Cortisol and 17-hydroxyprogesterone levels in saliva of healthy neonates - Normative data and relation to body mass index, arterial cord blood pH and time of sampling after birth

The measurement of cortisol and 17-hydroxyprogesterone (17-OHP) in saliva has become a reliable tool for both the scientist and the clinician for studying adrenal cortical function in the adult and the older child. We have now established in parallel normative data for salivary cortisol and 17-OHP levels in healthy neonates. We have asked whether or not there is a circadian rhythm of cortisol and 17-OHP saliva levels in neonates. Furthermore, we have asked whether salivary hormone levels correlated with auxologic and clinical data and time of sampling. Cortisol and 17-OHP levels in saliva samples from 119 healthy neonates (55 girls, 64 boys) were measured using in-house time-resolved fluorescent immunoassays. Saliva samples were obtained using a saliva collecting tube three times a day on the first or second day of life. Gender and gestational age did not influence salivary cortisol and 17-OHP levels. No significant circadian rhythm of salivary hormone levels was detected in this group of newborns. However, body mass index, arterial cord blood pH and time of saliva sampling significantly influenced salivary hormone levels. In conclusion, measurement of cortisol and 17-OHP in saliva is feasible in healthy neonates. The existence of normative data forms the basis for future studies on pathophysiologic states in the newborn period. Copyright (C) 2000 S. Karger AG, Basel

Studies on Regioselective Binding Mode of Steroid Molecules in Homology Modeled Cytochrome P450-2C11

In this study, we investigated the regioselective binding mode of steroid molecules and structure requirements for steroid molecules for 16[alpha]-hydroxylation by Cytochrome P450-2C11. Docking study by using the homology Cytochrome P450-2C11 indicated that 16[alpha]-hydroxylation is favored with steroidal molecules possessing the following components, 1) a bent A-B ring configuration (5[beta]-reduced), 2) C-3[alpha]-hydroxyl group, 3) C-17[beta]-acetyl group, and 4) methyl group at both the C-18 and C-19. These respective steroid components requirements such as A-B ring configuration and functional groups at C-3 and C-17 were defined as the inhibitory contribution factor. Overall results by rat CYP2C11 revealed that steroidal structure requirements resulted in causing an effective inhibition of [^3^H]progesterone 16[alpha]-hydroxylation by the adult male rat liver microsome. As far as docking of homology modeled CYP2C11 against investigated steroids is concerned, they are docked at the active site superimposed with flurbiprofen. It was also found that the distance between heme iron and C16[alpha]-H was between 4 to 6 Å and that the related angle was in the range of 180±45°

Individual 17-Hydroxyprogesterone Responses to hCG Are Not Correlated With Follicle Size in Polycystic Ovary Syndrome.

Context:In women with polycystic ovary syndrome (PCOS), 17-hydroxyprogesterone (17-OHP) responses to gonadotropin stimulation vary from increased to indistinguishable compared with normal controls. Objective:To determine whether 17-OHP responses to recombinant-human chorionic gonadotropin (r-hCG) are individually correlated to the size of antral follicles among women with PCOS. Design Setting and Participants:A prospective study conducted in 19 women with PCOS and 20 normal controls at an academic medical center. Interventions:Blood samples were obtained before and 24 hours after administration of 25 μg of r-hCG. Ovarian imaging was conducted with three-dimensional pelvic ultrasonography. Each subject underwent a 2-hour oral glucose tolerance test. Main Outcome Measures:Basal and stimulated levels of 17-OHP, androgens, estradiol, progesterone, anti-Mullerian hormone (AMH), insulin, glucose, follicle number, and size. Results:In women with PCOS, mean antral follicle count (AFC) was greater than that of controls, although the size of cohort follicles within individual subjects was not correlated to 17-OHP responses. The numbers of 2- to 3-mm and 3- to 4-mm follicles in PCOS were significantly greater than in controls, whereas differences between larger follicles were not observed. Increased AMH in PCOS was correlated to AFC, but not 17-OHP responses. Insulin sensitivity did not correlate to r-hCG‒stimulated 17-OHP after adjustment for body mass index. Conclusions:17-OHP responses to hCG in individuals with PCOS were not correlated to the distribution of antral follicles. Greater numbers of small antral follicles in women with PCOS than in controls suggest an extension of accelerated growth from the preantral stage

Gestational dating by metabolic profile at birth: a California cohort study.

BackgroundAccurate gestational dating is a critical component of obstetric and newborn care. In the absence of early ultrasound, many clinicians rely on less accurate measures, such as last menstrual period or symphysis-fundal height during pregnancy, or Dubowitz scoring or the Ballard (or New Ballard) method at birth. These measures often underestimate or overestimate gestational age and can lead to misclassification of babies as born preterm, which has both short- and long-term clinical care and public health implications.ObjectiveWe sought to evaluate whether metabolic markers in newborns measured as part of routine screening for treatable inborn errors of metabolism can be used to develop a population-level metabolic gestational dating algorithm that is robust despite intrauterine growth restriction and can be used when fetal ultrasound dating is not available. We focused specifically on the ability of these markers to differentiate preterm births (PTBs) (<37 weeks) from term births and to assign a specific gestational age in the PTB group.Study designWe evaluated a cohort of 729,503 singleton newborns with a California birth in 2005 through 2011 who had routine newborn metabolic screening and fetal ultrasound dating at 11-20 weeks' gestation. Using training and testing subsets (divided in a ratio of 3:1) we evaluated the association among PTB, target newborn characteristics, acylcarnitines, amino acids, thyroid-stimulating hormone, 17-hydroxyprogesterone, and galactose-1-phosphate-uridyl-transferase. We used multivariate backward stepwise regression to test for associations and linear discriminate analyses to create a linear function for PTB and to assign a specific week of gestation. We used sensitivity, specificity, and positive predictive value to evaluate the performance of linear functions.ResultsAlong with birthweight and infant age at test, we included 35 of the 51 metabolic markers measured in the final multivariate model comparing PTBs and term births. Using a linear discriminate analyses-derived linear function, we were able to sort PTBs and term births accurately with sensitivities and specificities of ≥95% in both the training and testing subsets. Assignment of a specific week of gestation in those identified as PTBs resulted in the correct assignment of week ±2 weeks in 89.8% of all newborns in the training and 91.7% of those in the testing subset. When PTB rates were modeled using the metabolic dating algorithm compared to fetal ultrasound, PTB rates were 7.15% vs 6.11% in the training subset and 7.31% vs 6.25% in the testing subset.ConclusionWhen considered in combination with birthweight and hours of age at test, metabolic profile evaluated within 8 days of birth appears to be a useful measure of PTB and, among those born preterm, of specific week of gestation ±2 weeks. Dating by metabolic profile may be useful in instances where there is no fetal ultrasound due to lack of availability or late entry into care

Growth Patterns in the First Three Years of Life in Children with Classical Congenital Adrenal Hyperplasia Diagnosed by Newborn Screening and Treated with Low Doses of Hydrocortisone

Background: Linear growth is the best clinical parameter for monitoring metabolic control in classical congenital adrenal hyperplasia (CAH). Objective: To analyze growth patterns in children with CAH diagnosed by newborn screening and treated with relatively low doses of hydrocortisone during the first year of life. Patients and Methods: 51 patients (27 females) were diagnosed with classical CAH by newborn screening. All patients were treated with relatively low doses of hydrocortisone (9-15 mg/m(2) body surface area). 47 patients were additionally treated with fludrocortisone. Results: At birth, height SDS (H-SDS) was 1.1 +/- 1 in girls and 0.9 +/- 1.5 in boys. After 3 months, H-SDS decreased to 0.4 +/- 0.9 in girls and to 0.1 +/- 1.3 in boys. Over the 3-year period, H-SDS further decreased to -0.4 +/- 1.8 in girls and to -0.8 +/- 1 in boys and approached the genetic height potential (target H-SDS of girls -0.5 +/- 0.3 and target H-SDS of boys -0.9 +/- 0.7). During the first 9 months of age, growth velocity was slightly decreased in girls (18.2 +/- 1.9 cm) and boys (17.3 +/- 1.6 cm) when compared to a healthy reference population (girls 19.0 +/- 3.9 cm and boys 18.7 +/- 4.7 cm). At the age of 3 years, bone age was appropriate for chronological age in both girls (2.7 +/- 0.5 years) and boys (2.9 +/- 0.5 years). Conclusion: Birth length is above average in children with classical CAH, which might be the result of untreated hyperandrogenism in utero. With relatively low doses of hydrocortisone treatment, growth velocity decreases slightly during the first 9 months and H-SDS then approaches the genetic height potential. Copyright (C) 2010 S. Karger AG, Base

إنتاج مادة 17 ألفا هيدروكسي البروجستيرون على مستوى المخمر المعملي بواسطة فطرة كاننجهاميلا إيكينولاتا

The mircrobiological transformation of progesterone by a local isolate of Cunninghamella echiiiulata using a laboratory fermentor was studied. Progresterone (10-50 g/1) wetted by Tween 80 was added to 48-hour old culture and the transformation was left to proceed for 72 hours. Thereafter, the different transformation products were resolved chromatog-raphically. The identity of each product was established through the determination of m.p., mixed m.p., optical rotation and ultraviolet as well as infrared absorption spectra. A comparison of the R{ values of each product with that of the corresponding reference using different solvent systems as well as their colour expressed with two spray reagents, was used as a further proof for the identity of the isolated products. With all concentrations of progesterone tested, maximum yield of 17ot -hydroxyprogesterone was obtained after 48 hours of fermentation Progesterone concentrations of 10 and 20 g/1 were almost quantitatively converted to the different transformation products after 72 hours of fermentation. Using a concentration of 20 g/1 and incubation period of 48 hours, the transformation product mixture consisted of unchanged progesterone (6%), 17 o< -hydroxyprogesterone (54%),llotrhydroxyprogesterone (29%) and llo<;,17<^-dihydroxy-progesterone (2.5%).تم استخدام مخمر صناعي سعة 2 لتر لاختيار مقدرة الفطرة على تكوين هذه المادة في ظروف تشبه تلك المطبقة في الصناعة . وبدراسة تركيزات متعددة فن مادة البروجستيرون تتراوح ما بين 10جرام /لتر إلى 50جرام /لتر ، وجد أن أنسب التركيزات المختبرة هو تركيز 20 جرام من البروجستيرون لكل لترمن الوسط الغذائي ، حيث تم تحويل كل البووجستيرون المضاف إلى المشتقات المختلفة خلال 72 ساعة من بدء الاضافة . ووجد أن أعلى معدل لتكوين مادة 17 ألفا - هيدروكسي البروجستيرون كان بعد 48 ساعة من بدأ إضافة البروجستيرون . عند فصل المواد الناتجة من تحول البروجستيرون بواسطة الفطرة المستخدمة وذلك بواسطة أعمدة الفصل باستخدام مادة الالومينا وجد أن البروجستيرون يتحول إلى : 17 ألفا - هيدروكسي البروجستيرون ( 54 %) 11 ألفا - هيدروكسي البروجستيرون (29%) 11 ألفا ، 17 ألفا - ثنائي هيدروكسي البروجستيرون (2.5%

Variations of Steroid Hormone Metabolites in Serum and Urine in Polycystic Ovary Syndrome after Nafarelin Stimulation: Evidence for an Altered Corticoid Excretion.

To evaluate the clinical relevance of testing pituitary-ovarian responses in patients suffering from polycystic ovary syndrome (PCOS) with the GnRH agonist nafarelin, a 1.2-mg dose of nafarelin was given intranasally to 19 women with PCOS and 15 healthy premenopausal women. The subsequent analysis of steroids in both serum and urine during the test was carried out at several time points for up to 24 h. Serum levels of 17 alpha-hydroxyprogesterone were elevated at all time points of the test in PCOS patients vs. controls [at baseline, 3.5 +/- 0.2 vs. 1.8 +/- 0.1 nmol/L (P < 0.001); at 24 h, 9.9 +/- 0.9 vs. 4.9 +/- 0.3 nmol/L (P < 0.001)]. Basal levels of androstenedione were higher in the patient group, but there was no significant change during the test in either group. Serum testosterone levels were also found to differ in PCOS patients compared with the control values at baseline (2.2 +/- 0.2 vs. 1.5 +/- 0.1 nmol/L; P < 0.05) and after nafarelin treatment (at 24 h, 3.2 +/- 0.4 vs. 1.8 +/- 0.2 nmol/L; P < 0.05). Serum estradiol levels rose significantly in both groups during the test; the posttest levels were significantly higher in PCOS than in controls. The PCOS patients displayed a significant increase in androgen and gestagen metabolites as well as in glucocorticoid metabolites excreted in the urine during the 24 h. In the control subjects, except for 17 alpha-hydroxypregnanolone, which rose significantly, none of the urinary steroids investigated showed relevant changes during the nafarelin test. The posttest excretion of allo-tetrahydrocortisol (1.4 +/- 0.2 vs. 0.3 +/- 0.1 mumol/g creatinine; P < 0.001) and the increase in 17 alpha-hydroxypregnanolone excretion (1.4 +/- 0.2 vs. 0.3 +/- 0.1 mumol/g creatinine; P < 0.001) were distinctly higher in PCOS patients than in the controls; the diagnostic sensitivity of the combination of both parameters was 89% at a 93% specificity. Thus, measurements of 17 alpha-hydroxyprogesterone levels in serum and of urinary allo-tetrahydrocortisol and 17 alpha-hydroxypregnanolone after nafarelin treatment make this stimulation test a valuable diagnostic tool for identifying PCOS patients. The significant changes in the excretion of urinary androgen and gestagen metabolites, unmasked by GnRH agonist stimulation, suggest a functional alteration of the pituitary-ovarian axis. The reason for the increased excretion of glucocorticoid metabolites after nafarelin stimulation remains to be clarified

Obstetrics and gynaecology

A review of publications relating to significant advances in the specialty of Obstetrics and Gynaecology over the past four years will be discussed: topics reviewed will have an important impact on reducing maternal/fetal morbidity and mortality and should improve on woman's health care.peer-reviewe

Homozygous disruption of P450 side-chain cleavage (CYP11A1) is associated with prematurity, complete 46,XY sex reversal, and severe adrenal failure

Disruption of the P450 side-chain cleavage cytochrome (P450scc) enzyme due to deleterious mutations of the CYP11A1 gene is thought to be incompatible with fetal survival because of impaired progesterone production by the fetoplacental unit. We present a 46, XY patient with a homozygous disruption of CYP11A1.The child was born prematurely with complete sex reversal and severe adrenal insufficiency. Laboratory data showed diminished or absent steroidogenesis in all pathways. Molecular genetic analysis of the CYP11A1 gene revealed a homozygous single nucleotide deletion leading to a premature termination at codon position 288. This mutation will delete highly conserved regions of the P450scc enzyme and thus is predicted to lead to a nonfunctional protein. Both healthy parents were heterozygous for this mutation.Our report demonstrates that severe disruption of P450scc can be compatible with survival in rare instances. Furthermore, defects in this enzyme are inherited in an autosomal-recessive fashion, and heterozygote carriers can be healthy and fertile. The possibility of P450scc-independent pathways of steroid synthesis in addition to the current concept of luteoplacental shift of progesterone synthesis in humans has to be questioned

Human P450 CYP17A1: Control of Substrate Preference by Asparagine 202

CYP17A1 is a key steroidogenic enzyme known to conduct several distinct chemical transformations on multiple substrates. In its hydroxylase activity, this enzyme adds a hydroxyl group at the 17α position of both pregnenolone and progesterone at approximately equal rates. However, the subsequent 17,20 carbon–carbon scission reaction displays variable substrate specificity in the numerous CYP17A1 isozymes operating in vertebrates, manifesting as different Kd and kcat values when presented with 17α-hydroxypregnenlone (OHPREG) versus 17α-hydroxyprogesterone (OHPROG). Here we show that the identity of the residue at position 202 in human CYP17A1, thought to form a hydrogen bond with the A-ring alcohol substituent on the pregnene- nucleus, is a key driver of this enzyme’s native preference for OHPREG. Replacement of asparagine 202 with serine completely reverses the preference of CYP17A1, more than doubling the rate of turnover of the OHPROG to androstenedione reaction and substantially decreasing the rate of formation of dehydroepiandrosterone from OHPREG. In a series of resonance Raman experiments, it was observed that, in contrast with the case for the wild-type protein, in the mutant the 17α alcohol of OHPROG tends to form a H-bond with the proximal rather than terminal oxygen of the oxy–ferrous complex. When OHPREG was a substrate, the mutant enzyme was found to have a H-bonding interaction with the proximal oxygen that is substantially weaker than that of the wild type. These results demonstrate that a single-point mutation in the active site pocket of CYP17A1, even when far from the heme, has profound effects on steroidogenic selectivity in androgen biosynthesis