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1.
BMC Vet Res ; 15(1): 336, 2019 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-31533706

RESUMO

BACKGROUND: The recognition of illegal administration of synthetic corticosteroids in animal husbandry has been recently challenged by the case of prednisolone, whose occasional presence in the urine of bovines under strong stressful conditions was attributed to endogenous biosynthesis, not to exogenous administration. The study of the natural stress sources possibly inducing endogenous prednisolone production represents a stimulating investigation subject. The biochemical effects of transportation and slaughtering were verified in untreated cows by studying the possible occurrence of prednisolone and its metabolites in urine, liver and adrenal glands, and the cortisol/cortisone quantification. RESULTS: Cortisol, cortisone, prednisolone and its metabolites were measured in urine, collected at farm under natural micturition and then at the slaughterhouse. The study was performed on 15 untreated cows reared in different farms at the end of their productive cycle. 2-3 days after the first urine collection, the animals were transported by trucks to the abattoir, slaughtered, and subjected to a second urine sampling from the bladder. Specimens of liver and adrenal gland were also collected and analysed by means of a liquid chromatography-tandem mass spectrometry (LC-MS/MS) validated method. The stressful conditions of transportation and slaughtering proved to increase considerably the urinary levels of cortisol and cortisone as compared to those collected at farm. Prednisolone was detected in the urine collected at the slaughterhouse of two cows only, at a concentration level (≈0.6 µg L- 1) largely below the official cut off (5.0 µg L- 1) established to avoid false non-compliances. These two animals exhibited the highest urinary cortisol levels of the series. Prednisolone and prednisone were also detected in the adrenal glands of a different cow. Prednisolone metabolites were not detected in any urine, liver, and adrenal gland sample. CONCLUSION: Within the constraints of the condition adopted, this study confirms the sporadic presence of prednisolone traces (2 samples out of 15) and the consistently increased concentration of cortisone and cortisol in the urines collected from cows subjected to truck transportation and subsequent slaughtering. No prednisolone metabolites were detected in any liver and adrenal gland samples, nor in urine specimens, unlike what was previously reported for cows artificially stressed by pharmacological treatment.


Assuntos
Matadouros , Prednisolona/urina , Transportes , Glândulas Suprarrenais/química , Animais , Bovinos , Cortisona/urina , Feminino , Hidrocortisona/urina , Fígado/química , Prednisolona/metabolismo , Estresse Fisiológico/fisiologia
2.
J Steroid Biochem Mol Biol ; 190: 193-197, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30959155

RESUMO

24-h urine free cortisol (UFC) is an indicator of integrated cortisol secretion and established screening tool for Cushing's syndrome (CS). Doubts have been raised regarding specificity of immunoassays, and mass spectrometric techniques have been proposed as an alternative. In the present study we compared diagnostic accuracy of UFC measured with LC-MS/MS vs. immunoassay in patient with CS and patients where CS has been excluded. We examined 24-h urine samples from patients with surgically confirmed CS (n = 77; Cushing's disease (n = 44), ectopic CS (n = 5), adrenal CS (n = 28)) and patients in whom Cushing's syndrome was excluded (n = 97) by long-term follow up. UFC was first measured by automated chemiluminescence immunoassays (ADVIA Centaur, Siemens; LIAISON, DiaSorin). Aliquots of all samples were also analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Statistics: Passing-Bablok Regression, Receiver operating characteristic (ROC) analysis with Youden's index calculation. UFC of CS patients were higher with both immunoassays compared to LC-MS/MS (913 +/- 235 vs. 303 +/- 155 µg/24 h (ADVIA) and 898 +/-216 vs. 399 +/- 196 µg/24 h (LIAISON)). Similarly, UFC were higher with immunoassays than with LC-MS/MS in the control group (223 +/- 10 vs. 23 +/- 2 µg/24 h (ADVIA) and 105 +/- 6 vs. 27 +/- 4 ug/24 h for (LIAISON)). Passing-Bablok regression showed good correlation between LC-MS/MS and ADVIA as well as between LCMS/MS and LIAISON (r = 0.96 and r = 0.99, p < 000.1) but less correlation in controls (r = 0.83 and r = 0.74, respectively, p < 000.1). ROC calculation revealed the highest ROC AUC (0.89) for the LIAISON immunoassay, followed by LC-MS/MS (0.82) and the ADVIA (0.80). In direct comparison, AUCs from LC-MS/MS and immunoassays in the same patient were not statistically different (p < 0,001). Best cut-off concentration to identify patients with CS was 234 µg/24 h (LIAISON), 51 µg/24 h for LC-MS/MS and 330 µg/24 h (ADVIA Centaur). In summary, UFC values were measured substantially higher by both immunoassays compared to LC-MS/MS. This is most likely due to cross-reactivity from interfering glucocorticoid metabolites. Nevertheless, all three methods correlated well. ROC analysis revealed the highest AUC for one of the immunoassays, although differences between the three methods were not significant. Direct comparison with LC-MS/MS indicates that high diagnostic accuracy can be obtained with suitable immunoassays.


Assuntos
Cortisona/urina , Síndrome de Cushing/urina , Adulto , Cromatografia Líquida/métodos , Síndrome de Cushing/diagnóstico , Feminino , Humanos , Imunoensaio/métodos , Masculino , Pessoa de Meia-Idade , Espectrometria de Massas em Tandem/métodos
3.
Ann Clin Biochem ; 56(1): 82-89, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-29848040

RESUMO

BACKGROUND: The analysis of steroids in biological matrices is challenging. One can apply immunoassay as well as gas and liquid chromatography with various types of detection, depending on the available equipment and the experience of the analyst. The question is how the methods are interchangeable between themselves. Doubts were reported having compared immunoassays and chromatography-mass spectrometry, but there are scarce data on chromatographic methods with detection types other than mass spectrometry. METHODS: Here, we present the detailed comparison of two liquid chromatographic methods for the determination of free urinary cortisol and cortisone: one with fluorescence detection (high-performance liquid chromatography [HPLC-FLD]) and the other with tandem mass spectrometry (HPLC-MS/MS). The comparison was made with 199 human urine samples. The data analysis included Passing-Bablok and Deming regression, Bland-Altman test, Wilcoxon test, mountain plot and Lin's concordance correlation coefficient. RESULTS: The validation data indicated that both methods met the requirements of the European Medicines Agency. However, the statistical analysis revealed the systematic bias between the two assays. The Passing-Bablok and the Deming tests showed that the HPLC-FLD method overestimated results for cortisol and underestimated measurements for cortisone. The Bland-Altman analysis estimated the mean differences between the methods: 18.8 nmol/L for cortisol and -16.9 nmol/L for cortisone measurement. CONCLUSIONS: Both methods' results led to the same conclusion in observational studies, but the techniques are not interchangeable. The literature data, the observations from the clinical setting and our experience clearly indicate that the future of steroid measurements will belong to chromatography coupled with mass spectrometry.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Cortisona/urina , Hidrocortisona/urina , Espectrometria de Massas em Tandem/métodos , Feminino , Humanos , Gravidez , Espectrometria de Fluorescência/métodos
4.
Endocr Pract ; 24(10): 875-881, 2018 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-29975580

RESUMO

OBJECTIVE: Pituitary patients with different etiologies of hypopituitarism exhibit differing phenotypes, despite similar replacement therapy strategies. We hypothesized that differential regulation of the isoenzyme 11ß-hydroxysteroid dehydrogenase 1 (11ß-HSD1), which mediates the net autocrine conversion of cortisone to cortisol in adipose tissues and liver, may play a role. METHODS: We studied 11ß-HSD1 activity (using urine cortisol/cortisone metabolites ratio) in 36 hypopituitary patients with treated craniopharyngiomas, treated remitted Cushing disease, and treated nonfunctioning pituitary adenomas + prolactinomas on and off growth hormone (GH) replacement. RESULTS: 11ß-HSD1 activity was higher in subjects with craniopharyngioma both on and off GH, as evidenced by increased tetrahydrocortisol to tetrahydrocortisone metabolite ratios compared to other diagnostic groups, but there was no difference in body mass index, insulin levels, serum hormone measurements, or hydrocortisone dose between groups. CONCLUSION: Craniopharyngiomas are associated with enhanced 11ß-HSD1 activity compared to other diagnostic hypopituitary groups, and this may contribute to the adverse phenotypic and metabolic features seen in this condition. ABBREVIATIONS: BMI = body mass index; Em = cortisone metabolites; Fm = cortisol metabolites; GH = growth hormone; 11ß-HSD1 = 11ß-hydroxysteroid dehydrogenase type 1; IGF-1 = insulin-like growth factor 1; NFPA = nonfunctioning pituitary adenoma; THE = tetrahydrocortisone; THF = tetrahydrocortisol.


Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 1/metabolismo , Hipopituitarismo/etiologia , Hipopituitarismo/metabolismo , Adulto , Estudos de Casos e Controles , Cortisona/metabolismo , Cortisona/urina , Craniofaringioma/complicações , Craniofaringioma/tratamento farmacológico , Craniofaringioma/metabolismo , Feminino , Transtornos do Crescimento/complicações , Transtornos do Crescimento/tratamento farmacológico , Transtornos do Crescimento/metabolismo , Terapia de Reposição Hormonal , Hormônio do Crescimento Humano/deficiência , Hormônio do Crescimento Humano/uso terapêutico , Humanos , Hidrocortisona/metabolismo , Hidrocortisona/urina , Hipopituitarismo/tratamento farmacológico , Masculino , Pessoa de Meia-Idade , Hipersecreção Hipofisária de ACTH/complicações , Hipersecreção Hipofisária de ACTH/tratamento farmacológico , Hipersecreção Hipofisária de ACTH/metabolismo , Neoplasias Hipofisárias/complicações , Neoplasias Hipofisárias/tratamento farmacológico , Neoplasias Hipofisárias/metabolismo , Prolactinoma/complicações , Prolactinoma/tratamento farmacológico , Prolactinoma/metabolismo
5.
J Korean Med Sci ; 33(53): e298, 2018 Dec 31.
Artigo em Inglês | MEDLINE | ID: mdl-30595680

RESUMO

Background: The renal function of individuals is one of the reasons for the variations in therapeutic response to various drugs. Patients with renal impairment are often exposed to drug toxicity, even with drugs that are usually eliminated by hepatic metabolism. Previous study has reported an increased plasma concentration of indoxyl sulfate and decreased plasma concentration of 4ß-hydroxy (OH)-cholesterol in stable kidney transplant recipients, implicating indoxyl sulfate as a cytochrome P450 (CYP) inhibiting factor. In this study, we aimed to evaluate the impact of renal impairment severity-dependent accumulation of indoxyl sulfate on hepatic CYP3A activity using metabolic markers. Methods: Sixty-six subjects were enrolled in this study; based on estimated glomerular filtration rate (eGFR), they were classified as having mild, moderate, or severe renal impairment. The plasma concentration of indoxyl sulfate was quantified using liquid chromatography-mass spectrometry (LC-MS). Urinary and plasma markers (6ß-OH-cortisol/cortisol, 6ß-OH-cortisone/cortisone, 4ß-OH-cholesterol) for hepatic CYP3A activity were quantified using gas chromatography-mass spectrometry (GC-MS). The total plasma concentration of cholesterol was measured using the enzymatic colorimetric assay to calculate the 4ß-OH-cholesterol/cholesterol ratio. The correlation between variables was assessed using Pearson's correlation test. Results: There was a significant negative correlation between MDRD eGFR and indoxyl sulfate levels. The levels of urinary 6ß-OH-cortisol/cortisol and 6ß-OH-cortisone/cortisone as well as plasma 4ß-OH-cholesterol and 4ß-OH-cholesterol/cholesterol were not correlated with MDRD eGFR and the plasma concentration of indoxyl sulfate. Conclusion: Hepatic CYP3A activity may not be affected by renal impairment-induced accumulation of plasma indoxyl sulfate.


Assuntos
Citocromo P-450 CYP3A/metabolismo , Fígado/metabolismo , Insuficiência Renal/patologia , Adulto , Idoso , Biomarcadores/sangue , Biomarcadores/urina , Colesterol/sangue , Cromatografia Líquida de Alta Pressão , Cortisona/química , Cortisona/urina , Feminino , Cromatografia Gasosa-Espectrometria de Massas , Taxa de Filtração Glomerular , Humanos , Hidrocortisona/química , Hidrocortisona/urina , Indicã/sangue , Rim/metabolismo , Masculino , Espectrometria de Massas , Pessoa de Meia-Idade , Insuficiência Renal/metabolismo , Índice de Gravidade de Doença
6.
J Clin Res Pediatr Endocrinol ; 9(4): 344-349, 2017 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-28720554

RESUMO

OBJECTIVE: The precise mechanisms behind the development of hypertension in overweight or obese children are not yet completely understood. Alterations in hypothalamic-pituitary-adrenal axis activity may play a role. We aimed to investigate the association between cortisol parameters and hypertension in overweight or obese children. METHODS: Random urine (n=180) and early-morning saliva samples (n=126) for assessment of cortisol and cortisone were collected from 1) hypertensive overweight children (n=50), 2) normotensive overweight children (n=145), and 3) normotensive non-overweight children (n=75). RESULTS: The age of participants was 10.4±3.3 years and 53% were boys. The urinary cortisol-to-cortisone ratio [ß 1.11, 95% confidence interval (CI) 1.05-1.19] as well as urinary cortisol/creatinine (ß 1.38, 95% CI 1.09-1.54), and cortisone/creatinine ratios (ß 1.26, 95% CI 1.17-1.36) were significantly higher in overweight or obese than in non-overweight children. After adjusting for body mass index-standard deviation score and urinary cortisone/creatinine ratio, but not cortisol/creatinine ratio, was significantly associated with presence of hypertension (ß 1.12, 95% CI 1.02-1.23). Salivary cortisol and cortisone levels were significantly lower in overweight or obese than in non-overweight children (ß -4.67, 95% CI -8.19- -1.15, and ß 0.89, 95% CI 0.80-0.97 respectively). There were no significant differences in cortisol parameters between hypertensive and normotensive overweight or obese children. CONCLUSION: This study provided further evidence for an increased cortisol production rate with decreased renal 11ß-hydroxysteroid dehydrogenase 2 activity and flattening of early-morning peak cortisol and cortisone in overweight or obese children. However, there were no significant differences in cortisol parameters between hypertensive and normotensive overweight and obese children.


Assuntos
Hidrocortisona/metabolismo , Hipertensão/epidemiologia , Sobrepeso/epidemiologia , Obesidade Pediátrica/epidemiologia , Adolescente , Estudos de Casos e Controles , Criança , Pré-Escolar , Ritmo Circadiano/fisiologia , Cortisona/metabolismo , Cortisona/urina , Creatinina/metabolismo , Creatinina/urina , Feminino , Humanos , Hidrocortisona/urina , Hipertensão/complicações , Hipertensão/metabolismo , Masculino , Sobrepeso/complicações , Sobrepeso/metabolismo , Sobrepeso/fisiopatologia , Obesidade Pediátrica/complicações , Obesidade Pediátrica/metabolismo , Obesidade Pediátrica/fisiopatologia , Saliva/química , Saliva/metabolismo
7.
Res Vet Sci ; 114: 423-429, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28755555

RESUMO

Natural corticosteroids include two families of substances: mineralocorticoids and glucocorticoids. Several drugs of similar structure and biological activity have been synthesized and are currently used in the clinical practice. Beside legal pharmacological treatments, these drugs have been consistently misused in animal breeding. One of the most abused corticosteroids is prednisolone. For many years, prednisolone has been considered of exclusive synthetic origin, but nowadays a debate about its possible endogenous production is under way. Several studies have been addressed to ascertain the potential relationship between stressful conditions, such as transportation and slaughtering, and endogenous production of prednisolone. In order to verify further the effect of stressful conditions, our laboratory analysed urine samples collected from the cows participating to the "Batailles des Reines" (a traditional contest based on ritual and spontaneous fights of pregnant cows), to verify if an endogenous prednisolone production may occur in these animals. We developed and validated a LC-MS/MS method for the simultaneous determination of cortisol, cortisone, prednisolone and five of its metabolites. The method was applied to the analysis of urine samples collected from "Batailles des Reines" competitions in 2012 and 2013. All these samples had been previously analysed within an anti-doping control program and tested compliant to all screenings.


Assuntos
Corticosteroides/urina , Agressão , Bovinos/fisiologia , Cromatografia Líquida/veterinária , Estresse Fisiológico , Espectrometria de Massas em Tandem/veterinária , Animais , Cromatografia Líquida/métodos , Cortisona/urina , Hidrocortisona/urina , Masculino , Prednisolona/urina , Espectrometria de Massas em Tandem/métodos
8.
J Chromatogr B Analyt Technol Biomed Life Sci ; 1061-1062: 163-168, 2017 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-28735224

RESUMO

Cortisol and cortisone are two important glucocorticoids in human body, their interconversion is controlled by two isotypes of 11ß-hydroxy steroid dehydrogenase (11ß-HSD1 and 11ß-HSD2). The ratio of urinary cortisol to cortisone can be used to assess the activity of 11ß-HSDs. An analytical method to quantify urinary cortisol and cortisone using high performance liquid chromatographic tandem mass spectrometry following a packed-fiber solid-phase extraction (PFSPE) was developed. The proposed method was validated and applied to determine the urinary cortisol and cortisone concentrations in infants. Linearity was observed in the range of 0.6-150ng/mL for cortisol and 0.8-200ng/mL for cortisone. The intra-day RSD was 2.4-4.5% for cortisol and 3.3-6.2% for cortisone. Inter-day RSD was 3.7-6.6% for cortisol and 4.3-8.2% for cortisone. The recovery was 97.8±4.6% for cortisol and 98.9±4.4% for cortisone. The established method is simple and efficient for the quantification of urinary cortisol and cortisone and for indirectly assessing the activity of 11ß-hydroxy steroid dehydrogenase.


Assuntos
Cromatografia Líquida de Alta Pressão/métodos , Cortisona/urina , Hidrocortisona/urina , Extração em Fase Sólida/métodos , Espectrometria de Massas em Tandem/métodos , Cortisona/isolamento & purificação , Humanos , Hidrocortisona/isolamento & purificação , Lactente , Limite de Detecção , Modelos Lineares , Reprodutibilidade dos Testes
9.
Clin Endocrinol (Oxf) ; 87(5): 500-507, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28590513

RESUMO

BACKGROUND AND AIM: Differential diagnosis between Cushing's Disease (CD) and Ectopic ACTH Syndrome (EAS) may be a pitfall for endocrinologists. The increasing use in clinical practice of chromatography and mass spectrometry improves the measurement of urinary free cortisol (UFF) and cortisone (UFE). We have recently observed that cortisol to cortisone ratio (FEr) was higher in a small series of EAS; in this study we collected a larger number of ACTH-dependent Cushing's Syndrome (CS) to study the role of FEr to characterize the source of corticotropin secretion. MATERIALS AND METHODS: High-pressure liquid chromatography with UV detection (HPLC-UV, n=35) or liquid chromatography-tandem mass spectrometry (LC-MS/MS, n=72) were used to measure UFF, UFE and FEr in 83 patients with CD and 24 with EAS. RESULTS: UFF, UFE and FEr levels were higher in EAS than in CD (UFF: 6671 vs 549 nmol/24 hours; UFE: 2069 vs 464 nmol/24 hours; FEr: 4.13 vs 0.97; all P<.001). FEr >1.15 (the best ROC-based threshold) was able to distinguish CD from EAS with 75% sensitivity (SE) and 75% specificity (SP), AUC 0.811; results were similar between HPLC-UV (SE 73%, SP 79%, AUC 0.708) and LC-MS/MS (SE 77%, SP 73%, AUC 0.834; P=.727). The diagnostic accuracy of FEr was similar to that of CRH test or high-dose dexamethasone suppression test (respectively P=.171 and P=.683), also combined. Finally, FEr was able to increase the number of correct diagnosis in patients with discordant dynamic tests. CONCLUSIONS: Urinary FEr >1.15 was able to suggest EAS, with a diagnostic accuracy similar to that of other dynamic tests proposed to study ACTH-dependent CS.


Assuntos
Síndrome de ACTH Ectópico/diagnóstico , Cortisona/urina , Hidrocortisona/urina , Hipersecreção Hipofisária de ACTH/diagnóstico , Adulto , Idoso , Cromatografia Líquida , Diagnóstico Diferencial , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Sensibilidade e Especificidade , Espectrometria de Massas em Tandem
10.
Artigo em Inglês | MEDLINE | ID: mdl-28505491

RESUMO

Levels of urinary glucocorticoids and their concentration ratios have been analyzed as potential markers for various pathological statuses. Large-scale studies may possibly accelerate the investigations; however, a suitable method needs to be established. Analytical conditions for measurement of urinary glucocorticoids with LCMS were examined. Electrospray ionization in the positive ion mode was applied for detection of cortisol (precursor>product ion: 363.3>121.0), cortisol-d4 (internal standard, IS, 367.4>121.1), and cortisone (361.2>163.2). To maximize ionization, acetic acid-ammonium acetate buffer (18mM) at pH 5.3 was employed as eluent A. A C18 column (100mm×2.1mm, 2.7µm) at 50°C was used for the 9.5min binary gradient separation starting with 60% eluent A with methanol being eluent B. Linear correlations were observed between the concentrations and the peak areas in the concentration range of 1-300ng/mL with correlation coefficients (r) of 0.998 and 0.997 for cortisol and cortisone, respectively, without IS adjustment, and 0.999 with IS adjustment for both cortisol and cortisone. Solid-phase extraction (SPE) using a 2mL centrifuge column was performed for the urine samples, with the original and final volumes being 100µL. The SPE of 12 urine specimens could be performed within 30min. The effect of the sample matrix on the quantification of endogenous compounds present in the urine extract was limited (coefficient of variation (CV) of IS-adjusted matrix factor: 4.4-8.1%; urine extracts of 8 individuals); however, substantial peak reduction of cortisol was observed at low concentrations. Exogenous contaminants originating from the SPE centrifuge column seemed to be a main cause for this phenomenon because the pure-water extract showed similar peak reduction. A recovery of ∼50% was obtained for both cortisol and cortisone. Adjustment with the IS improved the apparent recovery, with ∼100% being obtained for both cortisol and cortisone. The recovery rate decreased when the urine samples were concentrated in the SPE step; the reduction was greater for cortisol than for cortisone. The lower limit of quantification (LLOQ) was set at 2.5ng/mL when the injection volume was 10µL, based on the reproducibility of the standards which were measured (CV of 12 repetitions: 10.1% for 0.5ng/mL cortisol and 19.6% for 1ng/mL cortisone), the matrix effect (-55% at 2ng/mL concentrations of cortisol), and the recovery rate (∼50%). Furthermore an alternative approach for preparation of the cortisol standards was required for low concentration range (2.5-20ng/mL) because of the effect of the matrix. Degradation of original urine specimens at room temperature was minimal during the first 24h. The extracted urine samples degraded over time; however, their concentrations were corrected with the IS, allowing for analysis up to 5days after extraction. In conclusion, an analytical method for urinary glucocorticoids was established, which is fast, sensitive, and well suited for practical application to large-scale study.


Assuntos
Cromatografia Líquida de Alta Pressão , Cortisona/urina , Glucocorticoides/urina , Hidrocortisona/urina , Espectrometria de Massas , Biomarcadores/análise , Biomarcadores/urina , Calibragem , Humanos , Sensibilidade e Especificidade , Extração em Fase Sólida
11.
Med Sci Monit Basic Res ; 23: 25-30, 2017 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-28179618

RESUMO

BACKGROUND It has been unclear whether relatively high cortisol and cortisone levels are related to overweight in childhood, parental body mass index (BMI), and family dietary habits. The aim of this study was to compare cortisol and cortisone levels in urine and saliva from overweight and normal children, as well as correlations between children's BMI, parental BMI and family dietary behavior questionnaire score (QS). MATERIAL AND METHODS We analyzed the data from 52 overweight children and 53 age- and sex-matched normal-weight children aged 4-5 years. The concentrations of salivary cortisol (SF), salivary cortisone (SE), urinary cortisol (UF) and urinary cortisone (UE) were measured using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). The family dietary behavior QS was answered by the parent mainly responsible for the family diet. RESULTS Average cortisol and cortisone levels were significantly higher in overweight children. There was no significant difference in the ratio of cortisol to cortisone (Rcc) and the marker of 11b-hydroxysteroid dehydrogenase type 2 (11ß-HSD2) activities. The results displayed correlations among cortisol, cortisone, and Rcc. Positive correlations were weak-to-moderate between BMI and SF, SE, UF, and UE. There were correlations between BMI and maternal BMI (mBMI), and BMI was significantly associated with QS. CONCLUSIONS Our results suggest that cortisol and cortisone levels are associated with overweight in children, but the 11ß-HSD2 activities showed no significant differences. Unhealthy family diet was associated with higher BMI, UF, and UE, and families with maternal overweight or obesity had a higher prevalence of children's overweight or obesity.


Assuntos
Cortisona/metabolismo , Hidrocortisona/metabolismo , Obesidade/metabolismo , Sobrepeso/metabolismo , Saliva/metabolismo , Estudos de Casos e Controles , Pré-Escolar , Cortisona/urina , Feminino , Humanos , Hidrocortisona/urina , Masculino , Obesidade/urina , Sobrepeso/urina
12.
Am J Hypertens ; 30(5): 518-523, 2017 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-28180242

RESUMO

BACKGROUND AND OBJECTIVE: Decreased renal 11-beta dehydrogenase type 2 (11ß-HSD2) activity, as reflected by an increased urinary free cortisol to cortisone ratio (UFF/UFE), is associated with having hypertension (HTN). The current study was conducted to determine if reduced 11ß-HSD2 activity is also associated with having resistant HTN. METHOD: We evaluated 55 consecutive patients with RHTN, defined as blood pressure (BP) ≥140/90 mm Hg despite using ≥3 antihypertensive medications including a diuretic, and 38 patients whose BP was controlled on ≤3 medications to serve as a non-RHTN comparator group. All patients underwent biochemical evaluation, including measurement of 24-hour urinary UFF/UFE. RESULTS: The 2 study groups had similar demographic characteristics. Systolic, diastolic BP, and number of antihypertensive medications were greater in patients with uncontrolled RHTN vs. the control group (167.5 ± 28.2/91.2 ± 18.8 vs. 126.6 ± 11.4/77.8 ± 8.65 mm Hg and 4.31 ± 1.23 vs. 2.74 ± 0.6, respectively). The 24-hour UFF was 13.6 ± 11.8 vs. 14.3 ± 10.7 µg/24 h and UFE was 64.9 ± 36.3 vs. 76.1 ± 44 µg/24 h such that the UFF/UFE was 0.22 ± 0.16 vs. 0.19 ± 0.09 in RHTN vs. the control group. This ratio was not associated to age, race, gender, and body mass index. CONCLUSION: An elevated UFF/UFE was not present in this large cohort of patients with uncontrolled RHTN. This suggests that reduced conversion of cortisol to cortisone does not contribute to the development of RHTN.


Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 2/metabolismo , Pressão Sanguínea , Resistência a Medicamentos , Hipertensão/enzimologia , Adulto , Idoso , Anti-Hipertensivos/uso terapêutico , Biomarcadores/urina , Pressão Sanguínea/efeitos dos fármacos , Cortisona/urina , Estudos Transversais , Quimioterapia Combinada , Feminino , Humanos , Hidrocortisona/urina , Hipertensão/diagnóstico , Hipertensão/tratamento farmacológico , Hipertensão/fisiopatologia , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos
13.
J Anim Physiol Anim Nutr (Berl) ; 101(5): 893-903, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-27080058

RESUMO

The Chianina, one of the oldest and most important cattle breeds of Italy, is now reared all over the world. The Chianina has been known and appreciated since ancient times because, from a nutritional point of view, its meat has no proper rivals. To date, studies have been performed to evaluate the genetic profile of the breed, but knowledge about the chemical profile is generally lacking. Due to the increased interest from farmers regarding breeding of the Chianina, this study proposes a preliminary evaluation of main endogenous urinary corticosteroids (cortisol and cortisone) and most commonly used synthetic one (dexamethasone). Moreover, after recent findings regarding the presence of endogenous prednisolone in the urine of more popular breeds, particular attention was given to analysis of the presence of prednisolone and prednisone, as well. For this aim, the urine samples of 12 young cows and 30 young bulls was collected at the farms and analysed using a fit-for-purpose LC-MS/MS method. The preliminary results of this study show that prednisolone was found only in Chianina females (3 out of 12). Cortisol and cortisone were found at concentrations that showed a high inter-individual variability, and that were higher in female urine compared to that of males.


Assuntos
Bovinos/fisiologia , Cortisona/urina , Hidrocortisona/urina , Prednisolona/urina , Animais , Bovinos/genética , Bovinos/urina , Cromatografia Líquida , Cortisona/química , Feminino , Hidrocortisona/química , Masculino , Estrutura Molecular , Prednisolona/química , Espectrometria de Massas em Tandem
14.
J Steroid Biochem Mol Biol ; 165(Pt A): 151-153, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-26956190

RESUMO

Apparent mineralocorticoid excess is a syndrome reflecting the absent or impaired activity of the enzyme 11ß-hydroxysteroid dehydrogenase Type 2. It may be mild when the mutant enzyme retains some activity, or severe when activity is absolutely or essentially absent. Diagnosis relies on a triad of hypertension, hypokalemia and suppressed plasma aldosterone levels, plus an abnormal urinary cortisol to cortisone ratio, either free steroid or metabolites. Treatment is symptomatic in the mild form - correction of hypertension and hypokalemia - but needs to be prompt, vigorous and sustained in the severe form, which usually presents in neonates/infancy. Elucidation of the pathogenesis of apparent mineralocorticoid excess is an example of 'reverse translation', in that it proved prismatic for the demonstration of the physiologic mechanisms underlying the selective activation of epithelial mineralocorticoid receptors by aldosterone.


Assuntos
11-beta-Hidroxiesteroide Desidrogenase Tipo 2/genética , Síndrome de Excesso Aparente de Minerolocorticoides/diagnóstico , Aldosterona/sangue , Aldosterona/metabolismo , Anti-Hipertensivos/química , Cortisona/urina , Diagnóstico Diferencial , Dieta , Humanos , Hidrocortisona/urina , Hipertensão/complicações , Hipertensão/genética , Hipopotassemia/complicações , Hipopotassemia/genética , Síndrome de Excesso Aparente de Minerolocorticoides/terapia , Mineralocorticoides/metabolismo , Receptores de Mineralocorticoides/metabolismo , Esteroides/metabolismo
15.
Steroids ; 118: 61-67, 2017 02.
Artigo em Inglês | MEDLINE | ID: mdl-27998757

RESUMO

BACKGROUND: Cortisol, cortisone and the ratio of cortisol to cortisone in saliva, urine and hair are acute, short-term and long-term biomarkers to reliably assess the activity of hypothalamic-pituitary-adrenal (HPA) axis and 11ß-hydroxysteroid dehydrogenase (11ß-HSD). One key issue is whether these biomarkers have intraindividual relative stability. Salivary, urinary and hair cortisol was proven to show considerable long-term intraindividual relative stability. However, currently unknown is whether cortisone and the ratio in saliva, urine and hair show intraindividual relative stability. METHODS: The present study utilized a longitudinal design to validate long-term stability within two weeks of three biomarkers in saliva and urine, and long-term stability within twelve months of three hair biomarkers. Salivary, urinary and hair steroids were measured with high performance liquid chromatography tandem mass spectrometry. RESULTS: Three biomarkers in urine and hair showed moderate test-retest correlations with coefficient (r) ranging between 0.22 and 0.56 and good multiple-test consistencies with coefficient of intraclass correlation (ICC) ranging between 0.42 and 0.67. Three single-point salivary biomarkers showed weak to moderate test-retest correlations (r's between 0.01 and 0.38) and poor to fair multiple-test consistencies (ICC's between 0.29 and 0.53) within two weeks. Three single-day salivary biomarkers showed moderate test-retest correlations (r's between 0.23 and 0.53) and good multiple-test consistencies (ICC's between 0.56 and 0.66) within two weeks. CONCLUSIONS: Three biomarkers in urine and hair showed moderate long-term intraindividual relative stability. Three single-point salivary biomarkers showed weak to moderate short-term and long-term intraindividual relative stability, but three single-day salivary biomarkers showed moderate short-term and long-term intraindividual relative stability.


Assuntos
Biomarcadores/análise , Biomarcadores/urina , Cortisona/análise , Cortisona/urina , Cabelo/química , Hidrocortisona/análise , Hidrocortisona/urina , Saliva/química , Adolescente , Cromatografia Líquida de Alta Pressão , Feminino , Humanos , Masculino , Espectrometria de Massas em Tandem , Adulto Jovem
16.
Bioanalysis ; 8(23): 2429-2443, 2016 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-27855510

RESUMO

AIM: A UHPLC-MS/MS assay was developed to quantify urinary dehydroepiandrosterone (DHEA), 7ß-hydroxy-DHEA, cortisone and 6ß-hydroxycortisone as potential biomarkers to predict CYP3A activity. RESULTS: A sensitive assay at LLOQ of 0.500 ng/ml with good accuracy and precision was developed for the four analytes in human urine. This UHPLC-MS/MS assay was optimized by eliminating nonspecific loss of the analytes in urine, ensuring complete hydrolysis of the conjugates to unconjugated forms and use of the product ions of [M+H-H2O]+ for multiple reaction monitoring detection of DHEA and 7ß-hydroxy-DHEA. CONCLUSION: This assay was successfully applied to a pilot clinical study. It is also suitable for future drug-drug interaction studies to continue evaluating the potential of these steroids as biomarkers for CYP3A inhibition and induction.


Assuntos
Biomarcadores/urina , Cortisona/urina , Citocromo P-450 CYP3A/metabolismo , Desidroepiandrosterona/urina , Espectrometria de Massas em Tandem , Urinálise/métodos , Cromatografia Líquida de Alta Pressão/normas , Cortisona/metabolismo , Cortisona/normas , Citocromo P-450 CYP3A/química , Desidroepiandrosterona/metabolismo , Desidroepiandrosterona/normas , Interações Medicamentosas , Humanos , Hidroxilação , Limite de Detecção , Extração Líquido-Líquido , Controle de Qualidade , Espectrometria de Massas em Tandem/normas , Urinálise/instrumentação
17.
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
18.
Kidney Int ; 90(2): 325-333, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27165611

RESUMO

Metabolic acidosis induces elevated glucocorticoid (GC) levels. However, the influence of less strong daily acid loads on GCs is largely unexplored. To investigate this, we studied whether higher acid loads in children, fully within the normal range of habitual diets, associate with endogenous GCs. In a specific quasi-experimental design, we examined 200 6- to 10-year-old healthy participants of the Dortmund Nutritional and Anthropometric Longitudinally Designed (DONALD) Study equally divided to either high or low 24-hour renal net acid excretion. Major urinary GC metabolites were analyzed by gas chromatography-mass spectrometry to assess daily adrenal GC secretion and metabolites of tissue cortisol catabolism (6ß-hydroxycortisol and 20α-dihydrocortisol). Liquid chromatography-mass spectrometry was used to quantify urinary free cortisol and cortisone. After confounder adjustment, significant positive associations were unmasked for urinary potential renal acid load and net acid excretion with adrenal GC secretion, free cortisone, free cortisone plus cortisol, 6ß-hydroxycortisol, and 20α-dihydrocortisol. An inverse association emerged for an enzymatic marker (5ß-reductase) of irreversible GC inactivation. Our data suggest that existing moderate elevations in diet-dependent acid loads suffice to raise GCs and affect cortisol metabolism. Thus, potential detrimental effects of high acid loading appear to be mediated, in part, by increased GC activity via increased GC secretion and/or reduced GC inactivation. Higher cortisone levels, directly available for intracrine activation to cortisol may play a special role.


Assuntos
Acidose/metabolismo , Glucocorticoides/metabolismo , Rim/metabolismo , Eliminação Renal , Criança , Cortisona/metabolismo , Cortisona/urina , Dieta , Feminino , Cromatografia Gasosa-Espectrometria de Massas , Glucocorticoides/urina , Voluntários Saudáveis , Humanos , Hidrocortisona/análogos & derivados , Hidrocortisona/metabolismo , Hidrocortisona/urina , Masculino , Valores de Referência
19.
Acta Pharmacol Sin ; 37(4): 555-60, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26924289

RESUMO

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


Assuntos
Citocromo P-450 CYP3A/genética , Citocromo P-450 CYP3A/metabolismo , Imunossupressores/metabolismo , Transplante de Rim , Tacrolimo/metabolismo , Grupo com Ancestrais do Continente Asiático , Cortisona/análogos & derivados , Cortisona/sangue , Cortisona/urina , Humanos , Hidrocortisona/análogos & derivados , Hidrocortisona/sangue , Hidrocortisona/urina , Imunossupressores/administração & dosagem , Tacrolimo/administração & dosagem
20.
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
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