Search

Background: In adults, cortisol levels show a pronounced 24-hour rhythm with a peak in the early morning. It is unknown at what age this early-morning peak in cortisol emerges during infancy, hampering the establishment of optimal dosing regimens for hydrocortisone replacement therapy in infants with an inborn form of adrenal insufficiency. Therefore, we aimed to characterize daily variation in salivary cortisol concentration across the first year of life., Methods: We conducted a systematic review followed by an individual participant data meta-analysis of studies reporting on spontaneous (i.e., not stress induced) salivary cortisol concentrations in healthy infants aged 0-1 year. A one-stage approach using linear mixed-effects modelling was used to determine the interaction between age and time of day on cortisol concentrations., Findings: Through the systematic review, 54 eligible publications were identified, reporting on 29,177 cortisol observations. Individual participant data were obtained from 15 study cohorts, combining 17,079 cortisol measurements from 1,904 infants. The morning/evening cortisol ratio increased significantly from 1.7 (95% CI: 1.3-2.1) at birth to 3.7 (95% CI: 3.0-4.5) at 6-9 months (p < 0.0001). Cosinor analysis using all available data revealed the gradual emergence of a 24-hour rhythm during infancy., Interpretation: The early-morning peak in cortisol secretion gradually emerges from birth onwards to form a stable morning/evening ratio from age 6-9 months. This might have implications for hydrocortisone replacement therapy in infants with an inborn form of adrenal insufficiency., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society.)

Context: Adrenocortical carcinomas are very rare malignancies in childhood associated with poor outcome in advanced disease. Most adrenocortical tumors (ACT) are functional, causing signs and symptoms of adrenal hormone excess. In most studies, endocrine manifestations were reported 4 to 6 months prior to diagnosis., Objective: We sought to extend knowledge on endocrine manifestations with regard to age and sex to facilitate early diagnosis., Methods: We retrospectively analyzed features of adrenal hormone excess in children and adolescents with ACT registered with the GPOH-MET studies between 1997 and 2022. Stage of puberty was defined as prepubertal in females < 8 years of age and males < 9 years., Results: By December 2022, 155 patients (110 female, 45 male) with data on endocrine manifestations had been reported. Median age at ACT diagnosis was 4.2 years [0.1-17.8], median interval from first symptoms was 4.2 months [0-90.7]. In 63 girls of prepubertal age, the most frequently reported manifestations were pubarche (68.3%), clitoral hypertrophy (49.2%), and weight gain (31.7%); in 47 pubertal female patients, the most frequent manifestations were excessive pubic hair (46.8%), acne (36.2%), and hypertension (36.2%). Leading symptoms in 34 boys of prepubertal age were pubarche (55.9%), penile growth (47.1%), and acne (32.4%), while in 11 pubertal male patients, leading symptoms were weight gain (45.5%), hypertension (36.4%), excessive pubic hair (27.3%), and cushingoid appearance (27.3%). In pubertal patients, symptoms of androgen excess were mainly unrecognized as part of pubertal development, while symptoms of Cushing syndrome were more frequently apparent., Conclusion: The endocrine phenotype induced by pediatric ACT is age- and sex-dependent., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Symptoms of hyperandrogenism are common in patients with Cushing's disease (CD), yet they are not sufficiently explained by androgen concentrations. In this study, we analyzed the contribution of 11-oxygenated C19 steroids (11oxC19) to hyperandrogenemia in female patients with CD., Methods: We assessed saliva day profiles in females with CD pre (n = 23) and post (n = 13) successful transsphenoidal surgery, 26 female controls, 5 females with CD treated with metyrapone and 5 treated with osilodrostat for cortisol, cortisone, androstenedione (A4), 11-hydroxyandrostenedione (11OHA4), testosterone (TS), 11-ketotestosterone (11KT), as well as metabolites of classic and 11-oxygenated androgens in 24-h urine. In addition, morning baseline levels of gonadotropins and estradiol, sex hormone-binding globulin, cortisol and dehydroepiandrosterone sulfate (DHEAS) in serum and adrenocorticotrophic hormone in plasma in patients and controls were investigated., Results: Treatment-naïve females with CD showed a significantly elevated area under the curve of 11OHA4 and 11KT in saliva throughout the day compared to controls (11OHA4 mean rank difference (mrd) 18.13, P = 0.0002; 11KT mrd 17.42; P = 0.0005), whereas A4, TS and DHEAS were comparable to controls. Gonadotropin concentrations were normal in all patients with CD. After transsphenoidal surgery, 11oxC19 and their metabolites dropped significantly in saliva (11OHA4 P < 0.0001; 11KT P = 0.0010) and urine (11-oxo-androsterone P = 0.0011; 11-hydroxy-androsterone P < 0.0001), treatment with osilodrostat and metyrapone efficaciously blocked 11oxC19 synthesis., Conclusion: Hyperandrogenemia in CD is predominantly caused by excess of 11oxC19 steroids.

Congenital adrenal hyperplasia (CAH) is a group of autosomal recessive disorders affecting cortisol biosynthesis. Reduced activity of an enzyme required for cortisol production leads to chronic overstimulation of the adrenal cortex and accumulation of precursors proximal to the blocked enzymatic step. The most common form of CAH is caused by steroid 21-hydroxylase deficiency due to mutations in CYP21A2. Since the last publication summarizing CAH in Endocrine Reviews in 2000, there have been numerous new developments. These include more detailed understanding of steroidogenic pathways, refinements in neonatal screening, improved diagnostic measurements utilizing chromatography and mass spectrometry coupled with steroid profiling, and improved genotyping methods. Clinical trials of alternative medications and modes of delivery have been recently completed or are under way. Genetic and cell-based treatments are being explored. A large body of data concerning long-term outcomes in patients affected by CAH, including psychosexual well-being, has been enhanced by the establishment of disease registries. This review provides the reader with current insights in CAH with special attention to these new developments., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Context: Fetal zone steroids (FZSs) are excreted in high concentrations in preterm infants. Experimental data suggest protective effects of FZSs in models of neonatal disease., Objective: We aimed to characterize the postnatal FZS metabolome of well preterm and term infants., Methods: Twenty-four-hour urinary FZS excretion rates were determined in early preterm (<30 weeks' gestation), preterm (30-36 weeks), and term (>37 weeks) infants. Pregnenolone and 17-OH-pregnenolone metabolites (n = 5), and dehydroepiandrosterone sulfate and metabolites (n = 12) were measured by gas chromatography mass spectrometry. Postnatal concentrations of FZSs were compared with already published prenatal concentrations in amniotic fluid., Results: Excretion rates of total FZSs and most of the single metabolites were highest in early preterm infants. In this group, excretion rates approach those of term infants at term equivalent postmenstrual age. Preterm infants of 30-36 weeks had more than half lower median excretion rates of FZSs than early preterm infants at the same time of postmenstrual age. Postnatal concentrations of FZSs were partly more than 100-fold higher in all gestational age groups than prenatal concentrations in amniotic fluid at midgestation., Conclusion: The excretion rates of FZSs as a proxy of the involution of the fetal zone of the most immature preterm infants approached those of term infants at term equivalent. In contrast, the fetal zone in more mature preterm infants undergoes more rapid involution. These data in exclusively well neonates can serve as a basis to investigate the effects of illness on the FZS metabolome in future studies., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Context: Excess glucocorticoids impact fetal health. Maternal glucocorticoids peak in early morning. Fetoplacental 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2) inactivates cortisol to cortisone, protecting the fetus from high glucocorticoids. However, time-specific alterations of human fetoplacental 11β-HSD2 have not been studied., Objective: We hypothesized that fetoplacental 11β-HSD2 activity shows time-specific alteration and acute affective or anxiety disorders impact fetoplacental 11β-HSD2 activity., Methods: In this observational study we investigated 78 pregnant European women undergoing amniocentesis (15.9 ± 0.9 weeks of gestation). Amniotic fluid was collected (8:00 to 16:30 hours) for analysis of fetoplacental 11β-HSD2 activity, using cortisol (F):cortisone (E) ratio in amniotic fluid, E/(E + F). Fetoplacental 11β-HSD2 rhythm and association with "acute affective or anxiety disorder" (patients with at least one of: a major depressive episode, specific phobia, panic disorder, generalized anxiety disorder, mixed anxiety and depressive disorder) and "acute anxiety disorder" (one of: panic disorder, generalized anxiety disorder, mixed anxiety, depressive disorder), assessed using Mini International Neuropsychiatric Interview, were investigated., Results: Activity of 11β-HSD2 correlated with time of amniocentesis, peaking in the morning (r = -0.398; P < 0.001) and increased with acute affective or anxiety disorder (mean [M] = 0.70 vs M = 0.74; P = 0.037) and acute anxiety disorder (M = 0.70 vs M = 0.75; P = 0.016). These associations remained significant when controlling for confounders. 11β-HSD2 activity correlated negatively with pre-pregnancy body mass index (r = -0.225; P = 0.047)., Conclusion: Our study indicates a time-specific alteration of fetoplacental 11β-HSD2 activity with peaking levels in the morning, demonstrating a mechanism of fetal protection from the morning maternal glucocorticoid surge., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Endocrine Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Objective: Although low birthweight (bw) and unfavorable intrauterine conditions have been associated with metabolic sequelae in later life, little is known about their impact on steroid metabolism. We studied genetically identical twins with intra-twin bw-differences from birth to adolescence to analyze the long-term impact of bw on steroid metabolism., Methods: 68 monozygotic twin pairs with a bw-difference of <1 standard deviation score (SDS; concordant; n = 41) and ≥1 SDS (discordant; n = 27) were recruited. At 14.9 years (mean age), morning urine samples were collected and analyzed with gas chromatography-mass-spectrometry., Results: No significant differences were detected in the concordant group. In contrast, in the smaller twins of the discordant group, we found significantly higher concentrations not only of the dehydroepiandrosterone sulfate (DHEAS) metabolite 16α-OH-DHEA (P = 0.001, 656.11 vs 465.82 µg/g creatinine) but also of cumulative dehydroepiandrosterone and downstream metabolites (P = 0.001, 1650.22 vs 1131.92 µg/g creatinine). Relative adrenal (P = 0.002, 0.25 vs 0.18) and overall androgen production (P = 0.001, 0.79 vs 0.65) were significantly higher in the formerly smaller discordant twins. All twin pairs exhibited significant intra-twin correlations for all individual steroid metabolites, sums of metabolites, indicators of androgen production, and enzyme activities. Multiple regression analyses of the smaller twins showed that individual steroid concentrations of the larger co-twin were the strongest influencing factor among nearly all parameters analyzed., Conclusion: In monozygotic twin pairs with greater intra-twin bw-differences (≥1 SDS), we found that bw had a long-lasting impact on steroid metabolism, with significant differences regarding DHEAS metabolites and relative androgen production. However, most parameters showed significant intra-twin correlations, suggesting a consistent interrelationship between prenatal environment, genetic background, and steroid metabolism., (© Published by Oxford University Press on behalf of the Endocrine Society 2020.)

Background: Perinatal treatment with lopinavir boosted by ritonavir (LPV/r) is associated with steroidogenic abnormalities. Long-term effects in infants have not been studied., Methods: Adrenal-hormone profiles were compared at weeks 6 and 26 between human immunodeficiency virus (HIV)-1-exposed but uninfected infants randomly assigned at 7 days of life to prophylaxis with LPV/r or lamivudine (3TC) to prevent transmission during breastfeeding. LPV/r in vitro effect on steroidogenesis was assessed in H295R cells., Results: At week 6, 159 frozen plasma samples from Burkina Faso and South Africa were assessed (LPV/r group: n = 92; 3TC group: n = 67) and at week 26, 95 samples from Burkina Faso (LPV/r group: n = 47; 3TC group: n = 48). At week 6, LPV/r-treated infants had a higher median dehydroepiandrosterone (DHEA) level than infants from the 3TC arm: 3.91 versus 1.48 ng/mL (P < .001). Higher DHEA levels (>5 ng/mL) at week 6 were associated with higher 17-OH-pregnenolone (7.78 vs 3.71 ng/mL, P = .0004) and lower testosterone (0.05 vs 1.34 ng/mL, P = .009) levels in LPV/r-exposed children. There was a significant correlation between the DHEA and LPV/r AUC levels (ρ = 0.40, P = .019) and Ctrough (ρ = 0.40, P = .017). At week 26, DHEA levels remained higher in the LPV/r arm: 0.45 versus 0.13 ng/mL (P = .002). Lopinavir, but not ritonavir, inhibited CYP17A1 and CYP21A2 activity in H295R cells., Conclusions: Lopinavir was associated with dose-dependent adrenal dysfunction in infants. The impact of long-term exposure and potential clinical consequences require evaluation., Clinical Trials Registration: NCT00640263., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Background: Treatment of children with classic congenital adrenal hyperplasia (CAH) with glucocorticoids is a difficult balance between hypercortisolism and hyperandrogenism. Biochemical monitoring of treatment is not well defined. Achievement of a normal growth rate is the most important therapeutic goal., Methods: We retrospectively evaluated 123 24-hour gas chromatography-mass spectrometry urinary steroid metabolome analyses together with their corresponding 1-year height velocity (HV) z scores in 63 prepubertal children aged 7.2 ± 1.6 years with classic CAH due to 21-hydroxylase deficiency treated with hydrocortisone and fludrocortisone., Results: Multivariate linear mixed effects model analysis revealed a positive influence of CAH-specific z scores of summed urinary androgen metabolites (B = 0.97 ± 0.20, t = 4.87, P < 0.0001) and a negative influence of the cortisol metabolite tetrahydrocortisol (B = -1.75 ± 0.79, t = -2.20, P = 0.03) on HV z scores. Receiver operating characteristic analysis demonstrated that adrenal androgen excess, defined as HV >1.5 z, was best determined by a z score of all urinary androgen metabolites of >0.512 [accuracy, 66.2%; sensitivity, 57.1%; specificity, 74.4%; positive prediction value (PPV), 66.7%; negative prediction value (NPV), 65.9%]. Tetrahydrocortisol excretion >1480 µg/m2 BSA/d in conjunction with suppressed urinary androgen metabolites <0.163 z indicated overtreatment, defined as HV < -1.5 z (accuracy, 79.6%; sensitivity, 40.0%; specificity, 94.9%; PPV, 75.0%; NPV, 80.4%)., Conclusion: We established target values for urinary steroid metabolite excretions in children with CAH based on their growth rate. Urinary steroid metabolome analysis represents a highly suitable method for monitoring metabolic control in children with CAH., (Copyright © 2019 Endocrine Society.)

Context: Elevated bone marrow adipose tissue (BMAT) is associated with lower bone quality, higher fracture rates, and an unfavorable overall metabolic profile. Apart from age, particularly glucocorticoids (GC), body fat, and diet are discussed to influence BMAT. We hypothesized that already in healthy youths, higher fat intake, higher fat mass index (FMI), and higher GC secretion, still within the normal range, may associate with increased BMAT., Design: In a subsample of healthy 6- to 18-year-old participants of the Dortmund Nutritional and Anthropometric Longitudinally Designed Study, peripheral quantitative CT of the nondominant proximal forearm was used to determine bone marrow density of the distal radius as an inverse surrogate parameter for BMAT. In those participants (n = 172) who had collected two, 24-hour urines within around one year before bone measurement, major urinary GC metabolites were measured by gas chromatography-mass spectrometry and summed up to assess daily adrenal GC secretion (ΣC21). Dietary intake was assessed by 3-day weighed dietary records. FMI was anthropometrically calculated. Separate multiple linear regression models were used to analyze the relationships of ΣC21, FMI, and fat intake with BMAT., Results: After controlling for confounders, such as age, energy intake, and forearm muscle area, ΣC21 (β = -0.042) and FMI (β = -0.002) showed inverse relationships with bone marrow density (P < 0.05), whereas fat intake did not associate significantly., Conclusion: Our results indicate that already a moderately elevated GC secretion and higher body fatness during adolescence may adversely impact BMAT, an indicator for long-term bone health., (Copyright © 2019 Endocrine Society.)

Context: The profile of urinary steroids as measured by gas chromatography-mass spectrometry defines a subject's "steroidal fingerprint.", Objective: Here, we clustered steroidal fingerprints to characterize patients with nonsyndromic childhood obesity by "steroid metabolomic signatures.", Hypothesis: Nonsyndromic obesity is a symptom of different diseases and conditions, some of them will have their own signature., Design: A total of 31 steroid metabolites were quantified by gas chromatography-mass spectrometry, and their excretion rates were z-transformed. Using MetaboAnalyst 3.0, we divided the subjects into 5 distinctive groups by k-means clustering. Steroidal fingerprints and clinical/biochemical data of patients in each cluster were analyzed., Patients: A total of 87 obese children (44 females), aged 8.5-17.9 years, were clinically characterized, and their 24-hour urine was collected., Results: Cluster 1 (n = 39, 21 females) had normal steroid profile. Cluster 2 (n = 20, 11 females) showed mild, nonspecific elevation of C19 and C21 steroids, females' resistance to polycystic ovary morphology, and hirsutism. Cluster 3 (n = 7 female), with relative 21-hydroxylase insufficiency, was characterized by partial or full polycystic ovary syndrome. Cluster 4 (n = 4 males), showed markedly elevated C21 steroids and imbalance in the 11β-hydroxysteroid dehydrogenase system, higher insulin, increased frequency of glucose/insulin index more than 0.3, γ-glutamyl transpeptidase activity, systolic blood pressure, and tendency to liver steatosis. Cluster 5 (n = 17, 5 females) had elevated dehydroepiandrosterone and 17-OH-pregnenolone metabolites, suggesting 3β-hydroxysteroid dehydrogenase insufficiency but no clinically unique phenotype. Z-score body mass index values were not significantly different between the clusters., Conclusions: We defined a novel concept of disease-specific steroid metabolomic signature based on urinary steroidal gas chromatography-mass spectrometry. Clustering by software designed for metabolic data analysis reclassified childhood obesity into 5 groups with distinctive signatures; groups require further definition and may require cluster-specific therapeutic strategies.

Context: Aromatase deficiency due to a CYP19A1 defect leads to fetoplacental inability to convert androgens into estrogens. Pregnant mothers experience virilization caused by excess nonaromatized fetal androgens entering the maternal circulation. Biochemical normalization is believed to take place shortly after delivery., Objective: We report prolonged postnatal hyperandrogenism and enlarged multicystic ovaries in the mother of an affected 46,XX infant and hypothesize a possible pathogenetic mechanism., Patients and Methods: We investigated the mother on days 12 and 20 after delivery. FSH, LH, T, estradiol (E2), androstenedione (A), dehydroepiandrosterone-sulfate (DHEA-S), and human chorionic gonadotropin (hCG) plasma levels were obtained, and ovarian ultrasonography and magnetic resonance imaging were performed., Results: T (1040 ng/dL), A (6940 ng/dL), and E2 (2787 pg/mL) levels were markedly elevated on day 12 after delivery, whereas LH and FSH were suppressed (<0.1 IU/L). On day 20, all hormones had decreased significantly; however, T, A, and E2 still remained 3.5-, 2.2-, and 1.4-fold elevated, respectively, as compared to upper reference values. hCG (18.9 U/L) was still increased. DHEA-S was normal on both occasions. Sonography and magnetic resonance imaging revealed enlarged ovaries, with several cysts up to 4 cm. There was no history of polycystic ovary syndrome., Conclusions: We hypothesize that persistent ovarian overstimulation by hCG had occurred in the mother during pregnancy, leading to prolonged autonomous excess production of androgens during the first weeks after delivery. As a causative mechanism, we propose that gestational hyperandrogenism and hypoestrogenism reduced inhibition of placental GnRH and hCG secretion by progesterone, resulting in persistently elevated hCG.

Background: 17-Hydroxyprogesterone (17-OHP) can be converted to dihydrotestosterone (DHT) via an alternative "backdoor" route that bypasses the conventional intermediates androstenedione and testosterone. In this backdoor pathway, 17-OHP is converted to 5α-pregnane-3α,17α-diol-20-one (pdiol), which is an excellent substrate for the 17,20 lyase activity of CYP17A1 to produce androsterone. OBJECTIVE AND HYPOTHESES: The objective of this study was to obtain evidence for the presence of the backdoor pathway in patients with 21-hydroxylase deficiency (21-OHD)., Methods: We compared urinary steroid hormone profiles determined by gas chromatography-mass spectrometry of 142 untreated 21-OHD patients (age range, 1 d to 25.4 yr; 51 males) with 138 control subjects. The activity of the backdoor pathway was assessed using the ratios of the urinary concentrations of pdiol to those of the metabolites of the classic Δ4 and Δ5 pathways. In contrast to etiocholanolone, which originates almost exclusively from the classic pathways, androsterone may be derived additionally from the backdoor pathway. Therefore, the androsterone to etiocholanolone ratio can be used as an indicator for the presence of the backdoor pathway., Results: Untreated 21-OHD subjects showed increased urinary ratios of pdiol to the Δ4 and Δ5 pathway metabolites and a higher androsterone to etiocholanolone ratio., Conclusions: The elevated ratios of pdiol to the Δ4 and Δ5 pathway metabolites as well as the higher androsterone to etiocholanolone ratio in patients with 21-OHD indicate postnatal activity of the backdoor pathway with maximum activity during early infancy. Our data provide new insights into the pathophysiology of androgen biosynthesis of 21-OHD.

Early onset of puberty may confer adverse health consequences. Thus, modifiable factors influencing the timing of puberty are of public health interest. Childhood overweight as a factor in the earlier onset of menarche has been supported by prospective evidence; nonetheless, its overall contribution may have been overemphasized, since secular trends toward a younger age at menarche have not been a universal finding during the recent obesity epidemic. Current observational studies suggest notable associations between dietary intakes and pubertal timing beyond contributions to an energy imbalance: children with the highest intakes of vegetable protein or animal protein experience pubertal onset up to 7 months later or 7 months earlier, respectively. Furthermore, girls with high isoflavone intakes may experience the onset of breast development and peak height velocity approximately 7-8 months later. These effect sizes are on the order of those observed for potentially neuroactive steroid hormones. Thus, dietary patterns characterized by higher intakes of vegetable protein and isoflavones and lower intakes of animal protein may contribute to a lower risk of breast cancer or a lower total mortality., (© 2012 International Life Sciences Institute.)

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.

Background: Longitudinal diet assessment data in children suggest bone anabolic effects of protein intake and concurrent catabolic effects of dietary acid load. However, studies using valid biomarker measurements of corresponding dietary intakes are lacking., Objective: The aim of the study was to examine whether the association of long-term dietary acid load and protein intake with children's bone status can be confirmed using approved urinary biomarkers and whether these diet influences may be independent of potential bone-anabolic sex steroids., Method: Urinary nitrogen (uN), urinary net acid excretion (uNAE), and urinary potential renal acid load (uPRAL) were quantified in 789 24-h urine samples of 197 healthy children who had at least three urine collections during the 4 yr preceding proximal forearm bone analyses by peripheral quantitative computed tomography. uPRAL was determined by subtracting measured mineral cations (sodium + potassium + calcium + magnesium) from measured nonbicarbonate anions (chloride + phosphorus + sulfate). In a subsample of 167 children, dehydroepiandrosterone metabolites were quantified by gas chromatography-mass spectrometry. Multivariable regression models adjusted for age, sex, pubertal stage, forearm muscle area, forearm length, and urinary calcium were run with uN and/or uPRAL or uNAE as predictors., Results: uN was positively associated with bone mineral content, cortical area, periosteal circumference, and strength strain index. uPRAL (but not uNAE) showed negative associations with bone mineral content and cortical area (P < 0.05), both with and without adjustment for the dehydroepiandrosterone-derived sex steroid androstenediol., Conclusions: In line with dietary assessment findings, urinary biomarker analyses substantiate long-term positive effects of protein intake and concomitant negative effects of higher dietary acid load on bone status of children, independent of bone-anabolic sex steroid action.

Context: Whether prepubertal glucocorticoid status impacts on the timing of puberty is not clear., Objective: The objective of the study was to examine the relationship between prepubertal glucocorticoid status and early or late pubertal markers, independent of adrenarchal and nutritional status., Design and Participants: Prospective cohort study of healthy Caucasian children (n = 111, 56 boys) who provided both 24-h urine samples and weighed dietary records 1 and 2 yr before the start of pubertal growth spurt [age at take-off (ATO)]., Measurements: Major urinary glucocorticoid and androgen metabolites determined by gas chromatography-mass spectrometry analysis were summed to assess daily overall cortisol (ΣC21) and adrenal androgen secretion; urinary free cortisol and cortisone measured by RIA were summed (UFF+UFE) as an indicator of potentially bioactive free glucocorticoids., Main Outcomes: The main outcomes included ATO, age at peak height velocity, age at menarche/voice break, ages at Tanner stage 2 for breast (girls) and genital (boys) development, and pubic hair., Results: In girls ΣC21, but not UFF+UFE, was associated with pubertal markers after adjusting for overall adrenal androgen, urinary nitrogen, and body fat. Girls with higher ΣC21 (fourth quartile) reached ATO 0.7 yr (P = 0.01) and menarche 0.9 yr later (P = 0.006) than girls with lower ΣC21 (first quartile). The ΣC21 tended to be also positively associated with age at Tanner stage 2 for breast (P = 0.1), Tanner stage 2 for pubic hair (P = 0.1), and age at peak height velocity (P = 0.06). In boys, neither the ΣC21 nor UFF+UFE was related to pubertal timing., Conclusion: An individually higher prepubertal glucocorticoid secretion level, even in physiological range, appears to delay early and late pubertal timing of healthy girls, particularly their onset of pubertal growth spurt and menarche.

Context: Whether adrenarche impacts on pubertal development is controversial., Objective: The objective of the study was to examine the associations of adrenal androgen (AA) secretion with early and late pubertal markers, independent of potential influences of dietary animal protein intake., Design and Participants: This was a prospective cohort study of healthy free-living Caucasian children (n = 109) who provided both 24-h urine samples and 3-d weighed dietary records 1 and 2 yr before the biological age at take-off of the pubertal growth spurt (ATO)., Measurements: Twenty-four-hour excretion rates of androgen (C19) metabolites quantified by gas chromatography-mass spectrometry were measured., Main Outcomes: ATO, age at peak height velocity (APHV), age at menarche/voice break, duration of pubertal growth acceleration, and ages at Tanner stage 2 for breast (girls) and genital (boys) development (B2-G2) and pubic hair (PH2)., Results: Higher adrenarchal C19 steroids predicted earlier ages at Tanner stage 2 for pubic hair (P < 0.0001) and B2-G2 (P = 0.009) as well as a shorter pubertal growth acceleration period (P = 0.001), independently of animal protein intake. Children with a higher AA secretion had a 1.5-yr earlier beginning of pubarche and a 0.8-yr earlier beginning of B2-G2 than those with a lower AA excretion. Furthermore, animal protein intake was independently negatively associated with ATO and APHV (P < 0.05 each) and tended to be negatively associated with age at menarche/voice break (P = 0.07)., Conclusion: A higher animal protein intake may be involved in an earlier attainment of ATO and APHV, whereas a more intensive adrenarchal process may precipitate a shorter pubertal growth spurt and a notably earlier onset of breast and genital development in girls and boys, respectively.

Context: During the physiological process of adrenarche, the adrenal glands of healthy children secrete increasing amounts of weak androgenic steroids partly metabolized to potent sex steroids., Objective: The aim of the study was to examine whether adrenal androgen metabolite excretion rates before the onset of puberty may be prospectively associated with late-pubertal diaphyseal bone strength., Setting: We conducted the study in an auxological and metabolic child nutrition research facility., Study Population and Design: The sample included 45 healthy adolescents who underwent proximal forearm bone and muscle area measurements by peripheral quantitative computed tomography at the age of 16 yr (SD 1.5) and who had collected a 24-h urine sample 8 yr earlier, allowing to quantify the prepubertal urine metabolome. Prepubertal hormonal predictors quantified by gas chromatography-mass spectrometry were: dehydroepiandrosterone, its 16-hydroxylated downstream metabolites, 5-androstene-3beta,17beta-diol (androstenediol), sums of total androgen and glucocorticoid metabolites, cortisol, and 6beta-hydroxycortisol., Main Outcomes: Proximal forearm radius was measured., Results: Of all prepubertal hormones analyzed, only sex- and age-specific androstenediol levels significantly predicted pubertal stage-, height-, and muscularity-adjusted diaphyseal bone modeling (periosteal circumference, beta = 0.67, P = 0.002; cortical area, beta = 2.15, P = 0.02), bone mineral content (beta = 2.2; P = 0.04), and polar strength strain index (beta = 12.2; P = 0.002). Androstenediol explained 5-10% of the late-pubertal diaphyseal radius variability., Conclusions: Our prospective profiling of urinary steroid metabolites in 24-h urine samples collected before puberty suggests that androstenediol is an early predictor of the diaphyseal bone strength in late puberty. This predominantly peripheral conversion product of adrenarchal dehydroepiandrosterone by 17beta-hydroxysteroid dehydrogenase may hence be involved in a sustained improvement of radial bone accretion during growth.

Context: Very few patients have been described with isolated 17,20-lyase deficiency who have had their mutations in P450c17 (17alpha-hydroxylase/17,20-lyase) proven by DNA sequencing and in vitro characterization of the mutations. Most patients with 17,20-lyase deficiency have mutations in the domain of P450c17 that interact with the electron-donating redox partner, P450 oxidoreductase (POR)., Objective: Our objective was to clarify the genetic and functional basis of isolated 17,20-lyase deficiency in familial cases who were previously reported as having 17,20-lyase deficiency., Patients: Four undervirilized males of an extended Bedouin family were investigated. One of these has previously been reported to carry mutations in the CYP17A1 gene encoding P450c17 causing isolated 17,20-lyase deficiency., Methods: Serum hormones were evaluated before and after stimulation with ACTH. Urinary steroid metabolites were profiled by gas chromatography-mass spectrometry. Exons 1 and 8 of CYP17A1 previously reported to harbor mutations in one of these patients and all 15 coding exons of POR were sequenced., Results: Gas chromatography-mass spectrometry (GC-MS) urinary steroid profiling and serum steroid measurements showed combined deficiencies of 17,20-lyase and 21-hydroxylase. Sequencing of exons 1 and 8 of CYP17A1 in two different laboratories showed no mutations. Sequencing of POR showed that all four patients were homozygous for G539R, a previously studied mutation that retains 46% of normal capacity to support the 17alpha-hydroxylase activity but only 8% of the 17,20-lyase activity of P450c17., Conclusion: POR deficiency can masquerade clinically as isolated 17,20-lyase deficiency.

Context: The CYP17A1 gene encodes many enzymatic reactions including 17alpha-hydroxylase and 17,20-lyase activities. Mutations that selectively ablate the 17,20-lyase activity, causing isolated 17,20-lyase deficiency, are exceedingly rare and may belong to the rarest of all disorders of steroidogenesis. We have previously reported an E305G mutation in the active site of CYP17A1 that apparently causes isolated 17,20-lyase deficiency. Expression studies suggested intact 17alpha-hydroxylase activity which was at odds with subnormal tetracosactrin stimulated cortisol in the patients., Objectives: To investigate the in vivo activity of the adrenal enzymes, we used the metabolomics approach with urinary steroid profiling by gas chromatography-mass spectrometry., Patients: Of the 11 subjects investigated, 6 patients in the kindred were found to be homozygous, 4 members were asymptomatic heterozygous, and 1 was homozygous for the wild-type allele., Results: In the homozygous patients for E305G, both serum and urinary steroids showed a severe lack of androgens (C(19)-steroids) pointing to the absence of 17,20-lyase activities. Furthermore, precursor/product ratios of urinary steroid metabolites characterizing 17alpha-hydroxylase activity showed variable decreases in 17alpha-hydroxylase activities., Conclusions: The results confirm the complete absence of 17,20-lyase activity in vivo, as in the in vitro expression studies. On the other hand, in vivo 17alpha-hydroxylase activity was partially impaired. Thus, the in vivo metabolic data seem to be more sensitive than the expression study and suggests that this mutation also impairs 17alpha-hydroxylase activity.

Objective: The incidence of childhood obesity and type 2 diabetes has reached epidemic proportions. Glucocorticoid excess causes central obesity and diabetes mellitus as seen in Cushing's syndrome. The 11beta-hydroxysteroid dehydrogenase type 1 enzyme (11beta-HSD1) regenerates active cortisol from inactive cortisone. Altered 11beta-HSD1 may cause tissue-specific Cushing's syndrome with central obesity and impaired glucose homeostasis., Design, Patients, and Methods: Clinical and laboratory characteristics, and anthropometric measurements were determined in 15 male and 6 female obese pubertal children (aged 12-18 years, Tanner stages 2-5). In addition, analyses of 24-h excretion rates of glucocorticoids were also performed in 21 age-, sex-, and pubertal stage-matched non-obese children using gas chromatographic-mass spectrometric (GC-MS) analysis., Results: 11beta-HSD1 activity (urinary tetrahydrocortisol (THF) + 5alpha-THF/tetrahydrocortisone (THE) ratio) was lower in obese when compared with non-obese boys. In addition, obese children had a higher total cortisol metabolite excretion than non-obese children. 11beta-HSD1 activity was significantly related to age in lean and obese children. Standard deviation score (SDS)-body mass index did not correlate with 11beta-HSD1 activity, or with total cortisol metabolite excretion within each group. In obese children, 11beta-HSD1 activity and total cortisol metabolite excretion showed no correlation to waist-to-hip ratio, fat mass (percentage of body mass), or the homeostasis model assessment of insulin resistance index., Conclusions: In conclusion, our findings strongly suggest that 11beta-HSD1 activity increases with age, and is reduced in obese boys. In addition, obese children have a higher total cortisol metabolites excretion suggesting a stimulated hypothalamus-pituitary-adrenal axis.

Context: Whereas intrauterine growth and maturation depend on low cortisol levels, an adrenal stress response postnatally is thought to be mandatory in preterm infants., Objective: The goal of this study was to determine cortisol production rates (CPRs) in preterm infants during early life with extreme illness and, thereafter, during extrauterine growth and maturation., Design: We describe a longitudinal observational study., Setting: The study was conducted at a university neonatal intensive care unit., Patients and Methods: Seventeen well (27.9 +/- 1.8 wk) and 44 ill (27.3 +/- 1.6 wk) preterm infants were classified by the Score for Neonatal Acute Physiology. Glucocorticoid metabolites were profiled by gas chromatography-mass spectrometry in 24-h urinary samples. Urine was collected noninvasively using cellulose nappies and extracted by hydraulic press., Results: Medians of CPRs (microg kg(-1) d(-1) mg creatinine) in ill (well) preterm infants were as follows: at d 1, 35 (40); d 2, 35 (40); d 3, 48 (53); d 5, 47 (41); wk 2, 72 (48); wk 3, 73 (37); wk 4, 54 (26). Regression analysis revealed a significant inverse influence of gestational age (P < 0.005) on the maximum of CPRs but not of severity of illness (Score for Neonatal Acute Physiology; P = 0.72). A mature adrenal response was found in only 12 of 44 (27%) ill preterm infants, who had CPRs higher than the 3-fold median of CPRs of well infants. This mature adrenal response was associated with a significantly higher incidence of cerebral bleeding: 9 of 12 (75%) vs. 8 of 32 (25%) without such a response (P = 0.003). During growth, CPRs of ill (well) preterm infants decreased: at month 2, 30 (18); month 3, 18 (22); correlation between weight gain and decrease of CPRs in ill infants between wk 4 and month 3, r = -0.48 (P = 0.027)., Conclusions: Severity of illness did not have a significant influence on CPRs in preterm infants. However, the highest CPRs were associated with a significantly higher incidence of cerebral bleeding. During growth, CPRs decreased significantly, suggesting that preterm infants have the ability to regulate cortisol production. CPRs in ill preterm infants might reflect inadequate stress reaction, but this could also reveal persistence of fetal protective mechanisms against high catabolic cortisol concentrations.

Context: 17alpha-Hydroxylase/17,20-lyase deficiency, a rare autosomal recessive form of congenital adrenal hyperplasia, is caused by mutations in the cytochrome P450c17 (CYP17) gene. We report on a case of complete 17alpha-hydroxylase/17,20-lyase deficiency due to a novel homozygous mutation of CYP17., Design: A 20-yr-old female Turkish patient (46,XX) presented with primary amenorrhea, sexual infantilism, and easy fatigability., Results: The patient's steroid metabolism showed increased levels of mineralocorticoid precursors and low or undetectable plasma concentrations of 17alpha-hydroxycorticoids, androgens, and estrogens before and after ACTH stimulation. The gas chromatography-mass spectrometry urinary steroid profile was dominated by metabolites of corticosterone and its precursors, while cortisol and C(19)-steroid metabolites were lacking. ACTH, FSH, and LH levels were elevated. These hormonal findings were consistent with a combined and total 17alpha-hydroxylase/17,20-lyase deficiency. A therapy with hydrocortisone and a cyclic estrogen/gestagen substitution was initiated., Conclusion: The CYP17 gene analysis revealed homozygosity of the mutation Y27Stop (TAC-->TAA) in exon 1, a mutation that has not been previously described. This novel mutation leads to a stop codon causing a total loss of 17alpha-hydroxlyase/17,20-lyase activity, as reflected biochemically by the detected concentrations of the steroid metabolites.

Information on the urinary excretion of dehydroepiandrosterone (DHEA) and its direct metabolites is scarce for healthy subjects during growth. We used gas chromatography-mass spectrometry urinary steroid profiling to noninvasively study adrenarchal metabolome in 400 healthy subjects, aged 3-18 yr. Urinary 24-h excretion rates of DHEA did not increase significantly before age 7-8 yr. However, DHEA together with its 16alpha-hydroxylated downstream metabolites, 16alpha-hydroxy-DHEA and 3beta,16alpha,17beta-androstenetriol (DHEA&M), as well as the DHEA metabolite, 5-androstene-3beta,17beta-diol (ADIOL), and the sum of major urinary androgen metabolites (C19) rose consistently from the youngest to the oldest age group. The significant increases (P < 0.01) observed for 24-h excretion rates of C19, ADIOL, and DHEA&M were 2- to 4-fold in boys and girls between age 3 and 8 yr. DHEA&M, for example, rose from about 20 to 80 microg/d (P < 0.0001) during this period. Until the age of 16 yr, DHEA&M excretion also increased to nearly 1000 microg/d. Patterns of steroidogenic enzyme activities were assessed (from definite ratios of urinary steroid metabolites) for 21-hydroxylase, 3beta-hydroxysteroid dehydrogenase, 17beta-hydroxysteroid dehydrogenase, and 5alpha-reductase. Our results indicate for healthy boys and girls that adrenarche is a gradual process starting much earlier than hitherto believed. Efficient metabolism of DHEA, especially to 16-hydroxylated steroids, may explain the almost constant levels seen for this steroid until age 7-8 yr. The established reference values for DHEA, DHEA&M, ADIOL, C19 (including androsterone and etiocholanolone), and urinary parameters of steroidogenic enzyme activities could be useful to identify nutritional, environmental, and pathophysiological interrelations with the progressive maturational process of adrenarche. Our data may also be used as reference data for the diagnosis of steroid-related disorders.

Hyperandrogenism, a main clinical feature of polycystic ovary syndrome (PCOS), is thought to result from enhanced ovarian and adrenal androgen generation. To investigate the contribution of peripheral steroidogenesis, we used an oral challenge with dehydroepiandrosterone (DHEA) and analyzed its downstream conversion toward androgens in eight women with PCOS (age, 20-32 yr; body mass index, 20-41 kg/m(2)) and eight healthy women matched for age and body mass index. They underwent frequent serum sampling and urine collection for 8 h on three occasions: at baseline, and after 4 d of dexamethasone (Dex; 4 x 0.5 mg/d), followed by ingestion of 100 mg DHEA or placebo. Dex induced similar significant suppression of circulating steroids in both groups. The oral DHEA challenge led to similar significant increases in the area under the concentration-time curve (0-8 h after Dex) of serum DHEA, DHEA sulfate, androstenedione, and testosterone. However, after oral DHEA, PCOS women had significantly higher increases in serum 5 alpha-dihydrotestosterone (P < 0.01), its main metabolite androstanediol glucuronide (P < 0.05), and the 5 alpha-reduced urinary androgen metabolite androsterone (P < 0.05). PCOS women also had significantly higher baseline excretion of 5 alpha-reduced glucocorticoid (P < 0.01) and mineralocorticoid metabolites (P < 0.05). Taken together, these data indicate enhanced peripheral 5 alpha-reductase activity in PCOS. Thus, not only ovary and adrenal, but also liver and peripheral target tissues, significantly contribute to steroid alterations in PCOS.