Context: Several studies have highlighted the importance of the 11oxygenated 19carbon (11oxC19) adrenalderived steroids as potential biomarkers for monitoring patients with 21hydroxylase deficiency (21OHD).

Objective: To analyze circadian rhythmicity of 11oxC19 steroids in saliva profiles and evaluate their relevance as potential monitoring parameters in 21OHD.

Design, setting, and participants: Cross-sectional single center study including 59 patients with classic 21OHD (men=30; women=29) and 49 BMI- and agematched controls (men=19; women=30).

<strong class="sub-title"> Outcome measures: </strong> Salivary concentrations of the following steroids were analyzed by LCMS/MS: 17hydroxyprogesterone (<em>17OHP</em>), androstenedione (A4), testosterone (T), 11βhydroxyandrostenedione (11OHA4) and 11ketotestosterone (11KT).

<strong class="sub-title"> Results: </strong> Similar to the previously described rhythmicity of <em>17OHP</em>, 11OHA4 and 11KT concentrations followed a distinct diurnal rhythm in both patients and controls with highest concentrations in the early morning and declining throughout the day (11-OHA4: mean reduction of hormone concentrations between timepoint one and five (Δmean) in male patients = 66 %; male controls Δmean = 83 %; female patients Δmean = 47 %; female controls Δmean = 86 %; 11KT: male patients Δmean = 57 %; male controls Δmean = 63 %; female patients Δmean = 50 %; female controls Δmean = 76 %). Significant correlations between the area under the curve (AUC) for <em>17OHP</em> and 11KT (r pmale = 0.773 <0.0001; r pfemale = 0.737 <0.0001), and 11OHA4 (r pmale = 0.633 0.0002; r pfemale = 0.564 0.0014) were observed in patients but not present or reduced in controls.

Conclusions: Adrenal 11oxC19 androgens are secreted following a diurnal pattern. This should be considered when evaluating their utility for monitoring treatment control.

Keywords: 11-ketotestosterone; 11oxygenated androgens; Congenital adrenal hyperplasia; diurnal rhythm; saliva.

BACKGROUND

Reference intervals (RI) of serum 17α- hydroxyprogesterone (<em>17OHP</em>) are useful to confirm congenital adrenal hyperplasia (CAH) in neonates with abnormal screening results and nonclassical forms of CAH in symptomatic children. We aimed to establish serum <em>17OHP</em> RI in normal children and adolescents using a current <em>17OHP</em> radioimmunoassay (RIA).

METHODS

Serum <em>17OHP</em> was measured via a current RIA (Diasource) in children, i.e. 111 infants aged <1 year [before (NE-<em>17OHP</em>) and after extraction (E-<em>17OHP</em>)] and 216 children aged 1-17 years. Forty NE serum samples from subjects aged >1 year, covering the whole analytical range, were simultaneously measured to compare <em>17OHP</em> RIA from Diagnostic System Laboratories (DSL) (withdrawn) and Diasource by Passing Bablok linear regression and ratio plot. The equation obtained was used to correct our own previous RI (DSL RIA) for infancy for the Diasource RIA. Samples from infants aged <1 year were used to verify the calculated RI with evaluator protocol C28-A3. The influence of age, gender, and Tanner's classification (T) was assessed in children aged >1 year by ANOVA.

RESULTS

E-<em>17OHP</em> as measured via the Diasource RIA was significantly lower than NE-<em>17OHP</em> in infants aged <1 year (p < 0.0001). The <em>17OHP</em> measurement from the Diasource RIA was negatively biased compared to the value obtained using the DSL RIA (Diasource (ng/ml) = 0.85 DSL (ng/ml) -0.32 ng/ml, r = 0.952). Most infants (93%) had age- and gender-adjusted NE-<em>17OHP</em> and E-<em>17OHP</em> levels within the recalculated RI. Serum <em>17OHP</em> significantly increased throughout prepuberty (p < 0.001). Sexual dimorphism was only observed at T IV-V.

CONCLUSIONS

When evaluating <em>17OHP</em> during childhood, we recommend taking into account the extraction procedure in neonates, the method used, age, and the Tanner's stage.

<AbstractText>Serum 17α- hydroxyprogesterone (<em>17OHP</em>) and bilateral adrenal sizes are pivotal for clinical practice in both diagnosis and treatment of congenital adrenal disorders during the first month of life. Our aims were to determine the reference ranges for serum <em>17OHP</em> and bilateral adrenal gland sizes according to sex and age groups in healthy term newborns.</AbstractText><AbstractText>A total of 156 healthy newborns, aged 4-7 days (Group 1) or 26-30 days old (Group 2) were included in the study. Serum <em>17OHP</em> concentration was measured in the morning by radioimmunoassay. The right and left adrenal glands' width, length, and depth were measured with ultrasonography by the same radiologist and the volumes were calculated.</AbstractText><AbstractText>The clinical characteristics and serum <em>17OHP</em> concentrations were similar in male and female newborns. Percentiles for serum <em>17OHP</em> concentration and the volume of adrenal glands according to age groups and sexes were obtained. Mean <em>17OHP</em> concentration was 4.67 ± 2.6 ng/ml and 4.49 ± 2.7 ng/ml at the first and fourth week of life, respectively (p > .05). There was a significant decrease in adrenal sizes during the fourth week of life. There was no significant correlation between serum <em>17OHP</em> concentration and adrenal gland sizes.</AbstractText><AbstractText>We have determined reference intervals for serum <em>17OHP</em> concentration and bilateral adrenal gland sizes for healthy newborns. Although serum concentrations of <em>17OHP</em> did not change significantly through the first month of life, our reference intervals for serum <em>17OHP</em> concentration and adrenal sizes may improve clinical approach toward newborns who are suspected of adrenal disorder. We conclude that our reference intervals can guide for congenital adrenal screening regarding serum <em>17OHP</em> concentration besides diagnosis of adrenal hypoplasia or hyperplasia with ultrasonographic adrenal gland sizes.</AbstractText>

17Alpha-hydroxyprogesterone (<em>17OHP</em>) is the most important serum marker for congenital adrenal hyperplasia (CAH). <em>17OHP</em> is usually measured by immunoassay but its detection by mass spectrometry (MS) is a potentially superior method. An LC-MS (liquid chromatography-mass spectrometry) method was developed which utilizes 0.5 ml serum spiked with 6-alpha-methylprednisolone (6-MP) or deuterated <em>17OHP</em> (d8-IS) as the internal standard. The samples were extracted with ether/ethylacetate, and the extract was evaporated to dryness and analysed by LC-MS/MS operating in the positive mode after separation on a reversed-phase C18 column. The calibration curves for analysis of serum <em>17OHP</em> exhibited consistent linearity and reproducibility in the range of 5-250 nmol/l. Interassay CVs were 8.5 and 9.2% at mean concentrations of 7.9 and 23 nmol/l, respectively. The detection limit was 1 nmol/l (signal-to-noise ratio=3). The mean recovery of <em>17OHP</em> added to serum ranged from 76 to 89% and that of internal standards from 75 to 82%. The regression equation for the LC-MS/MS (x) and in-house radioimmunoassay (RIA) (y) methods was: y=0.87x+0.26 (r=0.97; n=100) and for a commercial RIA it was: y=1.32x+0.02 (r=0.97; n=26).

OBJECTIVE

To determine if the diuretic spironolactone cross reacts with 17alpha-hydroxyprogesterone (<em>17OHP</em>) in an enzyme linked immunosorbent assay (ELISA) kit used for the mass screening of congenital adrenal hyperplasia.

METHODS

Concentrations of <em>17OHP</em> on a blood filter paper disc were measured using an ELISA kit (kit C-7: ENZAPLATE N-17alpha -OHP-7; Chiron, Tokyo, Japan). The cross reactivity of spironolactone and its metabolites with <em>17OHP</em> was determined. The concentrations of spironolactone and its metabolites in blood were measured using HPLC (high performance liquid chromatography).

RESULTS

Spironolactone cross reacted with <em>17OHP</em> using kit C-7 (0.01%), by increasing <em>17OHP</em> concentration in a dose dependent manner. The blood concentration of spironolactone and its metabolites was nearly 900 ng/ml, high enough to show an additive effect on the <em>17OHP</em> concentration. About 12% of the false positive cases screened using the kit were due to the administration of spironolactone.

CONCLUSIONS

Spironolactone interferes with <em>17OHP</em> concentrations, leading to false positive test results for CAH.

3 alpha-androstanediol glucuronide (3 alpha diolG) is a marker of peripheral tissue androgen metabolism. There are no previous data regarding complete paediatric reference ranges for 3 alpha diolG. In order to obtain reference values for 3 alpha diolG we have measured serum levels of 3 alpha diolG in 283 healthy children and adolescents, 146 boys and 137 girls, age 1 month to 20 years and 28 adults. A non-extraction, solid phase radioimmunoassay employing a polyclonal antiserum that is specific for 3 alpha diolG was used to measure serum 3 alpha diolG levels (intra assay variation 5.1-10.1%, inter assay variation 2.7-9.0%). There was a strong sex and age dependence (r = 0.8; p < 0.0001) of 3 alpha diolG levels throughout childhood and adolescence with males showing significantly higher levels of the androgen than females (p < 0.05). 3 alpha diolG serum levels (nmol/l +/- SD) correlated significantly with pubertal stage (p < 0.01). Interestingly, in 35 children with CAH serum 3 alpha diolG levels correlated well with clinical and metabolic status, i.e. <em>17OHP</em> serum levels. In summary, we have established percentile curves for 3 alpha diolG levels in healthy children and adolescents. We hypothesize that on the basis of our reference values the single measurement of serum 3 alpha diolG could serve as a means to determine androgen status in children with disorders of puberty and sexual development.

Non-classical congenital adrenal hyperplasia (NC-CAH) includes a group of genetic disorders due to a broad class of CYP21A2 variants identifying a disease-causing 'C' genotype. The heterozygous carriers of CYP21 mutations are at increased risk of developing clinically evident hyperandrogenism, even though clinical and laboratory characteristics are still underestimated. With the aim of obtaining a more accurate delineation of the phenotype of heterozygous carrier of CAH, we analyzed clinical, biochemical and molecular characteristics in a cohort of Sicilian subjects. Fifty-seven females with biallelic and monoallelic CYP21A2 variants classifying NC-CAH (24) and heterozygous carriers of CAH (33), respectively were selected. Forty-four females age-matched healthy controls were also enrolled and genotyped for CYP21A2. Clinical, hormonal and genetic data were collected. CYP21A2 monoallelic mutations, defining the heterozygous carriers state, were identified in subjects with clinical features including hirsutism, oligomenorrhoea, overweight and a PCO-like phenotype, particularly occurring in the age of adolescence. Consistently, levels of <em>17OHP</em> and cortisol were found to be significantly different from NC-CAH. Overall, some clinical and laboratory findings including oligomenorrhea and <em>17OHP</em>/cortisol ratio were observed as independent markers associated with carriers of CAH. Here we report a high prevalence of late-onset signs of polycystic ovary syndrome (PCOS) and hyperandrogenism in heterozygous carriers. The <em>17OHP</em>/cortisol ratio may be a predictive tool to identify the carriers of CAH, even though specific cut-off values have not yet been identified.

Congenital Adrenal Hyperplasias (CAH) are genetic diseases transmitted in an autosomal recessive way and these diseases affect many aspects of human health. The majority of CAH cases is due to a deficiency in 21-hydroxylase as a result of the existence of mutations in both alleles of the CYP21A2 gene. Since the identification of mild, non-classic forms of this disease, CAH has been recognized to be one of the most common genetic diseases in human beings. This disease is generally associated with elevated secretion of androgens, sometimes resulting in virilizing syndromes, including genital ambiguity, precocious puberty in both sexes, or milder syndromes of androgen excess like precocious pubarche or the occurrence of hirsutism and oligomenorrhea in women. Accumulating precursors like 17-hydroxypregnenolone and 17-hydroxyprogesterone (<em>17OHP</em>) are directed to the synthesis of androgens through the enzyme 17-hydroxylase/17,20 lyase leading to the production of dehydroepiandrosterone that is then converted to testosterone and dihydrotestosterone (DHT) at the gonads and at other peripheral tissues. <em>17OHP</em>, the hallmark of 21-hydroxylase deficiency, can be converted to androstenedione (in a low efficiency molecular process) but can also be converted to DHT through an alternative pathway that becomes active due to the large amounts of accumulated <em>17OHP</em> - the backdoor pathway. Another important pathway that becomes significant in this disease is the 11-oxyandrogens pathway through which androstenedione is converted to 11β-hydroxyandrostenedione at the adrenal and from there to 11-ketotestosterone and 11-ketoDHT. The elevated androgens levels affect the hypothalamic-pituitary-gonadal axis and, in some cases, the ovary resulting in chronic anovulation and infertility.

Early diagnosis of congenital adrenal hyperplasia (CAH) can be lifesaving. With the advent of newborn screening programs employing blood 17-hydroxyprogesterone, fewer cases are missed. Because false positive results occur, especially in premature and low birth weight babies, infants with borderline elevations, although requiring follow-up, are often considered normal. We describe a newborn female that, despite severe virilization, only had a borderline elevation in 17-hydroxyprogesterone (<em>17OHP</em>) on newborn screening, as well as on initial confirmatory testing in our clinical laboratory. Our confirmatory method, which employs high performance liquid chromatography (HPLC) separation, because of its high specificity, yields steroid values from both normal children and those with CAH that are lower than found with older, less specific methods. Given the heterogeneity of phenotypes of CAH, less severe forms, especially in males, could result in marginally abnormal laboratory results early in life, with possible adverse effects later. Although in retrospect the diagnosis of the described patient was clear and not a novel entity, we consider it an important example for several reasons. It emphasizes the broad range of <em>17OHP</em> levels in CAH, the lack of correlation of these levels with clinical phenotype and the importance of the timing of both screening and confirmatory tests. Due to the complexity of interpreting these tests, any screening program for CAH should be controlled by an experienced pediatric endocrinologist.

Recent studies have described mild adrenal enzymatic defects in patients presenting with precocious pubarche. In order to identify these defects we have evaluated basal and ACTH- (25 IU iv) stimulated serum adrenal steroid levels in 19 girls, 2- to 8.3-year-old, with precocius pubarche (pubic hair Tanner II-III). Two patients had clitorial enlargement. Bone age was moderatly advanced in 10 patients and 2 to 3.7 yr in four others. Four patients had high basal serum levels of 17-hydroxyprogesterone (<em>17OHP</em>) (525 + 202 ng/dl, mean +SD), compatible with the diagnosis of nonclassical congenital adrenal hyperplasia due to 21-hydroxylase deficiency (NCCAH-21OH), which was confirmed by an increased response of <em>17OHP</em> to ACTH (3425 +/- 953 ng/dl). Fifteen patients had moderately elevated basal <em>17OHP</em> levels (56 + 38 ng/dl) but a normal 170HP response (191 +/- 71 ng/dl) to ACTH, compatible with the diagnosis of idiopathic precocious pubarche (IPP). The cortisol response to ACTH was normal in both groups. Basal values of DHEA-S were 651 +/- 256 and 506 + 462 ng/ml and of DHEA 380 +/- 24 ng/dl and 205 +/- 102 ng/dl, in NCCAH-210H and IPP, respectively. We conclude that: i) clinical findings and baseline levels of DHEA-S and DHEA in IPP can be indistinguishable from the late onset 21 hydroxylase deficiency; ii) baseline levels of <em>17OHP</em> are sufficient for the diagnosis of NCCAH-21OH; iii) the ACTH stimulation test is indicated only when baseline levels of <em>17OHP</em> are moderately elevated (100-300 ng/dl).

OBJECTIVE

To determine the variables affecting serum 17 hydroxyprogesterone (<em>17OHP</em>) in neonates born at a tertiary hospital in Mumbai, India.

METHODS

Serum <em>17OHP</em> was measured in peripheral venous blood between 3rd to 5th day of life by competitive radioimmunoassay and on follow up at 3 mo of age. Serum <em>17OHP</em> was compared among four groups [full term healthy(FT), full term stressed(FS), preterm healthy(PT), preterm stressed(PS)] by non-parametric tests (Kruskal Wallis (KW) test and Mann- Whitney (MW) test). Pearson's test was used to correlate natural log of serum <em>17OHP</em> (ln<em>17OHP</em>) with variables like gestational age, birth weight, stress factor, sex, antenatal administration of glucocorticoids to mothers, Apgar score at 5 min and mode of delivery. Linear regression analysis was done using significant variables in Pearson's test to determine best predictors of ln<em>17OHP</em>.

RESULTS

The initial median (number of cases, inter-quartile range) serum <em>17OHP</em> (ng/ml) for the four groups was as follows; FT 8.4 (33, 6-13); PT 20 (36, 11-29.5); FS 34 (29, 26-45) and PS 58 (24, 40.75-76.5) [total N = 122 newborns, p = 0.001]. Pearson's test showed that gestational age, birth weight, stress factor, Apgar score were negatively correlated with <em>17OHP</em> whereas stress factor, mode of delivery, use of antenatal steroids in mothers were significantly positively correlated. However, stress factor emerged as the most important significant positive predictor (multiple R = 0.643, P = <0.0001). On follow up at 3 mo age, the median <em>17OHP</em> (N = 73 newborns) had significantly decreased to normal level.

CONCLUSIONS

Stress due to neonatal illnesses like meconium aspiration, sepsis, birth asphyxia, etc. significantly elevate serum <em>17OHP</em> and may lead to false positives in newborn screening for congenital adrenal hyperplasia.

BACKGROUND

Basal levels of androgens, in particular 17-hydroxyprogesterone (<em>17OHP</em>), are widely debated as predictors of non-classical congenital adrenal hyperplasia (NCCAH) among patients with precocious pubarche (PP). Many authors have recommended the use of adrenocorticotropic hormone (ACTH) stimulation test in children with PP. The aim of our study was to identify clinical and biochemical predictors of NCCAH in children with PP.

METHODS

We conducted a prospective study of 92 patients with PP undergoing an ACTH stimulation test. We tested the association of basal clinical and biochemical parameters with NCCAH diagnosis. Patients were suspected to have NCCAH if their stimulated <em>17OHP</em> plasma levels were >10 ng/mL. In these patients, the diagnosis was confirmed by genetic test.

RESULTS

Seven (7.6%) patients resulted having NCCAH. The best basal biochemical predictor for NCCAH was <em>17OHP</em> level >2 ng/mL. In fact, a basal <em>17OHP</em> level >2 ng/mL had 100% (95% confidence interval (CI), 59.04-100) sensitivity and 93% (95% CI, 85.3-97.37) specificity. The area under the receiver-operating characteristic curve for <em>17OHP</em> was 0.99 (95% CI, 0.98-1.007).

CONCLUSIONS

Basal <em>17OHP</em> cut-off of 2 ng/mL was very effective in predicting NCCAH among our patients with PP. Assay-specific cut-off would probably be the best strategy to avoid unnecessary ACTH test.

OBJECTIVE

Preterm rupture of membranes (PROM) is associated with an increased risk of preterm birth and neonatal morbidity. Prophylactic 17-hydroxyprogesterone caproate (<em>17OHP</em>-C) reduces the risk of preterm birth in some women who are at risk for preterm birth. We sought to test whether <em>17OHP</em>-C would prolong pregnancy or improve perinatal outcome when given to mothers with preterm rupture of the membranes.

METHODS

This is a multicenter, double-blind, placebo-controlled, randomized clinical trial. The study included singleton pregnancies with gestational ages from 23(0/7) to 30(6/7) weeks at enrollment, documented PROM, and no contraindication to expectant management. Consenting women were assigned randomly to receive weekly intramuscular injections of <em>17OHP</em>-C (250 mg) or placebo. The primary outcome was continuation of pregnancy until a favorable gestational age, which was defined as either 34(0/7) weeks of gestation or documentation of fetal lung maturity at 32(0/7) to 33(6/7) weeks of gestation. The 2 prespecified secondary outcomes were interval from randomization to delivery and composite adverse perinatal outcome. The planned sample size was 222 total women.

RESULTS

From October 2011 to April 2014, 152 women were enrolled; 74 women were allocated randomly to <em>17OHP</em>-C, and 78 were allocated randomly to placebo. The trial was stopped when results of a planned interim analysis suggested that continuation was futile. The primary outcome was achieved in 3% of the <em>17OHP</em>-C group and 8% of the placebo group (P = .18). There was no significant between-group difference in the prespecified secondary outcomes, randomization-to-delivery interval (17.1 ± 16.1 vs 17.0 ± 15.8 days, respectively; P = .76) or composite adverse perinatal outcome (63% vs 61%, respectively; P = .93). No significant differences were found in other outcomes, which included rates of chorioamnionitis, postpartum endometritis, cesarean delivery, individual components of the composite outcome, or prolonged neonatal length of stay.

CONCLUSIONS

Compared with placebo, weekly <em>17OHP</em>-C injections did not prolong pregnancy or reduce perinatal morbidity in patients with PROM in this trial.

Because of the increasing use of 17-hydroxyprogesterone (<em>17OHP</em>) levels with the short adrenocorticotrophic hormone (ACTH) test in the detection of 21-hydroxylase deficiency, the diagnostic efficiency of the test was evaluated in patient and family studies of congenital adrenal hyperplasia due to 21-hydroxylase deficiency and of congenital adrenal hyperplasia due to 11-hydroxylase (11OH) deficiency (the latter disorder now overlaps basally with the milder non-classical 21-hydroxylase deficiency [NC-CAH]). Stimulated 17-hydroxyprogesterone level (<em>17OHP</em>30), 17-hydroxyprogesterone increase (delta 17 OHP) and the ratio 17-hydroxyprogesterone increase to cortisol increase (delta <em>17OHP</em>/delta cortisol) were the parameters from the short ACTH test derived for assessment. 17-OHP30 provided complete differentiation of NC-CAH from the controls and heterozygotes, but overlap between NC-CAH and 11-OH occurred. Complete differentiation of NC-CAH from 11-OH was achieved using delta <em>17OHP</em>. The heterozygotes showed best differentiation from the controls using delta <em>17OHP</em>/delta cortisol with a diagnostic accuracy of 70%, however marked overlap of heterozygotes and NC-CAH with 11-OH was found. The short ACTH test proved to be a valuable technique with the further detection of homozygotes (n = 3) and heterozygotes (n = 5) in the 13 families studied. However, when interpreting the short ACTH test a careful choice of parameters should be made. It should be kept in mind that mild NC-CAH patients can only be differentiated from 11OH patients by using delta <em>17OHP</em>.

Although a weekly injection of 17-hydroxyprogestone caproate is recommended for preventing recurrent preterm birth, clinical experience in North Carolina suggested that many eligible patients were not receiving the intervention.

Our study sought to assess how well practices delivering at 2 major hospitals were doing in providing access to 17-hydroxyprogesterone caproate treatment for eligible patients.

This retrospective cohort analysis studied all deliveries occurring between January 1, 2012, and December 31, 2013, at 2 large hospitals in North Carolina. Women were included if they had a singleton pregnancy and history of a prior spontaneous preterm birth. We extracted demographic, payer, and medical information on each pregnancy, including whether women had been offered, accepted, and received 17-hydroxyprogesterone caproate. Our outcome of 17-hydroxyprogesterone caproate coverage was defined as documentation of ≥1 injection of the drug.

Over the 2-year study period, 1216 women with history of a prior preterm birth delivered at the 2 study hospitals, of which 627 were eligible for 17-hydroxyprogesterone caproate eligible after medical record review. Only 296 of the 627 eligible women (47%; 95% confidence interval, 43-51%) received ≥1 dose of the drug. In multivariable analysis, hospital of delivery, later presentation for prenatal care, fewer prenatal visits, later gestation of prior preterm birth, and having had a term delivery immediately before the index pregnancy were all associated with failed coverage. Among those women who were "covered," the median number of 17-hydroxyprogesterone caproate injections was 9 (interquartile range, 4-15), with 84 of 296 charts (28%) not having complete information on the number of doses.

Even under our liberal definition of coverage, less than half of eligible women received 17-hydroxyprogesterone caproate in this sample. Low overall use suggests that there is opportunity for improvement. Quality improvement strategies, including population-based measurement of 17-hydroxyprogesterone caproate coverage, are needed to fully implement this evidence-based intervention to decrease preterm birth.

OBJECTIVE

The aim of this study was to examine the effects of 17-alpha-hydroxyprogesterone caproate (<em>17OHP</em>-C) on the activity and expression of several common hepatic cytochrome P450 (CYP) enzymes.

METHODS

Primary human hepatocytes were pretreated with vehicle or <em>17OHP</em>-C (0.1 and 1 μmol/L) for 72 hours, then incubated for 1 hour with a cocktail of CYP substrates. The activity of various CYP enzymes was determined by measuring the formation of the metabolites of specific CYP substrates, using liquid chromatography-tandem mass spectrometry. The messenger RNA expression of various CYP enzymes was determined by real-time polymerase chain reaction.

RESULTS

In primary cultures of human hepatocytes, <em>17OHP</em>-C minimally altered the activity or messenger RNA levels of CYP1A2, CYP2C9, CYP2D6, and CYP3A. However, <em>17OHP</em>-C at 1 μmol/L increased CYP2C19 activity by 2.8-fold (P < .01) and CYP2C19 expression by 2.4-fold (P < .001), compared with vehicle-treated cells. A strong positive correlation between activity and expression of CYP2C19 was also observed (r = 0.9, P < .001).

CONCLUSIONS

The activity and expression of hepatic CYP2C19 was significantly increased by <em>17OHP</em>-C in primary cultures of human hepatocytes. This suggests that exposure to medications that are metabolized by CYP2C19 may be decreased in pregnant patients receiving <em>17OHP</em>-C. Metabolism of substrates of CYP1A2, CYP2C9, CYP2D6, and CYP3A are not expected to be altered in patients receiving <em>17OHP</em>-C.

The plasma concentrations of 17 alpha-hydroxyprogesterone (17 alpha OHP) and 17 a'20 alpha-dihydroxy-4-pregnen-3-one (17 alpha 20 alpha OHP) have been measured in sheep during 5 days of ACTH administration at 20 micrograms/kg/day a rate of infusion known to produce hypertension. Five days of ACTH administration produced a progressive increase in plasma <em>17OHP</em> from 0.45 +/- 0.12 to 128.9 +/- 28.4 nmol/l and in 17 alpha 20 alpha OHP from 0.54 +/- 0.15 to 73.1 +/- 7.2 nmol/l. Calculation of the blood production rate of both steroids during ACTH treatment confirms that the rates of infusion of <em>17OHP</em> (3.0 mumol/h) and 17 alpha 20 alpha OHP (1.5 mumol/h) used to produce hypertension, when infused together with the other major ovine adrenocortical steroids, produced plasma concentrations in the range as found following administration at a rate to increase blood pressure.

The positive predictive value of newborn screening for congenital adrenal hyperplasia due to 21-hydroxylase deficiency was <2% in New Zealand. This is despite a bloodspot second-tier immunoassay method for 17-hydroxyprogesterone measurement with an additional solvent extract step to reduce the number of false positive screening tests. We developed a liquid chromatography tandem mass spectrometry (LCMSMS) method to measure 17-hydroxyprogesterone in bloodspots to replace our current second-tier immunoassay method. The method was assessed using reference material and residual samples with a positive newborn screening result. Correlation with the second-tier immunoassay was determined and the method was implemented. Newborn screening performance was assessed by comparing screening metrics 2 years before and 2 years after LCMSMS implementation. Screening data analysis demonstrated the number of false positive screening tests was reduced from 172 to 40 in the 2 years after LCMSMS implementation. The positive predictive value of screening significantly increased from 1.71% to 11.1% (X² test, <i>p</i> < 0.0001). LCMSMS analysis of <em>17OHP</em> as a second-tier test significantly improves screening specificity for CAH due to 21-hydroxylase deficiency in New Zealand.

Keywords: 17-hydroxyprogesterone; congenital adrenal hyperplasia; liquid chromatography tandem mass spectrometry; newborn screening.

Plasma testosterone (T), dihydrotestosterone (DHT), 17-hydroxyprogesterone (<em>17OHP</em>), androstenedione (A), estradiol (E2), and dehydroepiandrosterone sulfate (DHAS) were measured by radioimmunoassay after celite chromatography prior to and after a 3-hour infusion of the synthetic gonadotropin releasing factor, GnRH, in normal prepubertal and pubertal boys. Plasma T levels rose (p less than 0.001) in the pubertal but not prepubertal boys. <em>17OHP</em> concentrations increased in those boys who had an increment of T. A, DHT, E2 or DHAS levels did not increase after GnRH. Basal levels of T, DHT, A and DHAS correlated with the peak and mean serum LH levels attained during the GnRH infusion. These data confirm the greater Leydig cell responsivity to transient rises of endogenous gonadotropin in pubertal males and also suggest that there may be a relationship between adrenal androgen production and maturation of the hypothalamic-pituitary-gonadal system.

Serum sulphates of 5-androstene-3 beta, 17 beta-diol (5-ADIOL-S), 5 alpha-androstane-3 alpha,17 beta-diol (3 alpha-DIOL-S) and dehydroepiandrosterone (DHEA-S), as well as unconjugated androstenedione (AD), testosterone (T) and 17 alpha-hydroxyprogesterone (<em>17OHP</em>), sex hormone binding globulin (SHBG) and the free androgen index (FAI) were measured by specific radioimmunoassay in girls with premature adrenarche (n = 9-16), and in hirsute women with (1) late onset 21 hydroxylase deficiency (n = 14), (2) polycystic ovarian disease (n = 28) and (3) idiopathic hirsutism (n = 74). Levels were also determined in females with androgenic alopecia (n = 35-45), in normal prepubertal girls (n = 9-14) and in normal adult women (n = 50-73). Mean serum concentrations of 5-ADIOL-S, 3 alpha-DIOL-S, DHEA-S, AD, T, and FAI were elevated and SHBG depressed, in all patient groups compared with controls, except for DHEA-S and T in patients with alopecia. We conclude that in addition to DHEA-S, 5-ADIOL-S may have a role as a pro-hormone in the synthesis of more potent androgens (T, DHT) in peripheral tissues such as skin; in addition, 3 alpha-DIOL-S may be a marker of peripheral androgen metabolism.

The influence of aging upon serum concentrations of testicular steroids, sex hormone binding globulin (SHBG) and pituitary hormones and on adrenal steroid levels and adrenal steroid response to ACTH was studied in 81 healthy men aged 20-87 years. These endocrine variables were also compared in 43 patients with benign prostatic hyperplasia (BPH), aged 58-89 years and in a subgroup of 41 men, aged 58-87 years, from the above mentioned reference population. The normal endocrine aging was characterized by a rise in SHBG levels, decreasing levels of testicular steroids and non-SHBG-bound testosterone (NST) and increasing gonadotropin levels and decreasing concentrations of total estrone. Adrenal androgen levels decreased in the presence of unchanged levels of cortisol and the adrenal steroid response to ACTH changed by decreasing increments in dehydroepiandrosterone (DHA) and increasing increments in 17 alpha-hydroxyprogesterone (<em>17OHP</em>). With the exception of the alterations in SHBG and adrenal androgens, all these changes were finished before the seventh decade of life. BPH patients had elevated levels of testosterone and NST in the presence of normal SHBG and gonadotropin levels, elevated levels of DHA and DHA sulfate (DHAS) in the presence of normal cortisol levels, a "younger" pattern of adrenal steroid response to ACTH as judged from the increments in DHA and <em>17OHP</em>, elevated ratios between estrone and 4-androstene-3,17-dione suggesting an increased peripheral aromatization and subnormal prolactin levels. BPH patients may be considered as "endocrinologically younger" than healthy subjects. DHA and especially its proximate metabolite 5-androstene-3 beta, 17 beta-diol exert powerful estrogenic effects on the receptor level. Thus the elevated levels of DHA and DHAS in the BPH patients may create an hyperestrogenic condition in addition to the slight hyperandrogenicity caused by the elevated NST levels. Both endocrine aberrations may play a role in the etiology of BPH, in accordance with the dual sex steroid sensitivity of the periurethral glands.

The case of a 62-year-old woman with severe post-menopausal hirsutism is described. Her clinical history revealed regular menstrual periods until menopause at the age of 50, hysterectomy for fibromatosis at 58 years, non-insulin dependent diabetes mellitus, hypertension, obesity, severe hirsutism, which had developed in the previous 3 years, with a deeping of the voice. Examination showed android obesity, hypertension and severe hirsutism involving the face and the trunk. Endocrine evaluation pointed out regular adrenal function, serum total and free-testosterone in the adult male range, with normal androstenedione, DHEAS and <em>17OHP</em> levels. Estradiol was slightly increased and LH and FSH were inappropriately low for her post-menopausal age. Computed tomography of the abdomen showed regular adrenal glands, and a radio-labeled cholesterol scan was negative. A further pelvic transvaginal ultrasonography revealed a small cystic formation near the right ovary and a slight increase in the size of the left ovary. The patient underwent bilateral ovariectomy. Histological examination showed a lipoid cell tumor within the left ovary. Immunohistochemical studies were positive for inhibin and cytokeratin. After surgery, serum testosterone fell to normal levels, gonadotropins increased to menopausal levels, confirming that the tumor was able to produce both LH, and FSH-inhibiting factors, and hirsutism greatly improved. Periodic hormonal tests remained normal and CT of the abdomen and pelvic ultrasonography did not show alterations at a 3 years follow-up.

Hyperandrogenic manifestation in women, such as seborrhea, acne and increased hair growth are common reasons of psychological distress. Skin appearance is very important for young women. This study evaluated the hormonal and skin effects of two estroprogestins (EPs) containing ethinyl-estradiol (EE) 30 microg associated with drospirenone (DRSP) 3 mg or chlormadinone acetate (CMA) 2 mg, respectively. Fifty-five women with signs and symptoms of hyperandrogenism (seborrhea, acne and increased hair growth) were enrolled in the study; randomly, 30 women were treated with EE 30 microg + DRSP 3 mg and 25 with EE 30 microg + CMA 2 mg. Follicle-stimulating hormone (FSH), luteinising hormone (LH), 17-hydroxyprogesterone (<em>17OHP</em>), androstenedione (A), testosterone (T), dehydroepiandrosterone sulfate (DHEAS), sex hormone binding globulin (SHBG) and free androgen index (T x 100/SHBG, FAI) were assessed at baseline, and after 3 and 6 months of treatment with EPs. Effects on seborrhea, acne and increased hair growth (as Ferriman-Gallwey score) were also evaluated at the same time points. Finally, skin hydration, transepidermal water loss (TEWL) and skin homogeneity were studied with non-invasive technique during the study. Treatment for 6 months with both EPs decreased significantly the circulating androgen levels (A, T, DHEAS) and FAI, and increased SHBG levels; also skin pattern was improved. EP containing EE and DRSP was better than EP containing EE and CMA as for skin changes, as seborrhea, acne, increased hair, hydration, homogeneity and overall quality of the skin; moreover, hormonal changes (as FAI) under therapy were more pronounced with EE/DRSP than EE/CMA. These effects may be considered in EP choice and could be important in improving patient's compliance and quality of life in hyperandrogenic women.

OBJECTIVE

To explore the clinical and genetic characteristics of two patients with 11 β-hydroxylase deficiency (11 β-OHD).

METHODS

The clinical features and laboratory data were collected from the patients and their families. All exons of CYP11B1 gene were amplified by PCR. And the PCR product sequences were identified by a DNA analyzer.

RESULTS

Two patients presented with juvenile hypertension with bilateral adrenal hyperplasia and congenital hypospadias, hypertension for 17 years and periodic hematuria for 3 months after dexamethasone therapy respectively. Steroid analysis showed the typical pattern of 11 β-OHD: elevated plasma levels of adrenocorticotropic hormone (ACTH), 17-hydroxyprogesterone (<em>17OHP</em>), 11-deoxycortisol, androstenedione and testosterone and lowered levels of potassium, aldosterone and plasma renin activity (PRA). CT scan revealed the presence of bilateral nodular hyperplasia of adrenal glands. Sequencing analysis showed compound heterozygous mutations of [R453Q]+[R454C] at exon 8 in patient 1 and homozygous mutation of [R454C] at exon 8 in patient 2.

CONCLUSIONS

11 β-OHD is the second major cause of congenital adrenal hyperplasia. The classic characteristics are hypertension with low a level of PRA, hypokalemia, female pseudohermaphroditism and male sexual precocity. 11 β-OHD should be screened in the patients with juvenile onset hypertension accompanied by bilateral adrenal hyperplasia.