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1.
Anal Bioanal Chem ; 414(23): 6841-6853, 2022 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-35915250

RESUMO

The objective of the current research was to develop a liquid chromatography-MSn (LC-MSn) methodology for the determination of free cortisol and its 15 endogenous metabolites (6ß-hydroxycortisol, 20α-dihydrocortisol, 20α-dihydrocortisone, 20-ß-dihydrocortisol, 20ß-dihydrocortisone, prednisolone, cortisone, α-cortolone, ß-cortolone, allotetrahydrocortisol, 5α-dihydrocortisol, tetrahydrocortisol, allotetrahydrocortisone, 5ß-dihydrocortisol, tetrahydrocortisone) in human urine. Due to its optimal performance, a linear ion trap operating in ESI negative ion mode was chosen for the spectrometric analysis, performing MS3 and MS4 experiments. The method was validated for limit of detection (LOD) and limit of quantification (LOQ) (0.01 ng mL-1 and 0.05 ng mL-1, for all compounds, respectively), intra- and inter-day precision (CV = 1.4-9.2% and CV = 3.6-10.4%, respectively), intra- and inter-day accuracy (95-110%), extraction recovery (65-95%), linearity (R2 > 0.995), and matrix effect that was absent for all molecules. Additionally, for each compound, the percentage of glucuronated conjugates was estimated. The method was successfully applied to the urine (2 mL) of 50 healthy subjects (25 males, 25 females). It was also successfully employed on urine samples of two patients with Cushing syndrome and one with Addison's disease. This analytical approach could be more appropriate than commonly used determination of urinary free cortisol collected in 24-h urine. The possibility of considering the differences and relationship between cortisol and its metabolites allows analytical problems related to quantitative analysis of cortisol alone to be overcome. Furthermore, the developed method has been demonstrated as efficient for antidoping control regarding the potential abuse of corticosteroids, which could interfere with the cortisol metabolism, due to negative feedback on the hypothalamus-hypophysis-adrenal axis. Lastly, this method was found to be suitable for the follow-up of prednisolone that was particularly important considering its pseudo-endogenous origin and correlation with cortisol metabolism.


Assuntos
Hidrocortisona , Espectrometria de Massas em Tandem , Cromatografia Líquida/métodos , Feminino , Humanos , Masculino , Prednisolona , Espectrometria de Massas em Tandem/métodos , Tetra-Hidrocortisona/química , Tetra-Hidrocortisona/urina
2.
J Steroid Biochem Mol Biol ; 210: 105856, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33647522

RESUMO

To evaluate the diagnostic value of serum levels of adrenal steroids for diagnosing and subtyping Cushing's syndrome. Patients diagnosed with endogenous Cushing's syndrome (34 and 19 patients with adrenal and pituitary Cushing's syndrome, respectively) and healthy controls (n = 34) were consecutively enrolled at Seoul National University from 2016 to 2020. Morning serum samples were collected before and 3 months after treatment. Serum steroids were profiled using liquid chromatography-mass spectrometry. The diagnostic value of each and the combination of steroids were assessed using the area under the curve of receiver operating characteristic (AUROC) and decision tree analysis. Tetrahydrocortisone and 6ß-hydroxycortisol showed the highest AUROC (0.893 and 0.890, respectively) for the diagnosis of endogenous Cushing's syndrome. The decision tree composed of tetrahydrocortisone and 6ß-hydroxycortisol correctly classified 79/87 (90.8 %) subjects. For subtyping into adrenal or pituitary Cushing's syndrome, dehydroepiandrosterone sulfate (DHEA-S) showed the highest AUROC (0.988), which was similar to that of plasma ACTH (0.994, P = 0.458). The decision tree composed of only DHEA-S correctly classified 51/53 (96.2 %) of the Cushing's syndrome subtype. DHEA-S showed a significant linear correlation with the plasma ACTH level, but not with the 24 -h urine free cortisol or dexamethasone suppression test results. All steroids, except allo-tetrahydrocortisol and tetrahydrocortisone, decreased significantly at 3 months post-treatment with similar patterns in both adrenal and pituitary Cushing's syndrome. Serum steroid profiling using a single morning serum sample provides valuable information for diagnosing and subtyping Cushing's syndrome.


Assuntos
Síndrome de Cushing/sangue , Síndrome de Cushing/diagnóstico , Esteroides/sangue , Glândulas Suprarrenais/metabolismo , Adulto , Estudos de Casos e Controles , Síndrome de Cushing/tratamento farmacológico , Síndrome de Cushing/etiologia , Feminino , Humanos , Hidrocortisona/análogos & derivados , Hidrocortisona/sangue , Hidrocortisona/urina , Masculino , Pessoa de Meia-Idade , Hipófise/metabolismo , Curva ROC , Tetra-Hidrocortisona/sangue
3.
J Clin Endocrinol Metab ; 106(3): 814-825, 2021 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-33236103

RESUMO

CONTEXT: Oral once-daily dual-release hydrocortisone (DR-HC) replacement therapy has demonstrated an improved metabolic profile compared to conventional 3-times-daily (TID-HC) therapy among patients with primary adrenal insufficiency. This effect might be related to a more physiological cortisol profile, but also to a modified pattern of cortisol metabolism. OBJECTIVE: This work aimed to study cortisol metabolism during DR-HC and TID-HC. DESIGN: A randomized, 12-week, crossover study was conducted. INTERVENTION AND PARTICIPANTS: DC-HC and same daily dose of TID-HC were administered to patients with primary adrenal insufficiency (n = 50) vs healthy individuals (n = 124) as controls. MAIN OUTCOME MEASURES: Urinary corticosteroid metabolites were measured by gas chromatography/mass spectrometry at 24-hour urinary collections. RESULTS: Total cortisol metabolites decreased during DR-HC compared to TID-HC (P < .001) and reached control values (P = .089). During DR-HC, 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) activity measured by tetrahydrocortisol + 5α-tetrahydrocortisol/tetrahydrocortisone ratio was reduced compared to TID-HC (P < .05), but remained increased vs controls (P < .001). 11ß-HSD2 activity measured by urinary free cortisone/free cortisol ratio was decreased with TID-HC vs controls (P < .01) but normalized with DR-HC (P = .358). 5α- and 5ß-reduced metabolites were decreased with DR-HC compared to TID-HC. Tetrahydrocortisol/5α-tetrahydrocortisol ratio was increased during both treatments, suggesting increased 5ß-reductase activity. CONCLUSIONS: The urinary cortisol metabolome shows striking abnormalities in patients receiving conventional TID-HC replacement therapy, with increased 11ß-HSD1 activity that may account for the unfavorable metabolic phenotype in primary adrenal insufficiency. Its change toward normalization with DR-HC may mediate beneficial metabolic effects. The urinary cortisol metabolome may serve as a tool to assess optimal cortisol replacement therapy.


Assuntos
Doença de Addison , Hidrocortisona/farmacocinética , Esteroides/urina , Doença de Addison/tratamento farmacológico , Doença de Addison/metabolismo , Doença de Addison/urina , Adulto , Idoso , Cortisona/metabolismo , Cortisona/urina , Estudos Cross-Over , Preparações de Ação Retardada/farmacocinética , Preparações de Ação Retardada/uso terapêutico , Europa (Continente) , Feminino , Humanos , Hidrocortisona/uso terapêutico , Hidrocortisona/urina , Masculino , Metaboloma/efeitos dos fármacos , Pessoa de Meia-Idade , Pregnanos/metabolismo , Pregnanos/urina , Esteroides/metabolismo , Tetra-Hidrocortisol/metabolismo , Tetra-Hidrocortisol/urina , Tetra-Hidrocortisona/metabolismo , Tetra-Hidrocortisona/urina , Urinálise
4.
Clin Endocrinol (Oxf) ; 94(2): 150-157, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33030273

RESUMO

OBJECTIVE: Increasing magnesium intake might reduce the risk of cardiovascular disease (CVD). Whether potential effects on cortisol contribute to these beneficial effects on cardiovascular health remains unclear. We therefore studied effects of long-term oral magnesium supplementation on glucocorticoid metabolism, specifically on the excretion of urinary cortisol, cortisone and their metabolites, as well as on the ratios reflecting enzymatic activity of 11ß-hydroxysteroid dehydrogenases (11ß-HSDs) and A-ring reductases. DESIGN: A post-hoc analysis of a randomized trial with allocation to a magnesium supplement (350 mg/day) or a placebo for 24-week. PATIENTS: Forty-nine overweight men and women, aged between 45 and 70 years. MEASUREMENTS: Cortisol, cortisone and their metabolites (tetrahydrocortisol [THF], allo-tetrahydrocortisol [allo-THF] and tetrahydrocortisone [THE]) were measured in 24-h urine samples. Enzymatic activities of 11ß-HSD overall and of 11ß-HSD type 2 were estimated as the urinary (THF + allo-THF [THFs])/THE and cortisol/cortisone ratios, respectively. A-ring reductase activity was assessed by ratios of THF/allo-THF, allo-THF/cortisol, THF/cortisol and THE/cortisone. RESULTS: After 24-week, urinary cortisol excretion was decreased in the magnesium group as compared with the placebo group (-32 nmol/24-h, 95% CI: -59; -5 nmol/24-h, p = .021). Ratios of THFs/THE and cortisol/cortisone were decreased following magnesium supplementation by 0.09 (95% CI: 0.02; 0.17, p = .018) and 0.10 (95% CI: 0.03; 0.17, p = .005), respectively. No effects were observed on A-ring reductase activity. CONCLUSIONS: We observed a beneficial effect of magnesium supplementation towards a lower 24-h urinary cortisol excretion together with an increased activity of 11ß-HSD type 2. Our findings may provide another potential mechanism by which increased magnesium intake lowers CVD risk (ClinicalTrials.gov identifier: NCT02235805).


Assuntos
Cortisona , Glucocorticoides , Idoso , Suplementos Nutricionais , Feminino , Humanos , Hidrocortisona , Magnésio , Masculino , Pessoa de Meia-Idade , Tetra-Hidrocortisona
5.
ACS Nano ; 14(2): 2542-2552, 2020 02 25.
Artigo em Inglês | MEDLINE | ID: mdl-32049493

RESUMO

Successful translation of laboratory-based surface-enhanced Raman scattering (SERS) platforms to clinical applications requires multiplex and ultratrace detection of small biomarker molecules from a complex biofluid. However, these biomarker molecules generally exhibit low Raman scattering cross sections and do not possess specific affinity to plasmonic nanoparticle surfaces, significantly increasing the challenge of detecting them at low concentrations. Herein, we demonstrate a "confine-and-capture" approach for multiplex detection of two families of urine metabolites correlated with miscarriage risks, 5ß-pregnane-3α,20α-diol-3α-glucuronide and tetrahydrocortisone. To enhance SERS signals by 1012-fold, we use specific nanoscale surface chemistry for targeted metabolite capture from a complex urine matrix prior to confining them on a superhydrophobic SERS platform. We then apply chemometrics, including principal component analysis and partial least-squares regression, to convert molecular fingerprint information into quantifiable readouts. The whole screening procedure requires only 30 min, including urine pretreatment, sample drying on the SERS platform, SERS measurements, and chemometric analyses. These readouts correlate well with the pregnancy outcomes in a case-control study of 40 patients presenting threatened miscarriage symptoms.


Assuntos
Pregnanodiol/urina , Tetra-Hidrocortisona/urina , Calibragem , Teoria da Densidade Funcional , Feminino , Humanos , Estrutura Molecular , Tamanho da Partícula , Gravidez , Pregnanodiol/análogos & derivados , Pregnanodiol/metabolismo , Análise Espectral Raman , Propriedades de Superfície , Tetra-Hidrocortisona/metabolismo , Fatores de Tempo
6.
Clin Transplant ; 34(4): e13824, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32052523

RESUMO

BACKGROUND: Chronic corticosteroid treatment suppresses HPA-axis activity and might alter activity of 11ß hydroxysteroid dehydrogenases (11ß-HSD). We aimed to investigate whether the endogenous glucocorticoid production and 11ß-HSD activities are altered in prednisolone-treated renal transplant recipients (RTR) compared with healthy controls and whether this has implications for long-term survival in RTR. METHODS: In a longitudinal cohort of 693 stable RTR and 275 healthy controls, 24-hour urinary cortisol, cortisone, tetrahydrocorisol (THF), allotetrahydrocortisol (alloTHF), and tetrahydrocortisone (THE) were measured using liquid chromatography tandem-mass spectrometry. Twenty-four-hour urinary excretion of cortisol and metabolites were used as measures of endogenous glucocorticoid production; (THF + alloTHF)/THE and cortisol/cortisone ratios were used as measures of 11ß-HSD activity. RESULTS: Urinary cortisol and metabolite excretion were significantly lower in RTR compared with healthy controls (P < .001), whereas (THF + alloTHF)/THE and cortisol/cortisone ratios were significantly higher (P < .001 and P = .002). Lower total urinary metabolite excretion and higher urinary (THF + alloTHF)/THE ratios were associated with increased risk of mortality, independent of age, sex, estimated glomerular filtration rate, C-reactive protein, body surface area, and daily prednisolone dose, respectively. CONCLUSIONS: Endogenous glucocorticoid production and 11ß-HSD activities are altered in prednisolone-treated RTR. Decreased total urinary endogenous glucocorticoid metabolite excretion and increased urinary (THF + alloTHF)/THE ratios are associated with increased risk of mortality.


Assuntos
Cortisona , Transplante de Rim , Glucocorticoides/uso terapêutico , Humanos , Prednisolona/uso terapêutico , Tetra-Hidrocortisona
7.
J Pharm Biomed Anal ; 140: 174-181, 2017 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-28359965

RESUMO

Cortisol (F) and cortisone (E) are metabolized to A-ring reduced metabolites in the reactions catalyzed by 5α- and 5ß-reductase. 5α-tetrahydrocortisol (alloTHF) and 5ß-tetrahydrocortisol (THF) are produced from F. The metabolism of E takes place in analogy to form alloTHE and THE. Up to now, the analysis of endogenous glucocorticoids did not consider alloTHE, limiting the metabolism of E to THE only. Nevertheless, such simplification can generate inaccuracy in the assessment of the function of enzymes crucial for glucocorticoids metabolism: 11ß-hydroxysteroid dehydrogenase type 1 and type 2 (11ß-HSD1 and 11ß-HSD2), as well as 5α- and 5ß-reductase. The paper presents the new LC-MS/MS method for the simultaneous analysis of F and E with their tetrahydro- (THF and THE) and allo-tetrahydrometabolites (alloTHF and alloTHE) in urine. The method was fully validated and allows determining both the unconjugated and total concentrations of urinary glucocorticoids. The method meets the EMA's recommendations and was proved to be useful in the analysis of clinical samples. The LLOQ of 1ng/mL allows the determination of free urinary F, E, THF and THE, but not alloTHF and alloTHE, in samples obtained from pregnant women. The range of concentrations is wide enough for the analysis of total levels of F, E, THF, alloTHF, THE and alloTHE. The undisputed advantage of the method, distinguishing it among others, is the ability to determine F and E and their both 5α- and 5ß-metabolites. Taking alloTHE into consideration enables the thorough analysis of the glucocorticoid equilibrium in human.


Assuntos
Espectrometria de Massas em Tandem , Cromatografia Líquida , Cortisona , Feminino , Humanos , Hidrocortisona , Gravidez , Tetra-Hidrocortisol , Tetra-Hidrocortisona
8.
Br J Pharmacol ; 174(5): 396-408, 2017 03.
Artigo em Inglês | MEDLINE | ID: mdl-28012176

RESUMO

BACKGROUND AND PURPOSE: Reducing glucocorticoid exposure in the brain via intracellular inhibition of the cortisol-regenerating enzyme 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) has emerged as a therapeutic strategy to treat cognitive impairment in early Alzheimer's disease (AD). We sought to discover novel, brain-penetrant 11ß-HSD1 inhibitors as potential medicines for the treatment of AD. EXPERIMENTAL APPROACH: Medicinal chemistry optimization of a series of amido-thiophene analogues was performed to identify potent and selective 11ß-HSD1 inhibitors with optimized oral pharmacokinetics able to access the brain. Single and multiple ascending dose studies were conducted in healthy human subjects to determine the safety, pharmacokinetic and pharmacodynamic characteristics of the candidate compound. RESULTS: UE2343 was identified as a potent, orally bioavailable, brain-penetrant 11ß-HSD1 inhibitor and selected for clinical studies. No major safety issues occurred in human subjects. Plasma adrenocorticotropic hormone was elevated (a marker of systemic enzyme inhibition) at doses of 10 mg and above, but plasma cortisol levels were unchanged. Following multiple doses of UE2343, plasma levels were approximately dose proportional and the terminal t1/2 ranged from 10 to 14 h. The urinary tetrahydrocortisols/tetrahydrocortisone ratio was reduced at doses of 10 mg and above, indicating maximal 11ß-HSD1 inhibition in the liver. Concentrations of UE2343 in the CSF were 33% of free plasma levels, and the peak concentration in CSF was ninefold greater than the UE2343 IC50 . CONCLUSIONS AND IMPLICATIONS: UE2343 is safe, well tolerated and reaches the brain at concentrations predicted to inhibit 11ß-HSD1. UE2343 is therefore a suitable candidate to test the hypothesis that 11ß-HSD1 inhibition in brain improves memory in patients with AD.


Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 1/antagonistas & inibidores , Encéfalo/metabolismo , Inibidores Enzimáticos/administração & dosagem , Tiofenos/administração & dosagem , Tropanos/administração & dosagem , Adolescente , Adulto , Animais , Cães , Relação Dose-Resposta a Droga , Método Duplo-Cego , Inibidores Enzimáticos/efeitos adversos , Inibidores Enzimáticos/farmacocinética , Feminino , Meia-Vida , Humanos , Hidrocortisona/sangue , Concentração Inibidora 50 , Masculino , Pessoa de Meia-Idade , Ratos , Ratos Sprague-Dawley , Tetra-Hidrocortisol/urina , Tetra-Hidrocortisona/urina , Tiofenos/efeitos adversos , Tiofenos/farmacocinética , Distribuição Tecidual , Tropanos/efeitos adversos , Tropanos/farmacocinética , Adulto Jovem
9.
Artigo em Inglês | MEDLINE | ID: mdl-27310211

RESUMO

The effects of long-term administration of low doses of dexamethasone (DX) and prednisolone (PL) on the metabolism of endogenous corticosteroids were investigated in veal calves. In addition to cortisol (F) and cortisone (E), whose interconversion is regulated by 11ß-hydroxysteroid dehydrogenases (11ßHSDs), special attention was paid to tetrahydrocortisol (THF), allo-tetrahydrocortisol (aTHF), tetrahydrocortisone (THE) and allo-tetrahydrocortisone (aTHE), which are produced from F and E by catalytic activity of 5α and 5ß-reductases. A specifically developed HPLC-ESI-MS/MS method achieved the complete chromatographic separation of two pairs of diastereoisomers (THF/aTHF and THE/aTHE), which, with appropriate mass fragmentation patterns, provided an unambiguous conformation. The method was linear (r(2) > 0.9905; 0.5-25 ng ml(-1)), with LOQQ of 0.5 ng ml(-1). Recoveries were in range 75-114%, while matrix effects were minimal. The experimental study was carried out on three groups of male Friesian veal calves: group PL (n = 6, PL acetate 15 mg day(-1) p.o. for 31 days); group DX (n = 5, 5 mg of estradiol (E2) i.m., weekly, and 0.4 mg day(-1) of DX p.o. for 31 days) and a control group (n = 8). Urine was collected before, during (twice) and at the end of treatment. During PL administration, the tetrahydro-metabolite levels decreased gradually and remained low after the suspension of treatment. DX reduced urinary THF that persisted after the treatment, while THE levels decreased during the experiment, but rebounded substantially after the DX was withdrawn. Both DX and PL significantly interfered with the production of F and E, leading to their complete depletion. Taken together, the results demonstrate the influence of DX and PL administration on 11ßHSD activity and their impact on dysfunction of the 5-reductase pathway. In conclusion, profiling tetrahydro-metabolites of F and E might serve as an alternative, indirect but reliable, non-invasive procedure for assessing the impact of synthetic glucocorticosteroids administration.


Assuntos
Cortisona/urina , Dexametasona/urina , Hidrocortisona/urina , Prednisolona/urina , Tetra-Hidrocortisol/análogos & derivados , Tetra-Hidrocortisona/urina , 11-beta-Hidroxiesteroide Desidrogenases/antagonistas & inibidores , 11-beta-Hidroxiesteroide Desidrogenases/urina , Animais , Biomarcadores/urina , Biotransformação , Bovinos , Cromatografia Líquida de Alta Pressão , Dexametasona/farmacologia , Masculino , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/antagonistas & inibidores , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/urina , Prednisolona/farmacologia , Espectrometria de Massas por Ionização por Electrospray , Estereoisomerismo , Espectrometria de Massas em Tandem , Tetra-Hidrocortisol/urina
10.
J Steroid Biochem Mol Biol ; 156: 10-6, 2016 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-26493852

RESUMO

One major issue of newborn screening programs for 21-hydroxylase deficiency (21OHD) is the high rate of false-positive results, especially in preterm neonates. Urinary steroid metabolite analysis using gas chromatography-mass spectrometry (GC-MS) is suitable as a confirmatory diagnostic tool. The objective of this study was to analyze retrospectively diagnostic metabolite ratios in neonates and infants with and without 21OHD using GC-MS with emphasis on glucocorticoid metabolism, and to develop reference values for the steroid metabolite ratios for the diagnosis of 21OHD. We retrospectively analyzed urinary steroid hormone metabolites determined by GC-MS of 95 untreated neonates and infants with 21OHD (1-148 days), and 261 neonates and infants (100 preterms) without 21OHD (0-217 days). Metabolites of 17α-hydroxyprogesterone showed specificities below 98%, whereas the 21-deoxycortisol metabolite pregnanetriolone clearly separated 21OHD from non-21OHD subjects. The best diagnostic ratio for 21OHD was pregnanetriolone to 6α-hydroxy-tetrahydrocortisone. The lowest value of this ratio in the 21OHD group (0.47) was at least eight times higher than the highest values in the non-21OHD group (0.055). We have given appropriate reference values for steroid metabolite ratios in the largest 21OHD cohort so far described. Consideration of glucocorticoid metabolism, especially the use of typical neonatal 6α-hydroxylates metabolites, leads to improvement of diagnostic metabolite ratios.


Assuntos
Hiperplasia Suprarrenal Congênita/diagnóstico , Hiperplasia Suprarrenal Congênita/urina , Cromatografia Gasosa-Espectrometria de Massas/métodos , Metabolômica/métodos , Hiperplasia Suprarrenal Congênita/metabolismo , Estudos de Coortes , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Pregnanotriol/análogos & derivados , Pregnanotriol/metabolismo , Pregnanotriol/urina , Valores de Referência , Esteroides/metabolismo , Esteroides/urina , Tetra-Hidrocortisona/análogos & derivados , Tetra-Hidrocortisona/metabolismo , Tetra-Hidrocortisona/urina
11.
Endocrine ; 52(1): 139-47, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26349936

RESUMO

Low-grade metabolic acidosis (LGMA), as induced by high dietary acid load or sodium chloride (NaCl) intake, has been shown to increase bone and protein catabolism. Underlying mechanisms are not fully understood, but from clinical metabolic acidosis interactions of acid-base balance with glucocorticoid (GC) metabolism are known. We aimed to investigate GC activity/metabolism under alkaline supplementation and NaCl-induced LGMA. Eight young, healthy, normal-weight men participated in two crossover designed interventional studies. In Study A, two 10-day high NaCl diet (32 g/d) periods were conducted, one supplemented with 90 mmol KHCO3/day. In Study B, participants received a high and a low NaCl diet (31 vs. 3 g/day), each for 14 days. During low NaCl, the diet was moderately acidified by replacement of a bicarbonate-rich mineral water (consumed during high NaCl) with a non-alkalizing drinking water. In repeatedly collected 24-h urine samples, potentially bioactive-free GCs (urinary-free cortisol + free cortisone) were analyzed, as well as tetrahydrocortisol (THF), 5α-THF, and tetrahydrocortisone (THE). With supplementation of 90 mmol KHCO3, the marker of total adrenal GC secretion (THF + 5α-THF + THE) dropped (p = 0.047) and potentially bioactive-free GCs were reduced (p = 0.003). In Study B, however, GC secretion and potentially bioactive-free GCs did not exhibit the expected fall with NaCl-reduction as net acid excretion was raised by 30 mEq/d. Diet-induced acidification/alkalization affects GC activity and metabolism, which in case of long-term ingestion of habitually acidifying western diets may constitute an independent risk factor for bone degradation and cardiometabolic diseases.


Assuntos
Acidose/induzido quimicamente , Acidose/metabolismo , Álcalis/farmacologia , Glucocorticoides/metabolismo , Cloreto de Sódio , Equilíbrio Ácido-Base/efeitos dos fármacos , Adulto , Bicarbonatos/farmacologia , Cortisona/urina , Estudos Cross-Over , Dieta , Água Potável , Glucocorticoides/urina , Humanos , Hidrocortisona/urina , Masculino , Compostos de Potássio/farmacologia , Tetra-Hidrocortisol/urina , Tetra-Hidrocortisona/metabolismo , Adulto Jovem
12.
J Pharm Biomed Anal ; 107: 450-5, 2015 Mar 25.
Artigo em Inglês | MEDLINE | ID: mdl-25668797

RESUMO

Chronic stress as well as major depressive disorders is associated with cortisol metabolism. Two enzymes modulate cortisol (F) and cortisone (E) interconversion: 11ß-hydroxysteroid dehydrogenase type 1 and type 2 (11ß-HSD1 and 11ß-HSD2). Furthermore, F and E were inactivated by 5α and 5ß reductases to their tetrahydro-metabolites: tetrahydrocortisol (THF), allo-tetrahydrocortisol (5α-THF) and tetrahydrocortisone (THE). To better understand depression a LC-MS/MS method for simultaneous determination of F, E THF, 5α-THF and THE in human urine has been developed and validated. The quantification range was 0.1-160 ng mL(-1) for F and E, and 0.2-160 ng mL(-1) for the tetrahydro-metabolites, with >86.1% recovery for all analytes. The nocturnal urine concentrations of F, E and tetrahydro-metabolites in 12 apparently healthy male adult volunteers and 12 drug-free male patients (age range, 20-50 years) with a diagnosis of depression were analyzed. A series of significant changes in glucocorticoid metabolism can be detected: F/E ratios and (THF+5α-THF)/THE ratios as well as F and THF concentrations were significantly higher in depression patients than in healthy subjects (p<0.05); 5α-THF/F ratios, 5α-THF/THF ratios as well as 5α-THF concentrations were significantly lower in depression patients (p<0.05). The results pointed to the decreased 11ß-HSD2 activity and a dysfunction in the 5α-reductase pathway in depressed patients. This method allows the assessment of 11ß-HSD1/2 and 5α/ß-reductase activities in a single analytical run providing an innovative tool to explain the potential etiology of depression.


Assuntos
Cortisona/química , Cortisona/urina , Transtorno Depressivo Maior/urina , Hidrocortisona/química , Hidrocortisona/urina , Tetra-Hidrocortisona/química , Tetra-Hidrocortisona/urina , 11-beta-Hidroxiesteroide Desidrogenase Tipo 1/metabolismo , 11-beta-Hidroxiesteroide Desidrogenase Tipo 2/metabolismo , Adulto , Estudos de Casos e Controles , Colestenona 5 alfa-Redutase/metabolismo , Cromatografia Líquida/métodos , Cortisona/metabolismo , Transtorno Depressivo Maior/metabolismo , Glucocorticoides/química , Glucocorticoides/metabolismo , Glucocorticoides/urina , Humanos , Hidrocortisona/metabolismo , Masculino , Pessoa de Meia-Idade , Espectrometria de Massas em Tandem/métodos , Tetra-Hidrocortisol/análogos & derivados , Tetra-Hidrocortisol/química , Tetra-Hidrocortisol/metabolismo , Tetra-Hidrocortisol/urina , Tetra-Hidrocortisona/metabolismo , Adulto Jovem
13.
Zhonghua Yu Fang Yi Xue Za Zhi ; 48(9): 800-4, 2014 Sep.
Artigo em Chinês | MEDLINE | ID: mdl-25492292

RESUMO

OBJECTIVE: To study the association between phthalate esters (PAEs) metabolites in maternal urine and 11beta-hydroxysteroid dehydrogenase type 2 (11ß-HSD2 ) enzyme activity, explore the possible mechanism of PAEs effect on fetal development. METHODS: All of 33 cases of intrauterine growth retardation (IUGR) newborn were selected by random sampling in 2012. And 33 cases of normal control newborn were enrolled, use high performance liquid chromatography-tandem mass spectrometry method was used to detect 4 kinds of phthalate esters (PAEs) metabolites in maternal urine: mono-n-butyl phthalate ester (MBP), mono (2-ethylhexyl) phthalate (MEHP), mono (2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), mono (2-ethyl-5-oxohexyl) phthalate (MEOHP) and three kinds of cortisol corticosterone metabolites, tetrahydrocortisol (THF), allo-tetrahydrocortisol (allo-THF), tetrahydrocortisone (THE), and analyze the association between phthalate esters (PAEs) metabolites in maternal urine and 11ß-HSD2 enzyme activity. RESULTS: MBP, MEHP, MEHHP, MEOHP metabolites can be detected in 98% (65 cases) , 89% (59 cases), 91% (60 cases), 91% (60 cases) of all 66 maternal urine samples, respectively. The median concentrations of test material in case group were 31.20 ng/ml for MBP, 24.61 ng/ml for MEHHP, 11.72 ng/ml for MEOHP and 48.67 ng/ml for SumDEHP which were significantly higher than those of the control group (were 17.32, 12.03, 5.68 and 28.64 ng/ml); 11ß-HSD2 activity in case group ((THF+allo-THF)/THE = (0.79 ± 0.09) ng/ml) was significantly lower than that of the control group((THF+allo-THF)/THE = (0.58 ± 0.04) ng/ml); PAEs metabolites MBP (ß' = 1.12), MEHHP(ß' = 1.14), MEOHP(ß' = 1.10), SumDEHP(ß' = 1.08) in baby boy mother's urine was reversely correlated to 11ß-HSD2 activity. CONCLUSIONS: PAEs could affect fetal development by inhibit 11ß-HSD2 activity.


Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 2 , Dietilexilftalato/análogos & derivados , Desenvolvimento Fetal , Ácidos Ftálicos , Cromatografia Líquida , Humanos , Recém-Nascido , Masculino , Espectrometria de Massas , Tetra-Hidrocortisol/análogos & derivados , Tetra-Hidrocortisona
15.
Endocrine ; 47(3): 952-8, 2014 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-24532078

RESUMO

Endogenous glucocorticoids (GC) rapidly increase after acute exercise, and the phosphodiesterase's type 5 inhibitor (PDE5i) tadalafil influences this physiological adaptation. No data exist on acute effects of both acute exercise and PDE5i administration on 11ß-hydroxysteroid dehydrogenases (11ß-HSDs)-related GC metabolites. We aimed to investigate the rapid effects of exercise on serum GC metabolites, with and without tadalafil administration. A double blind crossover study was performed in eleven healthy male volunteers. After the volunteers randomly received a short-term administration of placebo or tadalafil (20 mg/die for 2 days), a maximal exercise test to exhaustion on cycle ergometer was performed. Then, after a 2-week washout period, the volunteers were crossed over. Blood samples were collected before starting exercise and at 5 and 30 min of recovery (+5-Rec, +30-Rec). Serum ACTH, corticosterone (Cn), cortisol (F), cortisone (E), tetrahydrocortisol (THF), tetrahydrocortisone (THE), cortols, cortolones and respective ratios were evaluated. Pre-Ex THF was higher after tadalafil. Exercise increased ACTH, Cn, F, E, THE, cortols and cortolones after both placebo and tadalafil, and THF after placebo. The F/E ratio increased at +5-Rec and decreased at +30-Rec after placebo. Compared to placebo, after tadalafil lower ACTH, F and Cn, higher THF/F and THE/E, and not E (at +5-Rec) and F/E modifications were observed. Acute exercise rapidly influences serum GC metabolites concentrations. Tadalafil influences both GC adaptation and 11ß-HSDs activity during acute exercise. Additional researches on the effects of both exercise and PDE5i on tissue-specific 11ß-HSDs activity at rest and during physiological adaptation are warranted.


Assuntos
11-beta-Hidroxiesteroide Desidrogenases/metabolismo , Carbolinas/farmacologia , Exercício Físico/fisiologia , Inibidores da Fosfodiesterase 5/farmacologia , Hormônio Adrenocorticotrópico/sangue , Adulto , Corticosterona/sangue , Cortisona/sangue , Estudos Cross-Over , Método Duplo-Cego , Humanos , Hidrocortisona/sangue , Masculino , Projetos Piloto , Tadalafila , Tetra-Hidrocortisol/sangue , Tetra-Hidrocortisona/sangue , Adulto Jovem
16.
J Psychiatr Res ; 52: 57-62, 2014 May.
Artigo em Inglês | MEDLINE | ID: mdl-24534618

RESUMO

Dysfunction of the hypothalamic-pituitary-adrenal (HPA) axis is suggested as a pathophysiological factor in bipolar disorder and schizophrenia. Increased clearance of cortisol was recently indicated as a component in the HPA axis hyperdrive. The aim of the present study was to test the model of increased cortisol metabolism in a new replication sample separately and combined with a previously published sample of bipolar disorder and schizophrenia. Spot urine was sampled from 212 healthy controls (HC) and 221 patients with a schizophrenia spectrum disorder (SCZ, n = 115) and bipolar disorder (BD, n = 106). Of these, a subsample of 169 HC and 155 patients was included in a previous report. Urinary free cortisol, cortisone and their metabolites were measured, and the activities of 5α-reductase, 5ß-reductase and 11ß-HSD were estimated and analyzed for differences between groups. In the new sample, there was increased enzyme activity in SCZ for 5ß-reductase (p = 0.024 vs HC; p = 0.027 vs BD) and 11ß-HSD2 (p = 0.014 vs HC; p = 0.004 vs BD). In the combined sample, there was increased activity in SCZ for 5α-reductase (p < 0.001 vs HC; p = 0.020 vs BD), 5ß-reductase (p < 0.001 vs HC; p = 0.045 vs BD) and 11ß-HSD2 (p < 0.001 vs HC; p = 0.043 vs BD), and in BD for 5ß-reductase (p = 0.002), 11ß-HSD2 (p = 0.039) and 5α-reductase (trend, p = 0.084) (all vs HC). The findings confirm increased systemic cortisol metabolism in BD and SCZ. This is most consistent in SCZ, with BD taking an intermediate position. The design makes it impossible to determine the direction of the effect. However, the findings merit further study of cortisol metabolism as a possible component in the HPA axis dysfunction and pathophysiology of BD and SCZ.


Assuntos
Transtorno Bipolar/metabolismo , Hidrocortisona/metabolismo , Esquizofrenia/metabolismo , Adolescente , Adulto , Análise de Variância , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Escalas de Graduação Psiquiátrica , Tetra-Hidrocortisol/análogos & derivados , Tetra-Hidrocortisol/metabolismo , Tetra-Hidrocortisona/metabolismo , Adulto Jovem
17.
J Steroid Biochem Mol Biol ; 135: 30-5, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23291109

RESUMO

Interconversion of hormonally active cortisol (F) into the corresponding inactive 11-keto form, cortisone (E), is catalyzed by 11beta-hydroxysteroid dehydrogenases (11ß-HSDs). With a view to estimating in vivo activities of some 11ß-HSD isoforms, the measurement of urinary F and E and their tetrahydro metabolites (tetrahydrocortisol, THF, allotetrahydrocortisol, ATHF, tetrahydrocortisone, THE) has been suggested. The basic knowledge of THF, ATHF and THE levels in farm cattle is limited. Therefore the aim of this study was first to optimize a simple and quick method to determine F and E tetrahydro-metabolites in bovine urine by HPLC-mass spectrometry with electrospray ionization (HPLC-ESI-MS) and then to apply the method to real urine of bovines treated with prednisolone. The samples underwent filtration, deconjugation, solid-phase extraction (SPE) and the relevant analytes were measured by HPLC-ESI-MS. The method described in this paper is simple and efficient, featuring good linearity (up to 0.996) and reproducibility (6.8-12.5%, CV). Especially, good LODs were obtained, from 1.63 to 2.67 ppb, depending on the analyte. The chromatographic conditions were optimized in order to obtain a resolution which would allow to simultaneously measure two diastereoisomers, i.e. THF and ATHF. In our study, ATHF turns out to be below the detection limit, while for 18 samples tested the contents of examinated metabolites were as followed: THF (12.5±4.8 ppb), THE (10.9±5.5 ppb), F (11.6±3.3 ppb) and E (5.0±2.2 ppb). When the method was applied to the subject treated with prednisolone a major increase in the concentration of tetrahydro metabolites was observed before the slaughter, mainly due to stress conditions; prednisolone treatment, most presumably, influenced the 11ß-HSD activity, as indicated by the decrease in the F/E ratio. This work may provide a useful methodological contribution to the future definition of F, E, THF, ATHF and THE urinary baseline values in order to obtain indirect evaluations of HSDs activity in farm cattle and possible applications in screenings for suspected abuse of synthetic corticosteroids in bovines.


Assuntos
Cortisona/metabolismo , Hidrocortisona/metabolismo , Tetra-Hidrocortisol/urina , Tetra-Hidrocortisona/urina , Animais , Bovinos , Cromatografia Líquida de Alta Pressão , Masculino , Espectrometria de Massas por Ionização por Electrospray , Tetra-Hidrocortisol/análogos & derivados , Urinálise
18.
Clin Endocrinol (Oxf) ; 78(5): 700-5, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-22233384

RESUMO

OBJECTIVE: Increased glucocorticoid metabolite excretion and enhanced expression and activity of 11ß-hydroxysteroid dehydrogenase type 1 in adipose tissue are closely correlated with obesity and its detrimental consequences. Weight loss ameliorates the latter. The aim of this study was to explore whether increased glucocorticoid exposure in obesity is improved with substantial weight loss and thus is a consequence rather than a cause of obesity. DESIGN AND PATIENTS: A prospective cohort study in 31 women. MEASUREMENTS: 11ß-HSD type 1 expression and activity, urinary glucocorticoid metabolite excretion, body composition including regional adipose tissue depots and insulin resistance by HOMA-IR before and 2 years after gastric bypass surgery. RESULTS: After weight loss, excretion of cortisol and cortisone metabolites decreased. Both cortisol and cortisone metabolite excretion correlated with central obesity, where the intraabdominal fat depot showed the strongest association. Cortisol metabolites correlated with 11ß-HSD type 1 activity in abdominal subcutaneous adipose tissue. The ratio of cortisol to cortisone metabolites [(5α-tetrahydrocortisol (5αTHF) + tetrahydrocortisol (THF) + α-cortol)/(tetrahydrocortisone (THE) + α-cortolone)] and the ratio of 5α-THF/THF both decreased after stable weight loss, reflecting a downregulation of the net activities of 11ß-HSD type 1 and 5α-reductase. CONCLUSION: Long-term weight loss in women is not only followed by reduced glucocorticoid production, but also favourably decreases the global and tissue-specific activity of the cortisol-activating enzyme 11 ß-HSD type 1, possibly contributing to the health benefits of bariatric surgery.


Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 1/metabolismo , Hidrocortisona/urina , Obesidade/metabolismo , Obesidade/urina , Redução de Peso/fisiologia , Adulto , Estudos de Coortes , Cortisona/urina , Feminino , Humanos , Pessoa de Meia-Idade , Estudos Prospectivos , Tetra-Hidrocortisol/metabolismo , Tetra-Hidrocortisona/metabolismo
19.
J Clin Endocrinol Metab ; 97(10): 3622-9, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22872687

RESUMO

BACKGROUND: The mineralocorticoid receptor is protected from excess of glucocorticoids by conversion of active cortisol to inactive cortisone by enzyme 11ß-hydroxysteroid dehydrogenase type 2 present in the kidney. The metabolites of cortisol and cortisone are excreted in the urine as tetrahydrocortisol (5αTHF+5ßTHF) and tetrahydrocortisone (THE), respectively. HYPOTHESIS: Patients with chronic kidney disease (CKD) and essential hypertension have a functional defect in their ability to convert cortisol to cortisone, thus leading to the activation of mineralocorticoid receptor. OBJECTIVE: The objective of the investigation was to study the ratio of urinary steroids (5αTHF+5ßTHF) to THE in patients with CKD, postrenal transplant, and essential hypertension and to compare the ratio with controls. DESIGN/METHODS: We enrolled 44 patients (17 with CKD, eight postrenal transplant, 19 with essential hypertension) and 12 controls. We measured spot urinary 5α-THF, 5ß-THF, THE, free active cortisol and inactive cortisone by gas chromatography/mass spectrometry. We collected data on age, sex, cause of kidney disease, height, weight, body mass index, blood pressure, serum electrolytes, aldosterone, and plasma renin activity. Blood pressure percentiles and z-scores were calculated. The glomerular filtration rate was calculated using the modified Schwartz formula. RESULTS: The ratios of 5αTHF+5ßTHF to THE were significantly higher in patients with CKD [mean±sd score (SDS)=1.31±1.07] as compared with essential hypertension (mean±SDS=0.59±0.23; P=0.02) and controls (mean±SDS=0.52±0.25; P=0.01). In the postrenal transplant group, the ratio was not significantly different (mean±SDS=0.71±0.55). The urinary free cortisol to free cortisone ratios were significantly higher in the hypertension and CKD groups as compared with the controls. The 5αTHF+5ßTHF to THE ratio negatively correlated with the glomerular filtration rate and positively correlated with systolic and diastolic blood pressure z-scores. The correlation of the blood pressure z-scores with ratios was stronger in the CKD group than the essential hypertension and posttransplant groups. CONCLUSIONS: We have elucidated a functional deficiency of 11ß-hydroxysteroid dehydrogenase type 2 in children with CKD and a subset of essential hypertension. Urinary 5α-THF, 5ß-THF, and THE analysis by gas chromatography/mass spectrometry should be a part of routine work-up of CKD and hypertensive patients.


Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 2/metabolismo , Hipertensão Renal/metabolismo , Insuficiência Renal Crônica/metabolismo , Adolescente , Criança , Pré-Escolar , Feminino , Taxa de Filtração Glomerular/fisiologia , Humanos , Hidrocortisona/metabolismo , Hipertensão Renal/cirurgia , Lactente , Transplante de Rim , Masculino , Receptores de Mineralocorticoides/metabolismo , Insuficiência Renal Crônica/cirurgia , Tetra-Hidrocortisol/análogos & derivados , Tetra-Hidrocortisol/urina , Tetra-Hidrocortisona/urina , Adulto Jovem
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