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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.
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Diabetes Mellitus Tipo 2 , Animales , Estudios de Cohortes , Factor Nuclear 1-alfa del Hepatocito/genética , Humanos , Hidrocortisona , Ratones , Estudios RetrospectivosRESUMEN
Steroid 5ß-reductase (AKR1D1) is highly expressed in human liver where it inactivates endogenous glucocorticoids and catalyses an important step in bile acid synthesis. Endogenous and synthetic glucocorticoids are potent regulators of metabolic phenotype and play a crucial role in hepatic glucose metabolism. However, the potential of synthetic glucocorticoids to be metabolised by AKR1D1 as well as to regulate its expression and activity has not been investigated. The impact of glucocorticoids on AKR1D1 activity was assessed in human liver HepG2 and Huh7 cells; AKR1D1 expression was assessed by qPCR and Western blotting. Genetic manipulation of AKR1D1 expression was conducted in HepG2 and Huh7 cells and metabolic assessments were made using qPCR. Urinary steroid metabolite profiling in healthy volunteers was performed pre- and post-dexamethasone treatment, using gas chromatography-mass spectrometry. AKR1D1 metabolised endogenous cortisol, but cleared prednisolone and dexamethasone less efficiently. In vitro and in vivo, dexamethasone decreased AKR1D1 expression and activity, further limiting glucocorticoid clearance and augmenting action. Dexamethasone enhanced gluconeogenic and glycogen synthesis gene expression in liver cell models and these changes were mirrored by genetic knockdown of AKR1D1 expression. The effects of AKR1D1 knockdown were mediated through multiple nuclear hormone receptors, including the glucocorticoid, pregnane X and farnesoid X receptors. Glucocorticoids down-regulate AKR1D1 expression and activity and thereby reduce glucocorticoid clearance. In addition, AKR1D1 down-regulation alters the activation of multiple nuclear hormone receptors to drive changes in gluconeogenic and glycogen synthesis gene expression profiles, which may exacerbate the adverse impact of exogenous glucocorticoids.
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Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Glucocorticoides/farmacología , Gluconeogénesis/efectos de los fármacos , Hígado/enzimología , Oxidorreductasas/efectos de los fármacos , Adulto , Células Cultivadas , Voluntarios Sanos , Hepatocitos , Humanos , MasculinoRESUMEN
OBJECTIVE: Non-alcoholic fatty liver disease (NAFLD) is the hepatic manifestation of metabolic syndrome. Steroid hormones and bile acids are potent regulators of hepatic carbohydrate and lipid metabolism. Steroid 5ß-reductase (AKR1D1) is highly expressed in human liver where it inactivates steroid hormones and catalyzes a fundamental step in bile acid synthesis. METHODS: Human liver biopsies were obtained from 34 obese patients and AKR1D1 mRNA expression levels were measured using qPCR. Genetic manipulation of AKR1D1 was performed in human HepG2 and Huh7 liver cell lines. Metabolic assessments were made using transcriptome analysis, western blotting, mass spectrometry, clinical biochemistry, and enzyme immunoassays. RESULTS: In human liver biopsies, AKR1D1 expression decreased with advancing steatosis, fibrosis and inflammation. Expression was decreased in patients with type 2 diabetes. In human liver cell lines, AKR1D1 knockdown decreased primary bile acid biosynthesis and steroid hormone clearance. RNA-sequencing identified disruption of key metabolic pathways, including insulin action and fatty acid metabolism. AKR1D1 knockdown increased hepatocyte triglyceride accumulation, insulin sensitivity, and glycogen synthesis, through increased de novo lipogenesis and decreased ß-oxidation, fueling hepatocyte inflammation. Pharmacological manipulation of bile acid receptor activation prevented the induction of lipogenic and carbohydrate genes, suggesting that the observed metabolic phenotype is driven through bile acid rather than steroid hormone availability. CONCLUSIONS: Genetic manipulation of AKR1D1 regulates the metabolic phenotype of human hepatoma cell lines, driving steatosis and inflammation. Taken together, the observation that AKR1D1 mRNA is down-regulated with advancing NAFLD suggests that it may have a crucial role in the pathogenesis and progression of the disease.
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Hepatocitos/metabolismo , Enfermedad del Hígado Graso no Alcohólico/metabolismo , Oxidorreductasas/fisiología , Fenotipo , Ácidos y Sales Biliares/metabolismo , Células Hep G2 , Humanos , Inflamación/etiología , Enfermedad del Hígado Graso no Alcohólico/patología , Obesidad , Oxidorreductasas/genética , ARN Mensajero/metabolismoRESUMEN
Steroid hormones, including glucocorticoids and androgens, have potent actions to regulate many cellular processes within the liver. The steroid A-ring reductase, 5ß-reductase (AKR1D1), is predominantly expressed in the liver, where it inactivates steroid hormones and, in addition, plays a crucial role in bile acid synthesis. However, the precise functional role of AKR1D1 to regulate steroid hormone action in vitro has not been demonstrated. We have therefore hypothesised that genetic manipulation of AKR1D1 has the potential to regulate glucocorticoid availability and action in human hepatocytes. In both liver (HepG2) and non-liver cell (HEK293) lines, AKR1D1 over-expression increased glucocorticoid clearance with a concomitant decrease in the activation of the glucocorticoid receptor and the down-stream expression of glucocorticoid target genes. Conversely, knockdown of AKR1D1 using siRNA decreased glucocorticoid clearance and reduced the generation of 5ß-reduced metabolites. In addition, the two 5α-reductase inhibitors finasteride and dutasteride failed to effectively inhibit AKR1D1 activity in either cell-free or hepatocellular systems. Through manipulation of AKR1D1 expression and activity, we have demonstrated its potent ability to regulate glucocorticoid availability and receptor activation within human hepatoma cells. These data suggest that AKR1D1 may have an important role in regulating endogenous (and potentially exogenous) glucocorticoid action that may be of particular relevance to physiological and pathophysiological processes affecting the liver.
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Carcinoma Hepatocelular/metabolismo , Glucocorticoides/metabolismo , Neoplasias Hepáticas/metabolismo , Oxidorreductasas/metabolismo , Receptores de Glucocorticoides/metabolismo , Células HEK293 , Células Hep G2 , Humanos , Hígado/metabolismoRESUMEN
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.
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11-beta-Hidroxiesteroide Deshidrogenasa de Tipo 1/metabolismo , Adiposidad/fisiología , Grasa Subcutánea/metabolismo , Adiposidad/genética , Corticoesteroides/metabolismo , Corticoesteroides/orina , Adulto , Femenino , Glucocorticoides/metabolismo , Glucocorticoides/orina , Humanos , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
BACKGROUND: Adrenal aldosterone excess is the most common cause of secondary hypertension and is associated with increased cardiovascular morbidity. However, adverse metabolic risk in primary aldosteronism extends beyond hypertension, with increased rates of insulin resistance, type 2 diabetes, and osteoporosis, which cannot be easily explained by aldosterone excess. METHODS: We performed mass spectrometry-based analysis of a 24-hour urine steroid metabolome in 174 newly diagnosed patients with primary aldosteronism (103 unilateral adenomas, 71 bilateral adrenal hyperplasias) in comparison to 162 healthy controls, 56 patients with endocrine inactive adrenal adenoma, 104 patients with mild subclinical, and 47 with clinically overt adrenal cortisol excess. We also analyzed the expression of cortisol-producing CYP11B1 and aldosterone-producing CYP11B2 enzymes in adenoma tissue from 57 patients with aldosterone-producing adenoma, employing immunohistochemistry with digital image analysis. RESULTS: Primary aldosteronism patients had significantly increased cortisol and total glucocorticoid metabolite excretion (all P < 0.001), only exceeded by glucocorticoid output in patients with clinically overt adrenal Cushing syndrome. Several surrogate parameters of metabolic risk correlated significantly with glucocorticoid but not mineralocorticoid output. Intratumoral CYP11B1 expression was significantly associated with the corresponding in vivo glucocorticoid excretion. Unilateral adrenalectomy resolved both mineralocorticoid and glucocorticoid excess. Postoperative evidence of adrenal insufficiency was found in 13 (29%) of 45 consecutively tested patients. CONCLUSION: Our data indicate that glucocorticoid cosecretion is frequently found in primary aldosteronism and contributes to associated metabolic risk. Mineralocorticoid receptor antagonist therapy alone may not be sufficient to counteract adverse metabolic risk in medically treated patients with primary aldosteronism. FUNDING: Medical Research Council UK, Wellcome Trust, European Commission.
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Context: The classic androgen synthesis pathway proceeds via dehydroepiandrosterone, androstenedione, and testosterone to 5α-dihydrotestosterone. However, 5α-dihydrotestosterone synthesis can also be achieved by an alternative pathway originating from 17α-hydroxyprogesterone (17OHP), which accumulates in congenital adrenal hyperplasia (CAH). Similarly, recent work has highlighted androstenedione-derived 11-oxygenated 19-carbon steroids as active androgens, and in CAH, androstenedione is generated directly from 17OHP. The exact contribution of alternative pathway activity to androgen excess in CAH and its response to glucocorticoid (GC) therapy is unknown. Objective: We sought to quantify classic and alternative pathway-mediated androgen synthesis in CAH, their diurnal variation, and their response to conventional GC therapy and modified-release hydrocortisone. Methods: We used urinary steroid metabolome profiling by gas chromatography-mass spectrometry for 24-hour steroid excretion analysis, studying the impact of conventional GCs (hydrocortisone, prednisolone, and dexamethasone) in 55 adults with CAH and 60 controls. We studied diurnal variation in steroid excretion by comparing 8-hourly collections (23:00-7:00, 7:00-15:00, and 15:00-23:00) in 16 patients with CAH taking conventional GCs and during 6 months of treatment with modified-release hydrocortisone, Chronocort. Results: Patients with CAH taking conventional GCs showed low excretion of classic pathway androgen metabolites but excess excretion of the alternative pathway signature metabolites 3α,5α-17-hydroxypregnanolone and 11ß-hydroxyandrosterone. Chronocort reduced 17OHP and alternative pathway metabolite excretion to near-normal levels more consistently than other GC preparations. Conclusions: Alternative pathway-mediated androgen synthesis significantly contributes to androgen excess in CAH. Chronocort therapy appears superior to conventional GC therapy in controlling androgen synthesis via alternative pathways through attenuation of their major substrate, 17OHP.
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Hiperplasia Suprarrenal Congénita/tratamiento farmacológico , Andrógenos/metabolismo , Ritmo Circadiano , Glucocorticoides/administración & dosificación , Hidrocortisona/administración & dosificación , 17-alfa-Hidroxipregnenolona/orina , Adolescente , Hiperplasia Suprarrenal Congénita/metabolismo , Hiperplasia Suprarrenal Congénita/orina , Adulto , Androsterona/análogos & derivados , Androsterona/orina , Cortodoxona/análogos & derivados , Cortodoxona/orina , Preparaciones de Acción Retardada , Dexametasona/uso terapéutico , Femenino , Cromatografía de Gases y Espectrometría de Masas , Glucocorticoides/uso terapéutico , Humanos , Hidrocortisona/uso terapéutico , Masculino , Persona de Mediana Edad , Prednisolona/uso terapéutico , Pregnanotriol/análogos & derivados , Pregnanotriol/orina , Adulto JovenRESUMEN
Androgen and estrogen biosynthesis in mammals requires the 17,20-lyase activity of cytochrome P450 17A1 (steroid 17-hydroxylase/17,20-lyase). Maximal 17,20-lyase activity in vitro requires the presence of cytochrome b5 (b5), and rare cases of b5 deficiency in human beings causes isolated 17,20-lyase deficiency. To study the consequences of conditional b5 removal from testicular Leydig cells in an animal model, we generated Cyb5(flox/flox):Sf1-Cre (LeyKO) mice. The LeyKO male mice had normal body weights, testis and sex organ weights, and fertility compared with littermates. Basal serum and urine steroid profiles of LeyKO males were not significantly different than littermates. In contrast, marked 17-hydroxyprogesterone accumulation (100-fold basal) and reduced testosterone synthesis (27% of littermates) were observed after human chorionic gonadotropin stimulation in LeyKO animals. Testis homogenates from LeyKO mice showed reduced 17,20-lyase activity and a 3-fold increased 17-hydroxylase to 17,20-lyase activity ratio, which were restored to normal upon addition of recombinant b5. We conclude that Leydig cell b5 is required for maximal androgen synthesis and to prevent 17-hydroxyprogesterone accumulation in the mouse testis; however, the b5-independent 17,20-lyase activity of mouse steroid 17-hydroxylase/17,20-lyase is sufficient for normal male genital development and fertility. LeyKO male mice are a good model for the biochemistry but not the physiology of isolated 17,20-lyase deficiency in human beings.
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Citocromos b5/genética , Células Intersticiales del Testículo/enzimología , Esteroide 17-alfa-Hidroxilasa/metabolismo , 17-alfa-Hidroxiprogesterona/sangre , Animales , Citocromos b5/metabolismo , Femenino , Fertilidad , Masculino , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , FenotipoRESUMEN
CONTEXT: 5α-Reductase 1 and 2 (SRD5A1, SRD5A2) inactivate cortisol to 5α-dihydrocortisol in addition to their role in the generation of DHT. Dutasteride (dual SRD5A1 and SRD5A2 inhibitor) and finasteride (selective SRD5A2 inhibitor) are commonly prescribed, but their potential metabolic effects have only recently been identified. OBJECTIVE: Our objective was to provide a detailed assessment of the metabolic effects of SRD5A inhibition and in particular the impact on hepatic lipid metabolism. DESIGN: We conducted a randomized study in 12 healthy male volunteers with detailed metabolic phenotyping performed before and after a 3-week treatment with finasteride (5 mg od) or dutasteride (0.5 mg od). Hepatic magnetic resonance spectroscopy (MRS) and two-step hyperinsulinemic euglycemic clamps incorporating stable isotopes with concomitant adipose tissue microdialysis were used to evaluate carbohydrate and lipid flux. Analysis of the serum metabolome was performed using ultra-HPLC-mass spectrometry. SETTING: The study was performed in the Wellcome Trust Clinical Research Facility, Queen Elizabeth Hospital, Birmingham, United Kingdom. MAIN OUTCOME MEASURE: Incorporation of hepatic lipid was measured with MRS. RESULTS: Dutasteride, not finasteride, increased hepatic insulin resistance. Intrahepatic lipid increased on MRS after dutasteride treatment and was associated with increased rates of de novo lipogenesis. Adipose tissue lipid mobilization was decreased by dutasteride. Analysis of the serum metabolome demonstrated that in the fasted state, dutasteride had a significant effect on lipid metabolism. CONCLUSIONS: Dual-SRD5A inhibition with dutasteride is associated with increased intrahepatic lipid accumulation.
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Inhibidores de 5-alfa-Reductasa/farmacología , Dutasterida/farmacología , Metabolismo de los Lípidos/efectos de los fármacos , Hígado/metabolismo , Proteínas de la Membrana/antagonistas & inhibidores , 3-Oxo-5-alfa-Esteroide 4-Deshidrogenasa , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/metabolismo , Adulto , Metabolismo de los Hidratos de Carbono/efectos de los fármacos , Finasterida/farmacología , Técnica de Clampeo de la Glucosa , Humanos , Resistencia a la Insulina , Hígado/efectos de los fármacos , Masculino , Metaboloma/efectos de los fármacos , Esteroides/metabolismoRESUMEN
CONTEXT: Patients with hypopituitarism have increased morbidity and mortality. There is ongoing debate about the optimum glucocorticoid (GC) replacement therapy. OBJECTIVE: To assess the effect of GC replacement in hypopituitarism on corticosteroid metabolism and its impact on body composition. DESIGN AND PATIENTS: We assessed the urinary corticosteroid metabolite profile (using gas chromatography/mass spectrometry) and body composition (clinical parameters and full body DXA) of 53 patients (19 female, median age 46 years) with hypopituitarism (33 ACTH-deficient/20 ACTH-replete) (study A). The corticosteroid metabolite profile of ten patients with ACTH deficiency was then assessed prospectively in a cross over study using three hydrocortisone (HC) dosing regimens (20/10âmg, 10/10âmg and 10/5âmg) (study B) each for 6 weeks. 11 beta-hydroxysteroid dehydrogenase 1 (11ß-HSD1) activity was assessed by urinary THF+5α-THF/THE. SETTING: Endocrine Centres within University Teaching Hospitals in the UK and Ireland. MAIN OUTCOME MEASURES: Urinary corticosteroid metabolite profile and body composition assessment. RESULTS: In study A, when patients were divided into three groups - patients not receiving HC and patients receiving HC≤20âmg/day or HC>20âmg/day - patients in the group receiving the highest daily dose of HC had significantly higher waist-to-hip ratio (WHR) than the ACTH replete group. They also had significantly elevated THF+5α-THF/THE (P=0.0002) and total cortisol metabolites (P=0.015). In study B, patients on the highest HC dose had significantly elevated total cortisol metabolites and all patients on HC had elevated THF+5α-THF/THE ratios when compared to controls. CONCLUSIONS: In ACTH-deficient patients daily HC doses of >20âmg/day have increased WHR, THF+5α-THF/THE ratios and total cortisol metabolites. GC metabolism and induction of 11ß-HSD1 may play a pivitol role in the development of the metabolically adverse hypopituitary phenotype.
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Hormona Adrenocorticotrópica/deficiencia , Composición Corporal/efectos de los fármacos , Glucocorticoides/metabolismo , Hidrocortisona/metabolismo , Hipopituitarismo/tratamiento farmacológico , Hipopituitarismo/metabolismo , Adulto , Anciano , Estudios Cruzados , Estudios Transversales , Femenino , Glucocorticoides/administración & dosificación , Glucocorticoides/orina , Humanos , Hidrocortisona/administración & dosificación , Hidrocortisona/orina , Hipopituitarismo/orina , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Relación Cintura-Cadera , Adulto JovenRESUMEN
CONTEXT: PAPSS2 (PAPS synthase 2) provides the universal sulfate donor PAPS (3'-phospho-adenosine-5'-phosphosulfate) to all human sulfotransferases, including SULT2A1, responsible for sulfation of the crucial androgen precursor dehydroepiandrosterone (DHEA). Impaired DHEA sulfation is thought to increase the conversion of DHEA toward active androgens, a proposition supported by the previous report of a girl with inactivating PAPSS2 mutations who presented with low serum DHEA sulfate and androgen excess, clinically manifesting with premature pubarche and early-onset polycystic ovary syndrome. PATIENTS AND METHODS: We investigated a family harboring two novel PAPSS2 mutations, including two compound heterozygous brothers presenting with disproportionate short stature, low serum DHEA sulfate, but normal serum androgens. Patients and parents underwent a DHEA challenge test comprising frequent blood sampling and urine collection before and after 100 mg DHEA orally, with subsequent analysis of DHEA sulfation and androgen metabolism by mass spectrometry. The functional impact of the mutations was investigated in silico and in vitro. RESULTS: We identified a novel PAPSS2 frameshift mutation, c.1371del, p.W462Cfs*3, resulting in complete disruption, and a novel missense mutation, c.809G>A, p.G270D, causing partial disruption of DHEA sulfation. Both patients and their mother, who was heterozygous for p.W462Cfs*3, showed increased 5α-reductase activity at baseline and significantly increased production of active androgens after DHEA intake. The mother had a history of oligomenorrhea and chronic anovulation that required clomiphene for ovulation induction. CONCLUSIONS: We provide direct in vivo evidence for the significant functional impact of mutant PAPSS2 on DHEA sulfation and androgen activation. Heterozygosity for PAPSS2 mutations can be associated with a phenotype resembling polycystic ovary syndrome.
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Andrógenos/metabolismo , Deshidroepiandrosterona/metabolismo , Hiperandrogenismo/genética , Complejos Multienzimáticos/genética , Mutación , Sulfato Adenililtransferasa/genética , Sulfatos/metabolismo , Adolescente , Adulto , Sulfato de Deshidroepiandrosterona/metabolismo , Familia , Femenino , Humanos , Hiperandrogenismo/metabolismo , Masculino , Persona de Mediana Edad , Complejos Multienzimáticos/metabolismo , Sulfato Adenililtransferasa/metabolismo , Adulto JovenRESUMEN
CONTEXT: Low birth weight is associated with adverse metabolic outcome in adulthood. Exposure to glucocorticoid (GC) excess in utero is associated with decreased birth weight, but the prospective longitudinal relationship between GC metabolism and growth has not been examined. OBJECTIVE: We have hypothesized that changes in GC metabolism leading to increased availability may impair growth. DESIGN: This was a prospective, longitudinal study with clinical measurements and 24-hour urinary steroid metabolite analysis at 1, 4, 12, 26, and 52 weeks after delivery in mothers and their babies. SETTING: The study was conducted with observations and samples collected in the volunteers' own homes. PARTICIPANTS: Healthy mothers and newborn babies/infants participated in the study. INTERVENTIONS: There were no interventions. MAIN OUTCOME MEASURES: Urinary steroid metabolite excretion quantified by gas chromatography/mass spectroscopy across the first year of life in relation to change in weight was measured. RESULTS: The total production of the GC metabolites quantified increased across the first year of life. Markers of 11ß-hydroxysteroid dehydrogenase type 1 activity increased from the age of 3 months as did those of 5α-reductase activity. After correcting for confounding variables, low markers of 11ß-hydroxysteroid dehydrogenase type 2 activity was associated with reduced absolute weight and decreased weight gain over the first year of life. In the mothers, 5α-reductase activity was low at birth and progressively increased to normal over the first 6 months postpartum. CONCLUSIONS: Increased GC exposure as a consequence of reduced 11ß-hydroxysteroid dehydrogenase type 2 activity is likely to be a critical determinant of growth in early life. This not only highlights the central role of GCs and their metabolism, but also emphasizes the need for detailed longitudinal analyses.
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11-beta-Hidroxiesteroide Deshidrogenasa de Tipo 2/metabolismo , Peso Corporal/fisiología , Desarrollo Infantil/fisiología , Aumento de Peso/fisiología , 11-beta-Hidroxiesteroide Deshidrogenasa de Tipo 1/metabolismo , Femenino , Humanos , Lactante , Recién Nacido , Masculino , Estudios ProspectivosRESUMEN
CONTEXT: Polycystic ovary syndrome (PCOS) is a triad of anovulation, insulin resistance, and hyperandrogenism. Androgen excess may correlate with metabolic risk and PCOS consensus criteria define androgen excess on the basis of serum T. Here we studied the utility of the androgen precursor serum androstenedione (A) in conjunction with serum T for predicting metabolic dysfunction in PCOS. PATIENTS AND METHODS: Eighty-six PCOS patients fulfilling Rotterdam diagnostic consensus criteria and 43 age- and body mass index-matched controls underwent measurement of serum androgens by tandem mass spectrometry and an oral glucose tolerance test with homeostatic model assessment of insulin resistance and insulin sensitivity index calculation. We analyzed 24-hour urine androgen excretion by gas chromatography/mass spectrometry. RESULTS: PCOS patients had higher levels of serum androgens and urinary androgen metabolites than controls (all P < .001). Within the PCOS cohort, both serum A and T were positively correlated with the free androgen index (T × 100/SHBG) and total androgen metabolite excretion (all P < .001). All subjects with T above the normal reference range [high T (HT)] also had high A (HA/HT group, n = 56). However, the remaining 30 patients had normal T levels, either in the presence of HA (HA/NT; n = 20) or normal A (NA/NT; n = 10). The groups did not differ in age or BMI. The HA/HT and HA/NT groups had higher total androgen excretion than NA/NT (P < .01 and P < .05, respectively). Multiple linear regression showed a strong negative association between serum androstenedione and insulin sensitivity. The incidence of dysglycemia according to an oral glucose tolerance test increased with the severity of androgen phenotype (NA/NT, 0%; HA/NT, 14%; HA/HT, 25%, P = .03). CONCLUSION: Simultaneous measurement of serum T and A represents a useful tool for predicting metabolic risk in PCOS women. HA levels are a sensitive indicator of PCOS-related androgen excess.
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Androstenodiona/sangre , Hiperandrogenismo/diagnóstico , Síndrome del Ovario Poliquístico/diagnóstico , Síndrome del Ovario Poliquístico/metabolismo , Adulto , Andrógenos/sangre , Andrógenos/orina , Androstenodiona/orina , Estudios de Casos y Controles , Estudios Transversales , Femenino , Humanos , Hiperandrogenismo/complicaciones , Hiperandrogenismo/metabolismo , Fenotipo , Síndrome del Ovario Poliquístico/complicaciones , Valor Predictivo de las Pruebas , Adulto JovenRESUMEN
CONTEXT: Despite lifelong steroid hormone replacement, there is excess morbidity and mortality associated with autoimmune Addison's disease. In health, adrenocortical cells undergo continuous self-renewal from a population of subcapsular progenitor cells, under the influence of ACTH, suggesting a therapeutic possibility. OBJECTIVE: We aimed to determine whether tetracosactide (synthetic ACTH1-24) could revive adrenal steroidogenic function in autoimmune Addison's disease. DESIGN, SETTING, AND PATIENTS: Thirteen patients (aged 16-65 y) with established autoimmune Addison's disease for more than 1 year were recruited at the Newcastle University Clinical Research Facility. INTERVENTION: The intervention included a 20-week study of regular sc tetracosactide (ACTH1-24) therapy. MAIN OUTCOME MEASURES: Serum and urine corticosteroids were measured during medication withdrawal at baseline and every 5 weeks during the study. RESULTS: Serum cortisol levels remained less than 100 nmol/L in 11 of 13 participants throughout the study. However, two women achieved peak serum cortisol concentrations greater than 400 nmol/L after 10 and 29 weeks of tetracosactide therapy, respectively, allowing withdrawal of corticosteroid replacement. Concurrently, urine glucocorticoid and mineralocorticoid metabolite excretion increased from subnormal to above the median of healthy controls. One of these responders remains well with improving peak serum cortisol (672 nmol/L) 28 months after stopping all treatments. The other responder showed a gradual reduction in serum cortisol and aldosterone over time, and steroid therapy was recommenced after a 28-week period without glucocorticoid replacement. CONCLUSION: This is the first study to demonstrate that established autoimmune Addison's disease is amenable to a regenerative medicine therapy approach.
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Enfermedad de Addison/tratamiento farmacológico , Enfermedad de Addison/fisiopatología , Glándulas Suprarrenales/efectos de los fármacos , Glándulas Suprarrenales/fisiología , Cosintropina/uso terapéutico , Enfermedad de Addison/metabolismo , Adolescente , Corticoesteroides/sangre , Corticoesteroides/orina , Adulto , Anciano , Femenino , Humanos , Bombas de Infusión , Masculino , Persona de Mediana Edad , Inducción de Remisión , Resultado del Tratamiento , Adulto JovenRESUMEN
Apparent mineralocorticoid excess syndrome (AME) is an autosomal recessive genetic disorder caused by a deficiency in the enzyme 11ß-hydroxysteroid dehydrogenase type 2 (11ß-HSD). We report a 36-year-old male who was hypertensive from birth and was diagnosed with AME at 8 years of age. There was continuous documentation of his hypertension and hypokalemic alkalosis throughout childhood, during which spironolactone and supplemental potassium were administered. At 33 years of age, the patient received a renal transplant, and following this the AME appears to have been cured clinically with remission of his low renin hypertension and hypokalemic alkalosis despite termination of treatment with spironolactone and potassium supplements.
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Electrólitos/sangre , Hipertensión/terapia , Trasplante de Riñón , Síndrome de Exceso Aparente de Mineralocorticoides/complicaciones , Adulto , Niño , Humanos , Hipertensión/complicacionesRESUMEN
CONTEXT: Mutations in the electron donor enzyme P450 oxidoreductase (POR) result in congenital adrenal hyperplasia with apparent combined 17α-hydroxylase/17,20 lyase and 21-hydroxylase deficiencies, also termed P450 oxidoreductase deficiency (PORD). Major clinical features present in PORD are disordered sex development in affected individuals of both sexes, glucocorticoid deficiency, and multiple skeletal malformations. OBJECTIVE: The objective of the study was to establish a noninvasive approach to prenatal diagnosis of PORD including assessment of malformation severity to facilitate optimized prenatal diagnosis and timely treatment. DESIGN: We analyzed 20 pregnancies with children homozygous or compound heterozygous for disease-causing POR mutations and 1 pregnancy with a child carrying a heterozygous POR mutation by recording clinical and biochemical presentations and fetal ultrasound findings. In 4 of the pregnancies (3 homozygous and 1 heterozygous for disease-causing POR mutations), prenatal analysis of steroid metabolite excretion in maternal urine was carried out by gas chromatography/mass spectrometry during gestational weeks 11-23. RESULTS: Pregnancy complications in our cohort included maternal virilization (6 of 20) with onset in the second trimester. Seven pregnant women presented with low unconjugated estriol at prenatal screening (triple or quadruple antenatal screening test). Overt dysmorphic features were noted in 19 of the 20 babies at birth but observed in only 5 by prenatal ultrasound. These 5 had the most severe malformation phenotypes and poor outcome, whereas the other babies showed normal development. Steroid profiling of maternal urine revealed significantly increased steroids of fetal origin, namely the pregnenolone metabolite epiallopregnanediol and the androgen metabolite androsterone, with concomitant low values for estriol. Diagnostic steroid ratios conclusively indicated PORD as early as gestational week 12. In the heterozygous pregnancy, steroid ratios were only slightly elevated and estriol excretion was normal. CONCLUSION: Prenatal diagnosis in PORD is readily established via urinary steroid metabolite analysis of maternal urine. Visible malformations at prenatal ultrasound predict a severe malformation phenotype.
Asunto(s)
Anomalías Múltiples/diagnóstico por imagen , Hiperplasia Suprarrenal Congénita , Tamizaje Masivo/métodos , Diagnóstico Prenatal/métodos , Esteroide 17-alfa-Hidroxilasa/orina , Esteroide 21-Hidroxilasa/orina , Anomalías Múltiples/genética , Hiperplasia Suprarrenal Congénita/diagnóstico , Hiperplasia Suprarrenal Congénita/genética , Hiperplasia Suprarrenal Congénita/orina , Androsterona/orina , Estriol/orina , Femenino , Heterocigoto , Homocigoto , Humanos , Masculino , Fenotipo , Valor Predictivo de las Pruebas , Embarazo , Segundo Trimestre del Embarazo/genética , Pregnanodiol/orina , Radiografía , Esteroide 17-alfa-Hidroxilasa/genética , Esteroide 21-Hidroxilasa/genética , Ultrasonografía Prenatal , Virilismo/diagnóstico , Virilismo/genéticaRESUMEN
CONTEXT: Mitotane [1-(2-chlorophenyl)-1-(4-chlorophenyl)-2,2-dichloroethane] is the first-line treatment for metastatic adrenocortical carcinoma (ACC) and is also regularly used in the adjuvant setting after presumed complete removal of the primary tumor. Mitotane is considered an adrenolytic substance, but there is limited information on distinct effects on steroidogenesis. However, adrenal insufficiency and male hypogonadism are widely recognized side effects of mitotane treatment. OBJECTIVE: Our objective was to define the impact of mitotane treatment on in vivo steroidogenesis in patients with ACC. SETTING AND DESIGN: At seven European specialist referral centers for adrenal tumors, we analyzed 24-h urine samples (n = 127) collected from patients with ACC before and during mitotane therapy in the adjuvant setting (n = 23) or for metastatic ACC (n = 104). Urinary steroid metabolite excretion was profiled by gas chromatography/mass spectrometry in comparison with healthy controls (n = 88). RESULTS: We found a sharp increase in the excretion of 6ß-hydroxycortisol over cortisol (P < 0.001), indicative of a strong induction of the major drug-metabolizing enzyme cytochrome P450 3A4. The contribution of 6ß-hydroxycortisol to total glucocorticoid metabolites increased from 2% (median, interquartile range 1-4%) to 56% (39-71%) during mitotane treatment. Furthermore, we documented strong inhibition of systemic 5α-reductase activity, indicated by a significant decrease in 5α-reduced steroids, including 5α-tetrahydrocortisol, 5α-tetrahydrocorticosterone, and androsterone (all P < 0.001). The degree of inhibition was similar to that in patients with inactivating 5α-reductase type 2 mutations (n = 23) and patients receiving finasteride (n = 5), but cluster analysis of steroid data revealed a pattern of inhibition distinct from these two groups. Longitudinal data showed rapid onset and long-lasting duration of the observed effects. CONCLUSIONS: Cytochrome P450 3A4 induction by mitotane results in rapid inactivation of more than 50% of administered hydrocortisone, explaining the need for doubling hydrocortisone replacement in mitotane-treated patients. Strong inhibition of 5α-reductase activity is in line with the clinical observation of relative inefficiency of testosterone replacement in mitotane-treated men, calling for replacement by 5α-reduced androgens.
Asunto(s)
3-Oxo-5-alfa-Esteroide 4-Deshidrogenasa/metabolismo , Neoplasias de la Corteza Suprarrenal/tratamiento farmacológico , Carcinoma Corticosuprarrenal/tratamiento farmacológico , Citocromo P-450 CYP3A/metabolismo , Mitotano/efectos adversos , Mitotano/uso terapéutico , Adolescente , Neoplasias de la Corteza Suprarrenal/metabolismo , Neoplasias de la Corteza Suprarrenal/orina , Carcinoma Corticosuprarrenal/metabolismo , Carcinoma Corticosuprarrenal/orina , Adulto , Anciano , Anciano de 80 o más Años , Andrógenos/administración & dosificación , Andrógenos/uso terapéutico , Antineoplásicos Hormonales/uso terapéutico , Regulación hacia Abajo/efectos de los fármacos , Activación Enzimática/efectos de los fármacos , Femenino , Glucocorticoides/administración & dosificación , Glucocorticoides/uso terapéutico , Necesidades y Demandas de Servicios de Salud , Terapia de Reemplazo de Hormonas/métodos , Terapia de Reemplazo de Hormonas/estadística & datos numéricos , Humanos , Masculino , Persona de Mediana Edad , Medicina de Precisión/métodos , Regulación hacia Arriba/efectos de los fármacos , Adulto JovenRESUMEN
CONTEXT: P450 oxidoreductase deficiency (PORD) is a unique congenital adrenal hyperplasia variant that manifests with glucocorticoid deficiency, disordered sex development (DSD), and skeletal malformations. No comprehensive data on genotype-phenotype correlations in Caucasian patients are available. OBJECTIVE: The objective of the study was to establish genotype-phenotype correlations in a large PORD cohort. DESIGN: The design of the study was the clinical, biochemical, and genetic assessment including multiplex ligation-dependent probe amplification (MLPA) in 30 PORD patients from 11 countries. RESULTS: We identified 23 P450 oxidoreductase (POR) mutations (14 novel) including an exonic deletion and a partial duplication detected by MLPA. Only 22% of unrelated patients carried homozygous POR mutations. p.A287P was the most common mutation (43% of unrelated alleles); no other hot spot was identified. Urinary steroid profiling showed characteristic PORD metabolomes with variable impairment of 17α-hydroxylase and 21-hydroxylase. Short cosyntropin testing revealed adrenal insufficiency in 89%. DSD was present in 15 of 18 46,XX and seven of 12 46,XY individuals. Homozygosity for p.A287P was invariably associated with 46,XX DSD but normal genitalia in 46,XY individuals. The majority of patients with mild to moderate skeletal malformations, assessed by a novel scoring system, were compound heterozygous for missense mutations, whereas nearly all patients with severe malformations carried a major loss-of-function defect on one of the affected alleles. CONCLUSIONS: We report clinical, biochemical, and genetic findings in a large PORD cohort and show that MLPA is a useful addition to POR mutation analysis. Homozygosity for the most frequent mutation in Caucasians, p.A287P, allows for prediction of genital phenotype and moderate malformations. Adrenal insufficiency is frequent, easily overlooked, but readily detected by cosyntropin testing.
Asunto(s)
Hiperplasia Suprarrenal Congénita/genética , NADPH-Ferrihemoproteína Reductasa/genética , Adolescente , Hiperplasia Suprarrenal Congénita/orina , Insuficiencia Suprarrenal/genética , Insuficiencia Suprarrenal/metabolismo , Insuficiencia Suprarrenal/orina , Adulto , Niño , Estudios de Cohortes , Análisis Mutacional de ADN/métodos , Trastornos del Desarrollo Sexual , Femenino , Estudios de Asociación Genética , Genitales/anomalías , Hormonas Esteroides Gonadales/orina , Humanos , Masculino , Metaboloma , Modelos Biológicos , Modelos Moleculares , Reacción en Cadena de la Polimerasa Multiplex/métodos , NADPH-Ferrihemoproteína Reductasa/deficiencia , NADPH-Ferrihemoproteína Reductasa/fisiología , Adulto JovenRESUMEN
CONTEXT: Abiraterone acetate is a small-molecule cytochrome P450 17A1 (CYP17A1) inhibitor that is active in castration-resistant prostate cancer. OBJECTIVE: Our objective was to determine the impact of abiraterone with and without dexamethasone treatment on in vivo steroidogenesis. DESIGN AND METHODS: We treated 42 castrate, castration-resistant prostate cancer patients with continuous, daily abiraterone acetate and prospectively collected blood and urine before and during abiraterone treatment and after addition of dexamethasone 0.5 mg daily. RESULTS: Treatment with single-agent abiraterone acetate was associated with accumulation of steroids with mineralocorticoid properties upstream of CYP17A1. This resulted in side effects, including hypertension, hypokalemia, and fluid overload, in 38 of 42 patients that were generally treated effectively with eplerenone. Importantly, serum and urinary androgens were suppressed by more than 90% from baseline. Urinary metabolites of 17-hydroxypregnenolone and 17-hydroxyprogesterone downstream of 17α-hydroxylase remained unchanged. However, 3α5α-17-hydroxypregnanolone, which can be converted via the backdoor pathway toward 5α-dihydrotestosterone, increased significantly and correlated with levels of the major 5α-dihydrotestosterone metabolite androsterone. In contrast, urinary metabolites of 11-deoxycortisol and active glucocorticoids declined significantly. Addition of dexamethasone to abiraterone acetate significantly suppressed ACTH and endogenous steroids, including 3α5α-17-hydroxypregnanolone. CONCLUSION: CYP17A1 inhibition with abiraterone acetate is characterized by significant suppression of androgen and cortisol synthesis. The latter is associated with a rise in ACTH that causes raised mineralocorticoids, leading to side effects and incomplete 17α-hydroxylase inhibition. Concomitant inhibition of 17,20-lyase results in diversion of 17-hydroxyprogesterone metabolites toward androgen synthesis via the backdoor pathway. Addition of dexamethasone reverses toxicity and could further suppress androgens by preventing a rise in substrates of backdoor androgen synthesis.
Asunto(s)
Androstenoles/administración & dosificación , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Carcinoma/tratamiento farmacológico , Glucocorticoides/administración & dosificación , Neoplasias de la Próstata/tratamiento farmacológico , Esteroide 17-alfa-Hidroxilasa/antagonistas & inhibidores , Androstenos , Androstenoles/efectos adversos , Androstenoles/farmacología , Antineoplásicos Hormonales/administración & dosificación , Antineoplásicos Hormonales/efectos adversos , Antineoplásicos Hormonales/farmacología , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Biomarcadores/análisis , Biomarcadores/metabolismo , Biomarcadores Farmacológicos/análisis , Biomarcadores Farmacológicos/metabolismo , Carcinoma/metabolismo , Carcinoma/cirugía , Quimioterapia Adyuvante , Progresión de la Enfermedad , Inhibidores Enzimáticos/administración & dosificación , Inhibidores Enzimáticos/efectos adversos , Inhibidores Enzimáticos/farmacología , Glucocorticoides/efectos adversos , Humanos , Masculino , Modelos Biológicos , Orquiectomía , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/cirugía , Esteroide 17-alfa-Hidroxilasa/metabolismo , Resultado del TratamientoRESUMEN
CONTEXT: Adrenal tumors have a prevalence of around 2% in the general population. Adrenocortical carcinoma (ACC) is rare but accounts for 2-11% of incidentally discovered adrenal masses. Differentiating ACC from adrenocortical adenoma (ACA) represents a diagnostic challenge in patients with adrenal incidentalomas, with tumor size, imaging, and even histology all providing unsatisfactory predictive values. OBJECTIVE: Here we developed a novel steroid metabolomic approach, mass spectrometry-based steroid profiling followed by machine learning analysis, and examined its diagnostic value for the detection of adrenal malignancy. DESIGN: Quantification of 32 distinct adrenal derived steroids was carried out by gas chromatography/mass spectrometry in 24-h urine samples from 102 ACA patients (age range 19-84 yr) and 45 ACC patients (20-80 yr). Underlying diagnosis was ascertained by histology and metastasis in ACC and by clinical follow-up [median duration 52 (range 26-201) months] without evidence of metastasis in ACA. Steroid excretion data were subjected to generalized matrix learning vector quantization (GMLVQ) to identify the most discriminative steroids. RESULTS: Steroid profiling revealed a pattern of predominantly immature, early-stage steroidogenesis in ACC. GMLVQ analysis identified a subset of nine steroids that performed best in differentiating ACA from ACC. Receiver-operating characteristics analysis of GMLVQ results demonstrated sensitivity = specificity = 90% (area under the curve = 0.97) employing all 32 steroids and sensitivity = specificity = 88% (area under the curve = 0.96) when using only the nine most differentiating markers. CONCLUSIONS: Urine steroid metabolomics is a novel, highly sensitive, and specific biomarker tool for discriminating benign from malignant adrenal tumors, with obvious promise for the diagnostic work-up of patients with adrenal incidentalomas.