic lipid homeostasis, triglyceride accumulation, and serum lipids. Additionally, no changes in the expression of genes involved in glucose or lipid homeostasis were observed. Liver-specific deletion of 11β-HSD1 reduces corticosterone regeneration and may be important for setting aspects of HPA axis tone, without impacting upon urinary steroid metabolite profile. These discordant data have significant implications for the use of these biomarkers of 11β-HSD1 activity in clinical studies. The paucity of metabolic abnormalities in LKO points to important compensatory effects by HPA activation and to a crucial role of extrahepatic 11β-HSD1 expression, highlighting the contribution of cross talk between GC target tissues in determining metabolic phenotype.

Aspergillus terreus is emerging as a causative agent of life-threatening invasive aspergillosis. Prognosis for affected patients is often worse than for A. fumigatus infections. To study A. terreus-mediated disease, we developed 3 infection models. In embryonated hen's eggs and leucopenic mice, the outcome of invasive aspergillosis was similar to that described for A. fumigatus. However, 10(2)- and 10(3)-fold higher conidia concentrations were required for 100% lethality. In corticosteroid-treated mice, only 50% mortality was observed, although bioluminescence imaging revealed transient disease in all infected animals. In surviving animals, we observed persistence of ungerminated but viable conidia. Cytokine levels in these mice were comparable to uninfected controls. In contrast to A. fumigatus infections, all mice infected with A. terreus developed fatty liver degeneration, suggesting the production of toxic secondary metabolites. Thus, at least in mice, persistence and subclinical liver damage are unique features of A. terreus infections.

Objectives: Amphotericin B inhalation powder (ABIP) is a novel dry-powder amphotericin B formulation that is directly delivered to the lung, resulting in elevated lung tissue drug concentrations of this polyene. We evaluated the prophylactic efficacy of single dose administration of ABIP in a guinea pig model of invasive pulmonary aspergillosis., Methods: Guinea pigs were immunosuppressed with cyclophosphamide and cortisone acetate and challenged with Aspergillus fumigatus conidia in an aerosol chamber. Guinea pigs received prophylaxis with a single inhaled dose of ABIP at 0.05, 0.5, 4 or 10 mg/kg administered 24 h prior to infection. Treatment with oral voriconazole at doses of 5 or 10 mg/kg twice daily beginning 24 h post-challenge served as the positive control., Results: Improvements in survival were observed with ABIP prophylaxis. A single inhaled dose of 4 mg/kg ABIP and treatment with 5 mg/kg voriconazole both improved median and percentage survival compared with untreated controls. In addition, pulmonary fungal burden, as assessed by cfu, quantitative PCR and galactomannan, was also reduced in a dose-dependent fashion with ABIP prophylaxis as well as with both doses of voriconazole treatment., Conclusions: Single-dose prophylaxis with inhaled ABIP as prophylaxis demonstrated a significant survival advantage and reductions in pulmonary fungal burden in this model of invasive pulmonary aspergillosis. Optimization of the dose and dosing frequency of ABIP dose may help to further enhance the anti-Aspergillus activity of this novel amphotericin B formulation.

Oropharyngeal candidiasis is a frequent cause of morbidity in patients with defects in cell-mediated immunity or saliva production. Animal models of this infection are important for studying disease pathogenesis and evaluating vaccines and antifungal therapies. Here we describe a simple mouse model of oropharyngeal candidiasis. Mice are rendered susceptible to oral infection by injection with cortisone acetate and then inoculated by placing a swab saturated with Candida albicans sublingually. This process results in a reproducible level of infection, the histopathology of which mimics that of pseudomembranous oropharyngeal candidiasis in humans. By using this model, data are obtained after 5-9 d of work.

Objective: Failure to mirror the diurnal cortisol profile could contribute to the impaired subjective health status in Addison's disease (AD). Some patients report benefit from the use of various nutritional compounds. The objective of this study was to investigate the impact of licorice and grapefruit juice (GFJ) on the absorption and metabolism of cortisone acetate (CA)., Design: Patients (n=17) with AD on stable CA replacement therapy were recruited from the outpatient clinic at Haukeland University Hospital, Norway. They were assessed on their ordinary CA medication and following two 3-day periods of co-administration of licorice or GFJ., Methods: Time series of glucocorticoids (GCs) in serum and saliva were obtained, and GCs in 24 h urine samples were determined. The main outcome measure was the area under the curve (AUC) for serum cortisol in the first 2.6 h after orally administered CA., Results: Compared with the ordinary treatment, the median AUC for serum cortisol increased with licorice (53 783 vs 50 882, P<0.05) and GFJ (60 661 vs 50 882, P<0.05). Median cortisol levels in serum were also elevated 2.6 h after tablet ingestion (licorice 223 vs 186 nmol/l, P<0.05; GFJ 337 vs 186 nmol/l, P<0.01). Licorice increased the median urinary cortisol/cortisone ratio (0.43 vs 0.21, P<0.00001), whereas GFJ increased the (allo-tetrahydrocortisol+tetrahydrocortisol)/tetrahydrocortisone ratio (0.55 vs 0.43, P<0.05)., Conclusion: Licorice and in particular GFJ increased cortisol available to tissues in the hours following oral CA administration. Both patients and physicians should be aware of these interactions.

Menopause is associated with central obesity, dyslipidemia, hypertension and insulin resistance, which are also shown in the patients with excess of glucocorticoids. However, the interaction of hypothalamic-pituitary-adrenal (HPA) axis activity and menopause has not been fully understood. In this study, 55 healthy non-obese women were recruited, and then divided into two groups, premenopausal group (n = 24) and postmenopausal group (n = 31). HPA axis function was evaluated by using dexamethasone suppression test (DST; 0.25 mg), and adrenocorticotrophic hormone (ACTH)-stimulation test. Moreover, 25 mg cortisone acetate test was applied to evaluate the hepatic 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) activity. We found that hepatic 11β-HSD1 activity and adrenal response to ACTH were increased in the postmenopausal group compared with the premenopausal group, whereas the negative feedback effect of dexamethasone did not show significant difference between pre- and postmenopausal women. These findings suggest that the adrenal sensitivity to ACTH and hepatic 11β-HSD1 activity are increased. The increased cortisol conversion and/or synthesis may be contributed to the dysmetabolic features in the postmenopausal women.

An endogenous probe for CYP3A activity would be useful for early identification of in vivo cytochrome P450 (CYP) 3A4 inhibitors. The aim of this study was to determine whether formation clearance (CL(f)) of the sum of 6β-hydroxycortisol and 6β-hydroxycortisone is a useful probe of CYP3A4 inhibition in vivo. In human liver microsomes (HLMs), the formation of 6β-hydroxycortisol and 6β-hydroxycortisone was catalyzed by CYP3A4, and itraconazole inhibited these reactions with half maximal inhibitory concentration (IC(50))(,u) values of 3.1 nmol/l and 3.4 nmol/l, respectively. The in vivo IC(50,u) value of itraconazole for the combined CL(f) of 6β-hydroxycortisone and 6β-hydroxycortisol was 1.6 nmol/l. The greater inhibitory potency in vivo is probably due to circulating inhibitory itraconazole metabolites. The maximum in vivo inhibition was 59%, suggesting that f(m,CYP3A4) for cortisol and cortisone 6β-hydroxylation is ~60%. Given the significant decrease in CL(f) of 6β-hydroxycortisone and 6β-hydroxycortisol after 200-mg and 400-mg single doses of itraconazole, this endogenous probe can be used to detect moderate and potent CYP3A4 inhibition in vivo.

Cortisone acetate test was performed in twelve young adult patients with diabetes mellitus type 1, after dexamethasone administration to suppress endogenous cortisol production. Previous screening revealed that all of the subjects had peak cortisol responses in the range from subnormal to normal, as determined by a low-dose Synacthen test. The aim was to find out whether these patients would exhibit different conversion of cortisone to cortisol by 11beta-hydroxysteroid dehydrogenase. Using multifactorial ANOVA the following significant relationships were obtained between cortisol or cortisol/cortisone ratio measured during the test and other parameters examined a) before dexamethasone suppression and b) during the test: a) Cortisol at 120(th) minute negatively correlated with daily insulin dose and positively with basal aldosterone. Cortisol/cortisone ratio at 60(th), 120(th), 180(th), and 240(th) minute negatively correlated with basal aldosterone/plasma renin activity ratio, urinary free cortisol/24 hours and positively with basal dehydroepindrosterone sulphate. b) Cortisol at 120(th) minute negatively correlated with suppressed basal serum glycemia; cortisol/cortisone ratio during the whole test negatively correlated with supressed basal ACTH. The examination of peripheral metabolism of cortisol using cortisone acetate test in patients with diabetes mellitus type 1 showed adaptive changes of 11beta-hydroxysteroid dehydrogenace activity associated with altered cortisol tissue supply.

Transformation of 16 delta5-3beta-hydroxy- and delta4-3-ketosteroids of androstane and pregnane classes was carried out using Curvularia lunata mycelium suspended in phosphate buffer with methyl-beta-cyclodextrine (MCD). As the result, 20 monohydroxy- and dihydroxy-metabolites, whose structure was determined using specters of proton magnetic resonance and mass-specters, have been isolated. Hydroxylation of delta5-3beta-hydroxy-steroids occurred mostly in the C-7alpha position whereas hydroxylation of delta4-3-ketosteroids was in the C-11beta position. Only androst-4-en-3,17-dione, 9alpha-hydroxyl-androstenedione, and androsts-1,4-diene-3,17-dione were hydroxylated at C-14alpha position. Besides main 11beta-derivatives, the 6beta- and 7beta-hydroxy-derivatives with yield 10 and 30%, respectively, were isolated during transformation of progesterone and hydroxymethyl pregnadienon. The ratio of MCD to transforming steroid was 1 : 1 (mol/mol). Hydroxycortisone and 7alpha-hydroxyandrostenolone with the yield 55 and 77%, respectively, were obtained at the maximal concentrations of cortexolone 20 g/l and androstenolone acetate 10 g/l in the presence of MCD. Absorption of steroids on mycelium, lower speed of their transformation, low concentrations of modifying substrates, and low yield of hydroxyderivatives have been observed in the absence of MCD.

Introduction: 21-hydroxylase deficiency is an autosomal recessive disorder that causes glucocorticoid deficiency and increased androgen production. Treatment is based on glucocorticoid replacement; however, interindividual variability in the glucocorticoid dose required to achieve adequate hormonal control has been observed., Objective: The present study aimed to evaluate the association between polymorphic variants involved inglucocorticoid action and/or metabolism and the mean daily glucocorticoid dose in 21-hydroxylase deficiency patients., Methods: We evaluated 53 patients with classical forms of 21-hydroxylase deficiency who were receiving cortisone acetate. All patients were between four and six years of age and had normal androgen levels., Results: The P450 oxidoreductase A503V, HSD11B1 rs12086634, and CYP3A7*1C variants were found in 19%, 11.3% and 3.8% of the patients, respectively. The mean ± SD glucocorticoid dose in patients with the CYP3A7*1C and wild-type alleles was 13.9 ± 0.8 and 19.5 ± 3.2 mg/m²/d, respectively. We did not identify an association between the P450 oxidoreductase or HSD11B1 allelic variants and the mean glucocorticoid dose., Conclusion: Patients carrying the CYP3A7*1C variant required a significantly lower mean glucocorticoid dose. Indeed, the CYP3A7*1C allele accounted for 20% of the variability in the cortisone acetate dose. The analysis of genes involved in glucocorticoid metabolism may be useful in the optimization of treatment of 21-hydroxylase deficiency.

Our aim was to investigate the usefulness of circulating levels of adrenocorticotropic hormone (ACTH) and also salivary cortisol to monitor cortisone substitution in patients with Addison's disease. 13 patients with primary adrenal insufficiency (8 women and 5 men, age 44 ± 11 years) received 12.5 mg cortisone acetate orally at 16:00 h and 25 mg at 07:00 h. Blood samples for cortisol and ACTH analysis were drawn every hour for 24 h, and also every half hour between 07:00 and 12:00 h. Samples for salivary cortisol were collected in parallel. Total ACTH levels showed large inter-individual variations and a diurnal rhythm with a nadir in the early evening at 19:00 (median 19 ng/l, range 2-434 ng/l) and high levels in the early morning, with a peak around 07:30 (median 844 ng/l, range 45-2,249 ng/l). Plasma cortisol concentrations showed 2 peaks distinct in time, but variable in height, 1-2 h after intake of cortisone. Plasma cortisol correlated significantly with ln(ACTH) at 17:00 h (r=-0.56), at 10:00 h (r=-0.51), and at 10.30 h (r=-0.57). When tested at different time points, ln(ACTH) at 10:00 to 12:00 h was negatively correlated with plasma cortisol between 08:30 and 12:00 h. Plasma cortisol was highly correlated to ln(salivary cortisol) most of the time points measured, but 30-60 min after intake of cortisone acetate the correlation disappeared. In conclusion, the large interindividual variation in ACTH levels most likely indicates varying sensitivity to cortisol with a need for individualized dosing schemes. Furthermore ACTH-determinations may be useful for dose titration of cortisol., (© Georg Thieme Verlag KG Stuttgart · New York.)

Atlantic cod Gadus morhua ovaries were incubated in vitro with tritiated 17-hydroxypregn-4-ene-3,20-dione (17-P) to determine whether 17,20beta-dihydroxypregn-4-en-3-one (17,20beta-P) or 17,20beta, 21-trihydroxypregn-4-en-3-one (17,20beta,21-P), or both, were more likely to be the steroid responsible for inducing oocyte final maturation (i.e. resumption of meiosis). Only 17,20beta,21-P was produced, in addition to 11-deoxycortisol (17,21-P), which is intermediate between 17-P and 17,20beta,21-P. Also, the 5beta-reduced forms of 17-P, 17,21-P and 17,20beta,21-P were all found. Some sulphation of 21-hydroxylated steroids was demonstrated. The ability of female G. morhua to make 17,20beta,21-P but not 17,20beta-P was confirmed by radioimmunoassay of plasma samples from spawning fish. Although small amounts of 17,20beta-P immunoreactivity were detected in a few plasma samples, this was shown, by thin-layer chromatography, to be mostly due to cross-reaction with other unidentified compounds. The evidence strongly suggests that 17,20beta,21-P is more likely than 17,20beta-P to be the maturation-inducing steroid in G. morhua.

Background: Inflammation and oedema of the facial nerve are implicated in causing Bell's palsy. Corticosteroids have a potent anti-inflammatory action which should minimise nerve damage., Objectives: The objective of this review was to assess the effect of corticosteroid therapy in Bell's palsy., Search Strategy: We searched the Cochrane Neuromuscular Disease Group Trials Specialized Register (9 December 2008) for randomised trials, as well as MEDLINE (January 1966 to December 2008), EMBASE (January 1980 to December 2008) and LILACS (9 December 2008). We contacted known experts in the field to identify additional published or unpublished trials., Selection Criteria: Randomised trials comparing different routes of administration and dosage schemes of corticosteroid or adrenocorticotrophic hormone therapy versus a control group where no therapy considered effective for this condition was administered, unless it was also given in a similar way to the experimental group., Data Collection and Analysis: Two authors independently assessed eligibility, trial quality, and extracted the data., Main Results: Eight trials with a total of 1569 participants were included. Allocation concealment was appropriate in six trials, and the data reported allowed an intention-to-treat analysis in four, while unpublished data from the fifth and sixth trials were provided by the authors. The data included in the main outcome of this meta-analysis were collected from seven trials with a total of 1507 participants. Overall 175/754 (23%) of the participants allocated to corticosteroids had incomplete recovery of facial motor function six months or more after randomisation, significantly less than 245/753 (33%) in the control group (risk ratio (RR) 0.71, 95% confidence interval (CI) 0.61 to 0.83). There was, also, a significant reduction in motor synkinesis during follow-up in those receiving corticosteroids (RR 0.6, 95% CI 0.44 to 0.81). The reduction in the proportion of patients with cosmetically disabling sequelae six months after randomisation, however, was not significant (RR 0.97, 95% CI 0.44 to 2.15). The trial not included in the primary outcome of this meta-analysis showed a non-significant difference in outcomes between the arms., Authors' Conclusions: The available evidence from randomised controlled trials shows significant benefit from treating Bell's palsy with corticosteroids.

An intensive and systematic investigation had been carried out on the Delta(1)-dehydrogenation of cortisone acetate (CA) to prednisone acetate (PA) by Arthrobacter simplex TCCC 11037 in the presence of native and modified beta-cyclodextrins (beta-CDs). The biotransformation was improved through the formation of the host-guest inclusion complex between CA and CDs in aqueous solution. The inclusion complexes of CDs with CA were investigated by means of phase solubility, 2D NMR spectroscopy and differential scanning calorimetry (DSC). The structural difference of CDs resulted in the stoichiometric differences between the complexes, the RM-beta-CD-CA, SBE-beta-CD-CA, HP-beta-CD-CA complexes were 1:1 whereas beta-CD-CA gave both 1:1 and 2:1 complexes, of which the 2:1 complex decreased the soluble CA concentration and inhibited the dissociation of beta-CD-CA in aqueous solution. The increase in solubility of CA was in the order of RM-beta-CD>SBE-beta-CD>HP-beta-CD>beta-CD. RM-beta-CD-CA, SBE-beta-CD-CA and HP-beta-CD-CA exhibited the higher biotransformation rate in comparison with native beta-CD. And the solubilization of CDs for CA in aqueous medium plays a key role in the biotransformation process. The article focuses on the various factors influencing the substrate water solubility, complex stability and biotransformation of CA through the addition of CDs in order to solve many problems associated with the process of drug delivery and biotransformation of different novel steroids.

The effect of the varying operating parameters on the separation of steroids was studied by oil-in-water microemulsion liquid chromatography (MELC). The parameters included the surfactant concentration, the type of oil phase, the nature of organic solvent additives, and the pore size of the stationary-phase. The optimized conditions for the separation of 6 steroids were as follows: a Venusil ASB C18 (T) column (5 microm, 30 nm, 250 mm x 4.6 mm) was used at 35 degrees C, the microemulsion mobile phase consisted of 30 g/L sodium dodecylsulfate (SDS), 0.8% (w/w) n-octane, 6.6% (w/w) n-butanol. The optimized method can be used for the separation, identification and simultaneous determination of steroids in bulk drugs and in pharmaceutical dose forms.

Background: Inflammation and oedema of the facial nerve are implicated in causing Bell's palsy. Corticosteroids have a potent anti-inflammatory action which should minimise nerve damage and thereby improve the outcome of patients suffering from this condition., Objectives: The objective of this review was to assess the effect of steroid therapy in the recovery of patients with Bell's palsy., Search Strategy: We searched the Cochrane Neuromuscular Disease Group register (searched November 2005) for randomised trials, as well as MEDLINE (January 1966 to November 2005), EMBASE (January 1980 to November 2005) and LILACS (January 1982 to November 2005). We contacted known experts in the field to identify additional published or unpublished trials., Selection Criteria: Randomised trials comparing different routes of administration and dosage schemes of corticosteroid or adrenocorticotrophic hormone therapy versus a control group where no therapy considered effective for this condition was administered, unless it was also given in a similar way to the experimental group., Data Collection and Analysis: Two reviewers independently assessed eligibility, trial quality, and extracted the data., Main Results: Four trials with a total of 179 patients were included. One trial compared cortisone acetate with placebo; one compared prednisone plus vitamins, with vitamins alone; one compared high-dose prednisone administered intravenously against saline solution, and one, not-placebo controlled, tested the efficacy of methylprednisolone. Allocation concealment was appropriate in two trials, and the data reported allowed an intention-to-treat analysis. The data included in the meta-analyses were collected from three trials with a total of 117 patients. Overall 13/59 (22%) of the patients allocated to steroid therapy had incomplete recovery of facial motor function six months after randomisation, compared with 15/58 (26%) in the control group. This reduction was not significant (relative risk 0.86, 95% confidence interval 0.47 to 1.59). The reduction in the proportion of patients with cosmetically disabling sequelae six months after randomisation was also not significant (relative risk 0.86, 95% confidence interval 0.38 to 1.98). The trial not included in the meta-analysis showed a non-significant difference in outcomes between the arms., Authors' Conclusions: The available evidence from randomised controlled trials does not show significant benefit from treating Bell's palsy with corticosteroids. More randomised controlled trials with a greater number of patients are needed to determine reliably whether there is real benefit (or harm) from the use of corticosteroid therapy in patients with Bell's palsy. One trial, with 551 participants, comparing prednisolone with acyclovir with both and with neither has just been published and will be included in an update of this review.

An HPLC-DAD method for determining corticoids in calf feed and in animal feeding water samples using a monolithic column has been developed and validated. The method optimization included the study of binary mobile phases of water and acetonitrile. The optimum separation was achieved at 40 degrees C, with acetonitrile:H(2)O 29:71 v/v used as mobile phase and a 3 ml/min flow-rate, which resulted in their separation in about 5 min. Two reported sample procedures were applied to feed and for animal feeding water samples prior to HPLC. Method validation was carried out according to the EU criteria established for quantitative screening methods. The results indicate that this method is highly specific, reproducible and accurate. The proposed method was found to be robust and unaffected by small variations in the extraction procedure and in HPLC conditions. The developed method for the determination of corticoids in feed and water samples was also found to be suitable for different kinds of feeds and waters.