Inhibition of the glucocorticoid-activating enzyme 11ß-hydroxysteroid dehydrogenase type 1 drives concurrent 11-oxygenated androgen excess.

Aldo-keto reductase 1C3 (AKR1C3) is a key enzyme in the activation of both classic and 11-oxygenated androgens. In adipose tissue, AKR1C3 is co-expressed with 11ß-hydroxysteroid dehydrogenase type 1 (HSD11B1), which catalyzes not only the local activation of glucocorticoids but also the inactivation of 11-oxygenated androgens, and thus has the potential to counteract AKR1C3. Using a combination of in vitro assays and in silico modeling we show that HSD11B1 attenuates the biosynthesis of the potent 11-oxygenated androgen, 11-ketotestosterone (11KT), by AKR1C3. Employing ex vivo incubations of human female adipose tissue samples we show that inhibition of HSD11B1 results in the increased peripheral biosynthesis of 11KT. Moreover, circulating 11KT increased 2-3 fold in individuals with type 2 diabetes after receiving the selective oral HSD11B1 inhibitor AZD4017 for 35 days, thus confirming that HSD11B1 inhibition results in systemic increases in 11KT concentrations. Our findings show that HSD11B1 protects against excess 11KT production by adipose tissue, a finding of particular significance when considering the evidence for adverse metabolic effects of androgens in women. Therefore, when targeting glucocorticoid activation by HSD11B1 inhibitor treatment in women, the consequently increased generation of 11KT may offset beneficial effects of decreased glucocorticoid activation.

Reduced plakoglobin increases the risk of sodium current defects and atrial conduction abnormalities in response to androgenic anabolic steroid abuse.

Androgenic anabolic steroids (AAS) are commonly abused by young men. Male sex and increased AAS levels are associated with earlier and more severe manifestation of common cardiac conditions, such as atrial fibrillation, and rare ones, such as arrhythmogenic right ventricular cardiomyopathy (ARVC). Clinical observations suggest a potential atrial involvement in ARVC. Arrhythmogenic right ventricular cardiomyopathy is caused by desmosomal gene defects, including reduced plakoglobin expression. Here, we analysed clinical records from 146 ARVC patients to identify that ARVC is more common in males than females. Patients with ARVC also had an increased incidence of atrial arrhythmias and P wave changes. To study desmosomal vulnerability and the effects of AAS on the atria, young adult male mice, heterozygously deficient for plakoglobin (Plako+/- ), and wild type (WT) littermates were chronically exposed to 5α-dihydrotestosterone (DHT) or placebo. The DHT increased atrial expression of pro-hypertrophic, fibrotic and inflammatory transcripts. In mice with reduced plakoglobin, DHT exaggerated P wave abnormalities, atrial conduction slowing, sodium current depletion, action potential amplitude reduction and the fall in action potential depolarization rate. Super-resolution microscopy revealed a decrease in NaV 1.5 membrane clustering in Plako+/- atrial cardiomyocytes after DHT exposure. In summary, AAS combined with plakoglobin deficiency cause pathological atrial electrical remodelling in young male hearts. Male sex is likely to increase the risk of atrial arrhythmia, particularly in those with desmosomal gene variants. This risk is likely to be exaggerated further by AAS use. KEY POINTS: Androgenic male sex hormones, such as testosterone, might increase the risk of atrial fibrillation in patients with arrhythmogenic right ventricular cardiomyopathy (ARVC), which is often caused by desmosomal gene defects (e.g. reduced plakoglobin expression). In this study, we observed a significantly higher proportion of males who had ARVC compared with females, and atrial arrhythmias and P wave changes represented a common observation in advanced ARVC stages. In mice with reduced plakoglobin expression, chronic administration of 5α-dihydrotestosterone led to P wave abnormalities, atrial conduction slowing, sodium current depletion and a decrease in membrane-localized NaV 1.5 clusters. 5α-Dihydrotestosterone, therefore, represents a stimulus aggravating the pro-arrhythmic phenotype in carriers of desmosomal mutations and can affect atrial electrical function.

COVID-19-related adrenal haemorrhage: Multicentre UK experience and systematic review of the literature.

OBJECTIVE: Adrenal haemorrhage (AH) is an uncommon, usually incidental imaging finding in acutely unwell patients. AH has been reported during coronavirus disease 2019 (COVID-19) infection and following ChAdOx1 nCoV-19 (Oxford-AstraZeneca) vaccination. The Society for Endocrinology (SfE) established a task force to describe the UK experience of COVID-19-related AH. DESIGN: A systematic literature review was undertaken. A survey was conducted through the SfE clinical membership to identify patients with COVID-19-related AH using a standardized data collection tool. RESULTS: The literature search yielded 25 cases of COVID-19-related AH (19 bilateral; 13 infection-related, and 12 vaccine-related). Eight UK centres responded to the survey with at least one case. A total of 18 cases were included in the descriptive study, including 11 from the survey and 7 UK-based patients from the systematic review. Seven patients (4 males; median age 53 (range 26-70) years), had infection-related AH (four bilateral). Median time from positive COVID-19 test to AH detection was 8 (range 1-30) days. Eleven cases of vaccine-related AH (eight bilateral) were captured (3 males; median age 47 (range 23-78) years). Median time between vaccination (nine Oxford-AstraZeneca and two Pfizer-BioNTech) and AH was 9 (range 2-27) days; 9/11 AH occurred after the first vaccine dose. Acute abdominal pain was the commonest presentation (72%) in AH of any cause. All 12 patients with bilateral AH and one patient with unilateral AH required glucocorticoid replacement. CONCLUSION: Adrenal haemorrhage with consequential adrenal insufficiency can be a complication of COVID-19 infection and vaccination. Adrenal function assessment is mandatory to avoid the potentially fatal consequences of unrecognized adrenal insufficiency.

Emotional and psychosexual well-being is influenced by ethnicity and birthplace in women and individuals with polycystic ovary syndrome in the UK and India.

OBJECTIVE: To assess the association of ethnicity and birthplace on emotional and psychosexual well-being in women with polycystic ovary syndrome (PCOS). DESIGN: Cross-sectional study. SETTING: Community recruitment via social media campaigns. POPULATION: Women with PCOS completing an online questionnaire in September-October 2020 (UK) and May-June 2021 (India). METHODS: The survey has five components, with a baseline information and sociodemographic section followed by four validated questionnaires: Hospital Anxiety and Depression Scale (HADS); Body Image Concern Inventory (BICI); Beliefs About Obese Persons Scale (BAOP); and Female Sexual Function Index (FSFI). MAIN OUTCOME MEASURES: We used adjusted linear and logistic regression models, adjusting for age, education, marital status and parity, to evaluate the impact of ethnicity and birthplace on questionnaire scores and outcomes (anxiety and/or depression, HADS ≥ 11; body dysmorphic disorder (BDD), BICI ≥ 72). RESULTS: A total of 1008 women with PCOS were included. Women of non-white ethnicity (613/1008) reported higher rates of depression (OR 1.96, 95% CI 1.41-2.73) and lower BDD (OR 0.57, 95% CI 0.41-0.79) than white women (395/1008). Women born in India (453/1008) had higher anxiety (OR 1.57, 95% CI 1.00-2.46) and depression (OR 2.20, 95% CI 1.52-3.18) but lower BDD rates (OR 0.42, 95% CI 0.29-0.61) than women born in the UK (437/1008). All sexual domains, excluding desire, scored lower for non-white women and women born in India. CONCLUSIONS: Non-white women and women born in India reported higher emotional and sexual dysfunction, whereas white women and women born in the UK reported higher body image concerns and weight stigma. Ethnicity and birthplace need to be considered for tailored, multidisciplinary care.

Cardiometabolic Disease Burden and Steroid Excretion in Benign Adrenal Tumors : A Cross-Sectional Multicenter Study.

BACKGROUND: Benign adrenal tumors are commonly discovered on cross-sectional imaging. Mild autonomous cortisol secretion (MACS) is regularly diagnosed, but its effect on cardiometabolic disease in affected persons is ill defined. OBJECTIVE: To determine cardiometabolic disease burden and steroid excretion in persons with benign adrenal tumors with and without MACS. DESIGN: Cross-sectional study. SETTING: 14 endocrine secondary and tertiary care centers (recruitment from 2011 to 2016). PARTICIPANTS: 1305 prospectively recruited persons with benign adrenal tumors. MEASUREMENTS: Cortisol excess was defined by clinical assessment and the 1-mg overnight dexamethasone-suppression test (serum cortisol: <50 nmol/L, nonfunctioning adrenal tumor [NFAT]; 50 to 138 nmol/L, possible MACS [MACS-1]; >138 nmol/L and absence of typical clinical Cushing syndrome [CS] features, definitive MACS [MACS-2]). Net steroid production was assessed by multisteroid profiling of 24-hour urine by tandem mass spectrometry. RESULTS: Of the 1305 participants, 49.7% had NFAT (n = 649; 64.1% women), 34.6% had MACS-1 (n = 451; 67.2% women), 10.7% had MACS-2 (n = 140; 73.6% women), and 5.0% had CS (n = 65; 86.2% women). Prevalence and severity of hypertension were higher in MACS-2 and CS than NFAT (adjusted prevalence ratios [aPRs] for hypertension: MACS-2, 1.15 [95% CI, 1.04 to 1.27], and CS, 1.37 [CI, 1.16 to 1.62]; aPRs for use of ≥3 antihypertensives: MACS-2, 1.31 [CI, 1.02 to 1.68], and CS, 2.22 [CI, 1.62 to 3.05]). Type 2 diabetes was more prevalent in CS than NFAT (aPR, 1.62 [CI, 1.08 to 2.42]) and more likely to require insulin therapy for MACS-2 (aPR, 1.89 [CI, 1.01 to 3.52]) and CS (aPR, 3.06 [CI, 1.60 to 5.85]). Urinary multisteroid profiling revealed an increase in glucocorticoid excretion from NFAT over MACS-1 and MACS-2 to CS, whereas androgen excretion decreased. LIMITATIONS: Cross-sectional design; possible selection bias. CONCLUSION: A cardiometabolic risk condition, MACS predominantly affects women and warrants regular assessment for hypertension and type 2 diabetes. PRIMARY FUNDING SOURCE: Diabetes UK, the European Commission, U.K. Medical Research Council, the U.K. Academy of Medical Sciences, the Wellcome Trust, the U.K. National Institute for Health Research, the U.S. National Institutes of Health, the Claire Khan Trust Fund at University Hospitals Birmingham Charities, and the Mayo Clinic Foundation for Medical Education and Research.

Abiraterone switches castration-resistant prostate cancer dependency from adrenal androgens towards androgen receptor variants and glucocorticoid receptor signalling.

INTRODUCTION: Castration-resistant prostate cancer (CRPC) remains dependent on androgen receptor (AR) signalling, which is largely driven by conversion of adrenal androgen precursors lasting after castration. Abiraterone, an inhibitor of the steroidogenic enzyme CYP17A1, has been demonstrated to reduce adrenal androgen synthesis and prolong CRPC patient survival. To study mechanisms of resistance to castration and abiraterone, we created coculture models using human prostate and adrenal tumours. MATERIALS AND METHODS: Castration-naïve and CRPC clones of VCaP were incubated with steroid substrates or cocultured with human adrenal cells (H295R) and treated with abiraterone or the antiandrogen enzalutamide. Male mice bearing VCaP xenografts with and without concurrent H295R xenografts were castrated and treated with placebo or abiraterone. Response was assessed by tumour growth and PSA release. Plasma and tumour steroid levels were assessed by LC/MS-MS. Quantitative polymerase chain reaction determined steroidogenic enzyme, nuclear receptor and AR target gene expression. RESULTS: In vitro, adrenal androgens induced castration-naïve and CRPC cell growth, while precursors steroids for de novo synthesis did not. In a coculture system, abiraterone blocked H295R-induced growth of VCaP cells. In vivo, H295R promoted castration-resistant VCaP growth. Abiraterone only inhibited VCaP growth or PSA production in the presence of H295R. Plasma steroid levels demonstrated CYP17A1 inhibition by abiraterone, whilst CRPC tumour tissue steroid levels showed no evidence of de novo intratumoural androgen production. Castration-resistant and abiraterone-resistant VCaP tumours had increased levels of AR, AR variants and glucocorticoid receptor (GR) resulting in equal AR target gene expression levels compared to noncastrate tumours. CONCLUSIONS: In our model, ligand-dependent AR-regulated regrowth of CRPC was predominantly supported via adrenal androgen precursor production while there was no evidence for intratumoural androgen synthesis. Abiraterone-resistant tumours relied on AR overexpression, expression of ligand-independent AR variants and GR signalling.

Polycystic ovary syndrome and risk of adverse obstetric outcomes: a retrospective population-based matched cohort study in England.

BACKGROUND: Polycystic ovary syndrome (PCOS) affects up to one in five women of childbearing age. Observational studies assessing the association between maternal PCOS and adverse obstetric outcomes have reported varying results, depending on patient population, diagnostic criteria for PCOS and covariates accounted for in their analyses. We aimed to assess the risk of obstetric outcomes among a population-based representative cohort of women with PCOS compared to an age-matched cohort of women without PCOS. METHODS: A retrospective cohort study was conducted of pregnancies of women in England aged 15-49 years identified from the Clinical Practice Research Datalink (CPRD) GOLD pregnancy register and linked Hospital Episodes Statistic (HES) data between March 1997 and March 2020. Pregnancies from the register that had a linked HES delivery record were included. Linked CPRD primary care data was used to ascertain maternal PCOS exposure prior to pregnancy. To improve detection of PCOS, in addition to PCOS diagnostic codes, codes for (1) polycystic ovaries or (2) hyperandrogenism and anovulation together were also considered. Sensitivity analysis was limited to only pregnant women with a diagnostic code for PCOS. Primary outcomes ascertained from linked HES data were (1) preterm delivery (gestation < 37 weeks), (2) mode of delivery, (3) high (> 4000 g) or low birthweight (< 2500 g) and (4) stillbirth. Secondary outcomes were (1) very preterm delivery (< 32 weeks), (2) extremely preterm delivery (< 28 weeks), (3) small and (4) large for gestational age. Conditional logistic regression models were performed adjusting for age, ethnicity, deprivation, dysglycaemia, hypertension, thyroid disorders, number of babies born at index pregnancy, and pre-gravid BMI. Multiple imputation was performed for missing outcome data. RESULTS: 27,586 deliveries with maternal PCOS were matched for age (± 1 year) to 110,344 deliveries without PCOS. In the fully adjusted models, maternal PCOS was associated with an increased risk of (1) preterm birth [aOR: 1.11 (95% CI 1.06-1.17)], and (2) emergency caesarean, elective caesarean and instrumental vaginal compared to spontaneous delivery [aOR: 1.10 (1.05-1.15), 1.07 (1.03-1.12) and 1.04 (1.00-1.09), respectively]. There was absence of association with low birthweight, high birthweight and stillbirth. In the sensitivity analysis, the association with preterm birth [aOR: 1.31 (95% CI 1.13-1.52)], emergency caesarean [aOR: 1.15 (95% CI 1.02-1.30)], and elective caesarean [aOR: 1.03 (95% CI 1.02-1.03)] remained. While there was no significant association with any of the secondary outcomes in the primary analysis, in the sensitivity analysis maternal PCOS was associated with increased risk of extremely preterm delivery [aOR: 1.86 (95% CI 1.31-2.65)], and lower risk of small for gestational age babies [aOR: 0.74 (95% CI 0.59-0.94)]. CONCLUSIONS: Maternal PCOS was associated with increased risk of preterm and caesarean delivery. Association with low birthweight may be largely mediated by lower gestational age at birth.

Alternative pathway androgen biosynthesis and human fetal female virilization.

Androgen biosynthesis in the human fetus proceeds through the adrenal sex steroid precursor dehydroepiandrosterone, which is converted to testosterone in the gonads, followed by further activation to 5α-dihydrotestosterone in genital skin, thereby facilitating male external genital differentiation. Congenital adrenal hyperplasia due to P450 oxidoreductase deficiency results in disrupted dehydroepiandrosterone biosynthesis, explaining undervirilization in affected boys. However, many affected girls are born virilized, despite low circulating androgens. We hypothesized that this is due to a prenatally active, alternative androgen biosynthesis pathway from 17α-hydroxyprogesterone to 5α-dihydrotestosterone, which bypasses dehydroepiandrosterone and testosterone, with increased activity in congenital adrenal hyperplasia variants associated with 17α-hydroxyprogesterone accumulation. Here we employ explant cultures of human fetal organs (adrenals, gonads, genital skin) from the major period of sexual differentiation and show that alternative pathway androgen biosynthesis is active in the fetus, as assessed by liquid chromatography-tandem mass spectrometry. We found androgen receptor expression in male and female genital skin using immunohistochemistry and demonstrated that both 5α-dihydrotestosterone and adrenal explant culture supernatant induce nuclear translocation of the androgen receptor in female genital skin primary cultures. Analyzing urinary steroid excretion by gas chromatography-mass spectrometry, we show that neonates with P450 oxidoreductase deficiency produce androgens through the alternative androgen pathway during the first weeks of life. We provide quantitative in vitro evidence that the corresponding P450 oxidoreductase mutations predominantly support alternative pathway androgen biosynthesis. These results indicate a key role of alternative pathway androgen biosynthesis in the prenatal virilization of girls affected by congenital adrenal hyperplasia due to P450 oxidoreductase deficiency.

SIMBA: using Kolb's learning theory in simulation-based learning to improve participants' confidence.

BACKGROUND: Simulation via Instant Messaging- Birmingham Advance (SIMBA) delivers simulation-based learning (SBL) through WhatsApp® and Zoom® based on Kolb's experiential learning theory. This study describes how Kolb's theory was implemented in practice during SIMBA adrenal session. METHODS: SIMBA adrenal session was conducted for healthcare professionals and replicated Kolb's 4-stage cycle: (a) concrete experience-online simulation of real-life clinical scenarios, (b) reflective observation-discussion and Q&A following simulation, (c) abstract conceptualisation-post-session MCQs, and (d) active experimentation-intentions to implement the acquired knowledge in future practice. Participants' self-reported confidence levels for simulated and non-simulated cases pre- and post-SIMBA were analysed using Wilcoxon Signed-Rank test. Key takeaway and feedback were assessed quantitatively and qualitatively in a thematic analysis. RESULTS: Thirty-three participants were included in the analysis. A Wilcoxon signed-rank test showed that the SIMBA session elicited a statistically significant change in participants' self-reported confidence in their approach to Cushing's syndrome (Z = 3.873, p = 0.0001) and adrenocortical carcinoma (Z = 3.970, p < 0.0001). 93.9% (n = 31/33) and 84.8% (n = 28/33) strongly agreed/agreed the topics were applicable to their clinical practice and accommodated their personal learning style, respectively. 81.8% (n = 27/33) reported increase in knowledge on patient management, and 75.8% (n = 25/33) anticipated implementing learning points in their practice. CONCLUSIONS: SIMBA effectively adopts Kolb's theory to provide best possible experience to learners, highlighting the advantages of utilising social media platforms for SBL in medical education. The ability to conduct SIMBA sessions at modest cost internationally paves way to engage more healthcare professionals worldwide.

Insights from the genetic characterization of central precocious puberty associated with multiple anomalies.

STUDY QUESTION: Is there an (epi)genetic basis in patients with central precocious puberty (CPP) associated with multiple anomalies that unmasks underlying mechanisms or reveals novel genetic findings related to human pubertal control? SUMMARY ANSWER: In a group of 36 patients with CPP associated with multiple phenotypes, pathogenic or likely pathogenic (epi)genetic defects were identified in 12 (33%) patients, providing insights into the genetics of human pubertal control. WHAT IS KNOWN ALREADY: A few studies have described patients with CPP associated with multiple anomalies, but without making inferences on causalities of CPP. Genetic-molecular studies of syndromic cases may reveal disease genes or mechanisms, as the presentation of such patients likely indicates a genetic disorder. STUDY DESIGN, SIZE, DURATION: This translational study was based on a genetic-molecular analysis, including genome-wide high throughput methodologies, for searching structural or sequence variants implicated in CPP and DNA methylation analysis of candidate regions. PARTICIPANTS/MATERIALS, SETTING, METHODS: A cohort of 197 patients (188 girls) with CPP without structural brain lesions was submitted to a detailed clinical evaluation, allowing the selection of 36 unrelated patients (32 girls) with CPP associated with multiple anomalies. Pathogenic allelic variants of genes known to cause monogenic CPP (KISS1R, KISS1, MKRN3 and DLK1) had been excluded in the entire cohort (197 patients). All selected patients with CPP associated with multiple anomalies (n = 36) underwent methylation analysis of candidate regions and chromosomal microarray analysis. A subset (n = 9) underwent whole-exome sequencing, due to presenting familial CPP and/or severe congenital malformations and neurocognitive abnormalities. MAIN RESULTS AND THE ROLE OF CHANCE: Among the 36 selected patients with CPP, the more prevalent associated anomalies were metabolic, growth and neurocognitive conditions. In 12 (33%) of them, rare genetic abnormalities were identified: six patients presented genetic defects in loci known to be involved with CPP (14q32.2 and 7q11.23), whereas the other six presented defects in candidate genes or regions. In detail, three patients presented hypomethylation of DLK1/MEG3:IG-DMR (14q32.2 disruption or Temple syndrome), resulting from epimutation (n = 1) or maternal uniparental disomy of chromosome 14 (n = 2). Seven patients presented pathogenic copy number variants: three with de novo 7q11.23 deletions (Williams-Beuren syndrome), three with inherited Xp22.33 deletions, and one with de novo 1p31.3 duplication. Exome sequencing revealed potential pathogenic variants in two patients: a sporadic female case with frameshift variants in TNRC6B and AREL1 and a familial male case with a missense substitution in UGT2B4 and a frameshift deletion in MKKS. LIMITATIONS, REASONS FOR CAUTION: The selection of patients was based on a retrospective clinical characterization, lacking a longitudinal inclusion of consecutive patients. In addition, future studies are needed, showing the long-term (mainly reproductive) outcomes in the included patients, as most of them are not in adult life yet. WIDER IMPLICATIONS OF THE FINDINGS: The results highlighted the relevance of an integrative clinical-genetic approach in the elucidation of mechanisms and factors involved in pubertal control. Chromosome 14q32.2 disruption indicated the loss of imprinting of DLK1 as a probable mechanism of CPP. Two other chromosomal regions (7q11.23 and Xp22.33) represented new candidate loci potentially involved in this disorder of pubertal timing. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by grant number 2018/03198-0 (to A.P.M.C.) and grant number 2013/08028-1 (to A.C.V.K) from the São Paulo Research Foundation (FAPESP), and grant number 403525/2016-0 (to A.C.L.) and grant number 302849/2015-7 (to A.C.L.) and grant number 141625/2016-3 (to A.C.V.K) from the National Council for Scientific and Technological Development (CNPq). The authors have nothing to disclose. TRIAL REGISTRATION NUMBER: N/A.

Altered cortisol metabolism in individuals with HNF1A-MODY.

OBJECTIVE AND CONTEXT: Maturity onset diabetes of the young due to variants in HNF1A (HNF1A-MODY) is the most common form of monogenic diabetes. Individuals with HNF1A-MODY usually have a lean phenotype which contrasts with type 2 diabetes (T2DM). Data from hepatocytes derived from Hnf1a knock-out mice demonstrated dysregulation of 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1), which regulates glucocorticoid availability and action in target tissues, together with 11ß-HSD2 and steroid A-ring reductases, 5α- and 5ß-reductase. We proposed that altered glucocorticoid metabolism might underpin some of the phenotypic differences between patients with HNF1A-MODY and those with T2DM. DESIGN: A retrospective matched cohort study. PATIENTS AND MEASUREMENTS: 24-hours urine steroid metabolome profiling was carried out by gas chromatography-mass spectrometry in 35 subjects with HNF1A-MODY, 35 individuals with T2DM and 35 healthy controls matched for age, sex and BMI. The steroid metabolites were expressed as µg/L in all groups and measured in mid-morning urine in diabetic subjects and 24-hour urine collection in healthy controls. Hence, only ratios were compared not the individual steroids. Established ratios of glucocorticoid metabolites were used to estimate 11ß-HSD1/2 and 5α- and 5ß-reductase activities. RESULTS: While 11ß-HSD1 activity was similar in all groups, 11ß-HSD2 activity was significantly lower in subjects with HNF1A-MODY and T2DM than in healthy controls. The ratio of 5ß- to 5α-metabolites of cortisol was higher in subjects with HNF1A-MODY than in T2DM and healthy controls, probably due to increased activity of the 5ß-reductase (AKR1D1) in HNF1A-MODY. CONCLUSIONS: This is the first report of steroid metabolites in HNF1A-MODY. We have identified distinct differences in steroid metabolism pathways in subjects with HNF1A-MODY that have the potential to alter steroid hormone availability. Further studies are required to explore whether these changes link to phenotype.

Human fetal adrenal cells retain age-related stem- and endocrine-differentiation potential in culture.

The adrenal gland is a multiendocrine organ with a steroidogenic mesenchymal cortex and an inner catecholamine-producing medulla of neuroendocrine origin. After embryonic development, this plastic organ undergoes a functional postnatal remodeling. Elucidating these complex processes is pivotal for understanding the early bases of functional endocrine disorders and tumors affecting the mature gland. We developed an in vitro human adrenal cell model derived from fetal adrenal specimens at different gestational ages, consisting of neuroendocrine and cortical components and expressing the zona and functional markers of the original fetal organ. These cortical and neuroendocrine progenitor cells retain in vitro an intrinsic gestational-age-related differentiation and functional program. In vitro these cells spontaneously form 3-dimensional structure organoids with a structure similar to the fetal gland. The organoids show morphofunctional features and adrenal steroidogenic factor, steroid acute regulatory, cytochrome-P450-17A1, dosage-sensitive, sex-reversal, adrenal hypoplasia-critical region on chromosome X protein , NOTCH1, and nephroblastoma overexpressed/cysteine-rich protein 61/connective tissue growth factor/nephroblastoma overexpressed gene-3; stem (BMI1, nestin); and chromaffin (chromogranin A, tyrosine hydroxylase) markers similar to those of the populations of origin. This in vitro human adrenal system represents a unique but preliminar model for investigating the pathophysiological processes underlying physiologic adrenal remodeling and pathologic alterations involved in organ hypo- and hyperplasia and cancer.-Poli, G., Sarchielli, E., Guasti, D., Benvenuti, S., Ballerini, L., Mazzanti, B., Armignacco, R., Cantini, G., Lulli, M., Chortis, V., Arlt, W., Romagnoli, P., Vannelli, G. B., Mannelli, M., Luconi, M. Human fetal adrenal cells retain age-related stem- and endocrine-differentiation potential in culture.

The contribution of serum cortisone and glucocorticoid metabolites to detrimental bone health in patients receiving hydrocortisone therapy.

BACKGROUND: Glucocorticoid therapy is the most common cause of iatrogenic osteoporosis. Less is known regarding the effect of glucocorticoids when used as replacement therapy on bone remodelling in patients with adrenal insufficiency. Enhanced intracellular conversion of inactive cortisone to active cortisol, by 11 beta-hydroxysteroid dehydrogenase type 1(11ß-HSD1) and other enzymes leading to alterations in glucocorticoid metabolism, may contribute to a deleterious effect on bone health in this patient group. METHODS: Study design: An open crossover prospective study randomizing ten hypopituitary men, with severe ACTH deficiency, to three commonly used hydrocortisone dose regimens. MEASUREMENTS: Following 6 weeks of each regimen, patients underwent 24-h serum cortisol/cortisone sampling, measurement of bone turnover markers, and a 24-h urine collection for measurement of urinary steroid metabolites by gas chromatography-mass spectrometry (GC-MS). Serum cortisone and cortisol were analysed by liquid chromatography-mass spectrometry (LC-MS). RESULTS: Dose-related and circadian variations in serum cortisone were seen to parallel those for cortisol, indicating conversion of ingested hydrocortisone to cortisone. The median area under the curve (AUC) of serum cortisone was significantly higher in patients on dose A (20 mg/10 mg) [670.5 (IQR 621-809.2)] compared to those on dose C (10 mg/5 mg) [562.8 (IQR 520.1-619.6), p = 0.01]. A negative correlation was observed between serum cortisone and bone formation markers, OC [1-49] (r = - 0.42, p = 0.03), and PINP (r = - 0.49, p = 0.01). There was a negative correlation between the AUC of night-time serum cortisone levels with the bone formation marker, OC [1-49] (r = - 0.41, p = 0.03) but there were no significant correlations between day-time serum cortisone or cortisol with bone turnover markers. There was a negative correlation between total urinary cortisol metabolites and the bone formation markers, PINP (r = - 0.39, p = 0.04), and OC [1-49] (r = - 0.35, p = 0.06). CONCLUSION: Serum cortisol and cortisone and total urinary corticosteroid metabolites are negatively associated with bone turnover markers in patients receiving replacement doses of hydrocortisone, with nocturnal glucocorticoid exposure having a potentially greater influence on bone turnover. TRIAL REGISTRATION: Irish Medicines Board Clinical Trial Number - CT900/459/1 and EudraCT Number - 2007-005018-37 . Registration date: 07-09-2007.

Natural History of Adrenal Incidentalomas With and Without Mild Autonomous Cortisol Excess: A Systematic Review and Meta-analysis.

Background: Adrenal incidentalomas are mostly benign nonfunctioning adrenal tumors (NFATs) or adenomas causing mild autonomous cortisol excess (MACE), but their natural history is unclear. Purpose: To summarize the follow-up data of adults with NFAT or MACE to determine the proportions of tumor growth, malignant transformation, and incident changes in hormone function; the prevalence of incident cardiometabolic comorbid conditions; and mortality. Data Sources: MEDLINE, Embase, Cochrane, and Scopus (January 1990 to February 2019) and bibliographies of identified articles, without language restriction. Study Selection: Studies that included 20 or more conservatively managed patients with NFAT or MACE and reported outcomes at baseline and after at least 12 months of follow-up. Data Extraction: Pairs of reviewers extracted outcomes and assessed methodological quality. Data Synthesis: Thirty-two studies reported outcomes of 4121 patients with NFAT or MACE, 61.5% of whom were women; the mean age was 60.2 years, and mean follow-up was 50.2 months. Mean tumor growth was 2 mm over 52.8 months. Clinically significant tumor enlargement (≥10 mm) occurred in 2.5% of patients, and none developed adrenal cancer. Clinically overt hormone excess was unlikely to develop (<0.1%) in patients with NFAT or MACE. Only 4.3% of patients with NFAT developed MACE, and preexisting MACE was unlikely to resolve (<0.1%). Hypertension, obesity, dyslipidemia, and type 2 diabetes were highly prevalent (60.0%, 42.0%, 33.7%, and 18.1% of patients, respectively) and were more likely to develop and worsen in MACE than NFAT. New cardiovascular events were more prevalent in MACE (15.5%) than NFAT (6.4%). Mortality was 11.2% and was similar between NFAT and MACE. Limitation: Evidence was scarce, and definitions of MACE and comorbid conditions were heterogeneous. Conclusion: During follow-up, NFAT and MACE do not show clinically relevant changes in size or hormonal function, but they may carry an increased risk for cardiometabolic comorbid conditions. Primary Funding Source: None.

Simulation via instant messaging-Birmingham advance (SIMBA) model helped improve clinicians' confidence to manage cases in diabetes and endocrinology.

BACKGROUND: Simulation-based learning (SBL) has been increasingly used in both undergraduate and postgraduate medical training curricula. The aim of Simulation via Instant Messaging-Birmingham Advance (SIMBA) is to create a simple virtual learning environment to improve trainees' self-reported confidence in diabetes and Endocrinology. METHODS: This study was done as part of the continuous professional development for Health Education England West Midlands speciality trainees in diabetes and Endocrinology. Standardized transcripts of anonymized real-life endocrinology (endocrine session) and diabetes cases (diabetes session) were used in the simulation model. Trainees interacted with moderators through WhatsApp® in this model. All cases were then discussed in detail by a consultant endocrinologist with reference to local, national and international guidelines. Trainee acceptance rate and improvement in their self-reported confidence levels post-simulation were assessed. RESULTS: 70.8% (n = 17/24) and 75% (n = 18/24) strongly agreed the simulation session accommodated their personal learning style and the session was engaging. 66.7% (n = 16/24) strongly felt that the simulation was worth their time. In the endocrine session, there was a significant improvement in trainees' confidence in the management of craniopharyngioma (p = 0.0179) and acromegaly (p = 0.0025). There was a trend towards improved confidence levels to manage Cushing's disease and macroprolactinoma. In diabetes session, there was a significant improvement in trainees' confidence to interpret continuous glucose monitor readings (p = 0.01). There was a trend towards improvement for managing monogenic diabetes, hypoglycaemic unawareness and interpreting Libre readings. Overall, there was a significant improvement in trainees' confidence in managing cases that were discussed post-simulation. CONCLUSION: SIMBA is an effective learning model to improve trainees' confidence to manage various diabetes and endocrine case scenarios. More sessions with a variety of other speciality case scenarios are needed to further assess SIMBA's effectiveness and application in other areas of medical training.

Human DHEA sulfation requires direct interaction between PAPS synthase 2 and DHEA sulfotransferase SULT2A1.

The high-energy sulfate donor 3'-phosphoadenosine-5'-phosphosulfate (PAPS), generated by human PAPS synthase isoforms PAPSS1 and PAPSS2, is required for all human sulfation pathways. Sulfotransferase SULT2A1 uses PAPS for sulfation of the androgen precursor dehydroepiandrosterone (DHEA), thereby reducing downstream activation of DHEA to active androgens. Human PAPSS2 mutations manifest with undetectable DHEA sulfate, androgen excess, and metabolic disease, suggesting that ubiquitous PAPSS1 cannot compensate for deficient PAPSS2 in supporting DHEA sulfation. In knockdown studies in human adrenocortical NCI-H295R1 cells, we found that PAPSS2, but not PAPSS1, is required for efficient DHEA sulfation. Specific APS kinase activity, the rate-limiting step in PAPS biosynthesis, did not differ between PAPSS1 and PAPSS2. Co-expression of cytoplasmic SULT2A1 with a cytoplasmic PAPSS2 variant supported DHEA sulfation more efficiently than co-expression with nuclear PAPSS2 or nuclear/cytosolic PAPSS1. Proximity ligation assays revealed protein-protein interactions between SULT2A1 and PAPSS2 and, to a lesser extent, PAPSS1. Molecular docking studies showed a putative binding site for SULT2A1 within the PAPSS2 APS kinase domain. Energy-dependent scoring of docking solutions identified the interaction as specific for the PAPSS2 and SULT2A1 isoforms. These findings elucidate the mechanistic basis for the selective requirement for PAPSS2 in human DHEA sulfation.

Novel methods in adrenal research: a metabolomics approach.

Metabolic alterations have implications in a spectrum of tissue functions and disease. Aided by novel molecular biological and computational tools, our understanding of physiological and pathological processes underpinning endocrine and endocrine-related disease has significantly expanded over the last decade. Herein, we focus on novel metabolomics-related methodologies in adrenal research: in situ metabolomics by mass spectrometry imaging, steroid metabolomics by gas and liquid chromatography-mass spectrometry, energy pathway metabologenomics by liquid chromatography-mass spectrometry-based metabolomics of Krebs cycle intermediates, and cellular reprogramming to generate functional steroidogenic cells and hence to modulate the steroid metabolome. All four techniques to assess and/or modulate the metabolome in biological systems provide tremendous opportunities to manage neoplastic and non-neoplastic disease of the adrenal glands in the era of precision medicine. In this context, we discuss emerging clinical applications and/or promising metabolic-driven research towards diagnostic, prognostic, predictive and therapeutic biomarkers in tumours arising from the adrenal gland and extra-adrenal paraganglia as well as modern approaches to delineate and reprogram adrenal metabolism.

Glucocorticoid activation by 11ß-hydroxysteroid dehydrogenase enzymes in relation to inflammation and glycaemic control in chronic kidney disease: A cross-sectional study.

OBJECTIVE: Patients with chronic kidney disease (CKD) have dysregulated cortisol metabolism secondary to changes in 11ß-hydroxysteroid dehydrogenase (11ß-HSD) enzymes. The determinants of this and its clinical implications are poorly defined. METHODS: We performed a cross-sectional study to characterize shifts in cortisol metabolism in relation to renal function, inflammation and glycaemic control. Systemic activation of cortisol by 11ß-HSD was measured as the metabolite ratio (tetrahydrocortisol [THF]+5α-tetrahydrocortisol [5αTHF])/tetrahydrocortisone (THE) in urine. RESULTS: The cohort included 342 participants with a median age of 63 years, median estimated glomerular filtration rate (eGFR) of 28 mL/min/1.73 m2 and median urine albumin-creatinine ratio of 35.5 mg/mmol. (THF+5αTHF)/THE correlated negatively with eGFR (Spearman's ρ = -0.116, P = 0.032) and positively with C-reactive protein (ρ = 0.208, P < 0.001). In multivariable analysis, C-reactive protein remained a significant independent predictor of (THF+5αTHF)/THE, but eGFR did not. Elevated (THF+5αTHF)/THE was associated with HbA1c (ρ = 0.144, P = 0.008) and diabetes mellitus (odds ratio for high vs low tertile of (THF+5αTHF)/THE 2.57, 95% confidence interval 1.47-4.47). Associations with diabetes mellitus and with HbA1c among the diabetic subgroup were independent of eGFR, C-reactive protein, age, sex and ethnicity. CONCLUSIONS: In summary, glucocorticoid activation by 11ß-HSD in our cohort comprising a spectrum of renal function was associated with inflammation and impaired glucose control.

Serum testosterone, sex hormone-binding globulin and sex-specific risk of incident type 2 diabetes in a retrospective primary care cohort.

OBJECTIVE: Previous studies suggest that androgens have a sexually dimorphic impact on metabolic dysfunction. However, the sex-specific link between circulating androgens and risk of type 2 diabetes mellitus (T2DM) has not been examined in a large scale, longitudinal cohort, a task we undertook in this study. DESIGN: A retrospective cohort study in a UK primary care database. PATIENTS: We included men and women with available serum testosterone and sex hormone-binding globulin (SHBG) results. MEASUREMENTS: We categorized serum concentrations according to clinically relevant cut-off points and calculated crude and adjusted T2DM Incidence Rate Ratios (IRRs and aIRRs). RESULTS: Serum testosterone concentrations were available in 70 541 men and 81 889 women; serum SHBG was available in 15 907 men and 42 034 women. In comparison to a reference cohort with serum testosterone ≥20 nmol/L, men with lower serum testosterone had a significantly increased risk of T2DM, with the highest risk in those with serum testosterone <7 nmol/L (aIRR 2.71, 95% CI 2.34-3.14, P < 0.001). In women, the risk of T2DM started to increase significantly when serum testosterone concentrations exceeded 1.5 nmol/L, with the highest risk in women with serum testosterone ≥3.5 nmol/L (aIRR 1.98, 95% CI 1.55-2.52, P < 0.001). These observations were verified in a continuous rather than categorized analysis. The risk of T2DM increased in men and women with serum SHBG <40 and <50 nmol/L, respectively. CONCLUSIONS/INTERPRETATION: In this longitudinal study, we found sexually dimorphic associations between serum testosterone and risk of incident T2DM. Androgen deficiency and excess should be considered important risk factors for diabetes in men and women, respectively.

Increased central adiposity and decreased subcutaneous adipose tissue 11ß-hydroxysteroid dehydrogenase type 1 are associated with deterioration in glucose tolerance-A longitudinal cohort study.

OBJECTIVE AND CONTEXT: Increasing adiposity, ageing and tissue-specific regeneration of cortisol through the activity of 11ß-hydroxysteroid dehydrogenase type 1 have been associated with deterioration in glucose tolerance. We undertook a longitudinal, prospective clinical study to determine if alterations in local glucocorticoid metabolism track with changes in glucose tolerance. DESIGN, PATIENTS, AND MEASUREMENTS: Sixty-five overweight/obese individuals (mean age 50.3 ± 7.3 years) underwent oral glucose tolerance testing, body composition assessment, subcutaneous adipose tissue biopsy and urinary steroid metabolite analysis annually for up to 5 years. Participants were categorized into those in whom glucose tolerance deteriorated ("deteriorators") or improved ("improvers"). RESULTS: Deteriorating glucose tolerance was associated with increasing total and trunk fat mass and increased subcutaneous adipose tissue expression of lipogenic genes. Subcutaneous adipose tissue 11ß-HSD1 gene expression decreased in deteriorators, and at study completion, it was highest in the improvers. There was a significant negative correlation between change in area under the curve glucose and 11ß-HSD1 expression. Global 11ß-HSD1 activity did not change and was not different between deteriorators and improvers at baseline or follow-up. CONCLUSION: Longitudinal deterioration in metabolic phenotype is not associated with increased 11ß-HSD1 activity, but decreased subcutaneous adipose tissue gene expression. These changes may represent a compensatory mechanism to decrease local glucocorticoid exposure in the face of an adverse metabolic phenotype.