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Cyclooxygenase (COX)-derived prostaglandins stimulate uterine contractions and prepare the cervix for parturition. Prior reports suggest Cox-1 knockout (KO) mice exhibit delayed parturition due to impaired luteolysis, yet the mechanism for late-onset delivery remains unclear. Here, we examined key factors for normal onset of parturition to determine whether any could account for the delayed parturition phenotype. Pregnant Cox-1KO mice did not display altered timing of embryo implantation or postimplantation growth. Although messenger RNAs of contraction-associated proteins (CAPs) were differentially expressed between Cox-1KO and wild-type (WT) myometrium, there were no differences in CAP agonist-induced intracellular calcium release, spontaneous or oxytocin (OT)-induced ex vivo uterine contractility, or in vivo uterine contractile pressure. Delayed parturition in Cox-1KO mice persisted despite exogenous OT treatment. Progesterone (P4) withdrawal, by ovariectomy or administration of the P4-antagonist RU486, diminished the delayed parturition phenotype of Cox-1KO mice. Because antepartum P4 levels do not decline in Cox-1KO females, P4-treated WT mice were examined for the effect of this hormone on in vivo uterine contractility and ex vivo cervical dilation. P4-treated WT mice had delayed parturition but normal uterine contractility. Cervical distensibility was decreased in Cox-1KO mice on the day of expected delivery and reduced in WT mice with long-term P4 treatment. Collectively, these findings show that delayed parturition in Cox-1KO mice is the result of impaired luteolysis and cervical dilation, despite the presence of strong uterine contractions.
Copyright © 2018 Endocrine Society.
Cytochrome P450 (P450, CYP) 17A1 plays a critical role in steroid metabolism, catalyzing both the 17α-hydroxylation of pregnenolone and progesterone and the subsequent 17α,20-lyase reactions to form dehydroepiandrosterone (DHEA) and androstenedione (Andro), respectively, critical for generating glucocorticoids and androgens. Human P450 17A1 reaction rates examined are enhanced by the accessory protein cytochrome (), but the exact role of in P450 17A1-catalyzed reactions is unclear as are several details of these reactions. Here, we examined in detail the processivity of the 17α-hydroxylation and lyase steps. did not enhance reaction rates by decreasing the rates of any of the steroids. Steroid binding to P450 17A1 was more complex than a simple two-state system. Pre-steady-state experiments indicated lag phases for Andro production from progesterone and for DHEA from pregnenolone, indicating a distributive character of the enzyme. However, we observed processivity in pregnenolone/DHEA pulse-chase experiments. ()-Orteronel was three times more inhibitory toward the conversion of 17α-hydroxypregnenolone to DHEA than toward the 17α-hydroxylation of pregnenolone. IC values for ()-orteronel were identical for blocking DHEA formation from pregnenolone and for 17α-hydroxylation, suggestive of processivity. Global kinetic modeling helped assign sets of rate constants for individual or groups of reactions, indicating that human P450 17A1 is an inherently distributive enzyme but that some processivity is present, some of the 17α-OH pregnenolone formed from pregnenolone did not dissociate from P450 17A1 before conversion to DHEA. Our results also suggest multiple conformations of P450 17A1, as previously proposed on the basis of NMR spectroscopy and X-ray crystallography.
© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.
Cytochrome P450 (P450) reactions can involve C-C bond cleavage, and several of these are critical in steroid and sterol biosynthesis. The mechanisms of P450s 11A1, 17A1, 19A1, and 51A1 have been controversial, in the context of the role of ferric peroxide (FeO2 (-)) versus perferryl (FeO(3+), compound I) chemistry. We reinvestigated the 17α-hydroxyprogesterone and 17α-hydroxypregnenolone 17α,20-lyase reactions of human P450 17A1 and found incorporation of one (18)O atom (from (18)O2) into acetic acid, consonant with proposals for a ferric peroxide mechanism (Akhtar, M., Lee-Robichaud, P., Akhtar, M. E., and Wright, J. N. (1997) J. Steroid Biochem. Mol. Biol. 61, 127-132; Akhtar, M., Wright, J. N., and Lee-Robichaud, P. (2011) J. Steroid Biochem. Mol. Biol. 125, 2-12). However, the reactions were supported by iodosylbenzene (a precursor of the FeO(3+) species) but not by H2O2 We propose three mechanisms that can involve the FeO(3+) entity and that explain the (18)O label in the acetic acid, two involving the intermediacy of an acetyl radical and one a steroid 17,20-dioxetane. P450 17A1 was found to perform 16-hydroxylation reactions on its 17α-hydroxylated products to yield 16,17α-dihydroxypregnenolone and progesterone, suggesting the presence of an active perferryloxo active species of P450 17A1 when its lyase substrate is bound. The 6β-hydroxylation of 16α,17α-dihydroxyprogesterone and the oxidation of both 16α,17α-dihydroxyprogesterone and 16α,17α-dihydroxypregnenolone to 16-hydroxy lyase products were also observed. We provide evidence for the contribution of a compound I mechanism, although contribution of a ferric peroxide pathway in the 17α,20-lyase reaction cannot be excluded.
© 2016 by The American Society for Biochemistry and Molecular Biology, Inc.
Soy food intake has previously been associated with reduced breast cancer risk. Epidemiological evidence for subgroups of breast cancer, particularly by menopausal and hormone receptor status, is less consistent. To evaluate the role of hormone receptor and menopausal status on the association between soy food intake and breast cancer risk, we measured usual soy food intake in adolescence and adulthood via food frequency questionnaire in 70,578 Chinese women, aged 40-70 years, recruited to the Shanghai Women's Health Study (1996-2000). After a median follow-up of 13.2 years (range: 0.01-15.0), 1,034 incident breast cancer cases were identified. Using Cox models, we found that adult soy intake was inversely associated with breast cancer risk [hazard ratio (HR) for fifth versus first quintile soy protein intake = 0.78; 95% confidence interval (CI):0.63-0.97]. The association was predominantly seen in premenopausal women (HR = 0.46; 95% CI:0.29-0.74). Analyses further stratified by hormone receptor status showed that adult soy intake was associated with significantly decreased risk of estrogen receptor (ER)+/progesterone receptor (PR)+ breast cancer in postmenopausal women (HR = 0.72; 95% CI:0.53-0.96) and decreased risk of ER-/PR- breast cancer in premenopausal women (HR = 0.46; 95% CI:0.22-0.97). The soy association did not vary by human epidermal growth factor-2 (HER2) status. Furthermore, we found that high soy intake during adulthood and adolescence was associated with reduced premenopausal breast cancer risk (HR = 0.53; 95% CI: 0.32-0.88; comparing third vs. first tertile) while high adulthood soy intake was associated with postmenopausal breast cancer only when adolescent intake was low (HR = 0.63; 95% CI: 0.43-0.91). Our study suggests that hormonal status, menopausal status and time window of exposure are important factors influencing the soy-breast cancer association.
© 2016 UICC.
Sickle cell anemia is one of the most common genetic blood disorders worldwide. Individuals with sickle cell disease (SCD) experience clinical manifestations such as chronic anemia, developmental delay, vaso-occlusive pain, acute chest syndrome, and neurological complications. Adolescent girls with SCD face unique gynecological challenges including delayed puberty marked by a later onset in menarche, vaso-occlusive pain associated with their menstrual cycle, and underdiagnosed abnormal uterine bleeding. This review focuses on these challenges with particular emphasis on delayed menarche and vaso-occlusive pain episodes associated with menstruation, in addition to the evaluation and initial management of heavy menstrual bleeding for adolescents with SCD. We highlight research opportunities in this neglected area to help enhance the comprehensive care model for this population.
Cytochrome P450 21A2 is a key player in steroid 21-hydroxylation and converts progesterone to 11-deoxycorticosterone and 17α-hydroxy progesterone to 11-deoxycortisol. More than 100 mutations in P450 21A2 have been established in patients thus far; these account for the vast majority of occurrences of congenital adrenal hyperplasia (CAH), which is among the most common heritable metabolic diseases in humans. CAH phenotypes range from the most severe, salt-wasting (SW), to the simple virilizing (SV), and nonclassical (NC) CAH forms. We recently determined the crystal structure of human P450 21A2 in complex with progesterone. To gain more insight into the structural and stability changes underlying the phenotypes of individual mutations, we analyzed 24 SW, SV, and NC mutants in the context of the crystal structure of the human enzyme. Our analysis reveals clear differences in the localization of SW, SV, and NC mutations, with many of the first type mapping to the active site and near the heme and/or substrate and mostly resulting in complete loss of enzyme activity. Conversely, NC mutations are often found near the periphery and close to the surface of the protein, and mutant enzymes retain partial activity. The main conclusion from the mutation-structure-activity study is that the severity of the CAH clinical manifestations can be directly correlated with the degree of mutation-induced damage in terms of protein fold stability and active site changes in the structural model. Thus, the NC phenotype is typically associated with mutations that have a compensatory effect, ie, H-bonding replacing hydrophobic interactions and vice versa.
Cytochrome P450 (P450) 21A2 is the major steroid 21-hydroxylase, and deficiency of this enzyme is involved in ∼95% of cases of human congenital adrenal hyperplasia, a disorder of adrenal steroidogenesis. A structure of the bovine enzyme that we published previously (Zhao, B., Lei, L., Kagawa, N., Sundaramoorthy, M., Banerjee, S., Nagy, L. D., Guengerich, F. P., and Waterman, M. R. (2012) Three-dimensional structure of steroid 21-hydroxylase (cytochrome P450 21A2) with two substrates reveals locations of disease-associated variants. J. Biol. Chem. 287, 10613-10622), containing two molecules of the substrate 17α-hydroxyprogesterone, has been used as a template for understanding genetic deficiencies. We have now obtained a crystal structure of human P450 21A2 in complex with progesterone, a substrate in adrenal 21-hydroxylation. Substrate binding and release were fast for human P450 21A2 with both substrates, and pre-steady-state kinetics showed a partial burst but only with progesterone as substrate and not 17α-hydroxyprogesterone. High intermolecular non-competitive kinetic deuterium isotope effects on both kcat and kcat/Km, from 5 to 11, were observed with both substrates, indicative of rate-limiting C-H bond cleavage and suggesting that the juxtaposition of the C21 carbon in the active site is critical for efficient oxidation. The estimated rate of binding of the substrate progesterone (kon 2.4 × 10(7) M(-1) s(-1)) is only ∼2-fold greater than the catalytic efficiency (kcat/Km = 1.3 × 10(7) M(-1) s(-1)) with this substrate, suggesting that the rate of substrate binding may also be partially rate-limiting. The structure of the human P450 21A2-substrate complex provides direct insight into mechanistic effects of genetic variants.
© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.
BACKGROUND - Hormonal contraception use is common among human immunodeficiency virus (HIV)-infected women. Risk of psychiatric and other noninfectious complications of hormonal contraception use has not been described in this population.
METHODS - We performed a retrospective cohort study of HIV-infected women receiving care in Tennessee from 1998 to 2008 to examine the risks of incident psychiatric and other noncommunicable diseases (NCDs), including cardiovascular, hepatic, renal, and malignant diseases, and hormonal contraception use, including depot medroxyprogesterone acetate (DMPA) and combined estrogen- and progestin-containing hormonal contraceptives. We used marginal structural models with inverse probability weights to account for time-varying confounders associated with hormonal contraception use.
RESULTS - Of the 392 women included, 94 (24%) used hormonal contraception during the study period. Baseline psychiatric disease was similar between women who received and did not receive hormonal contraception. There were 69 incident psychiatric diagnoses and 72 NCDs. Only time-varying DMPA use was associated with increased risk of psychiatric disease (adjusted odds ratio [aOR] 3.70; 95% confidence interval [95% CI] 1.32-10.4) and mood disorders, specifically (aOR 4.70 [1.87-11.8]). Time-varying and cumulative combined hormonal contraception use were not statistically associated with other NCDs (aOR 1.64, 95% CI 0.64-4.12 and aOR 1.16, 95% CI 0.86-1.56, respectively). However, risk of incident NCDs was increased with cumulative DMPA exposure (per year exposure aOR 1.45, 95% CI 1.01-2.08).
CONCLUSIONS - Among HIV-infected women, DMPA was associated with risk of incident psychiatric diseases, particularly mood disorders, during periods of use. Cumulative DMPA exposure was also associated with risk of other NCDs. However, combined estrogen and progestin-containing hormonal contraception use was not statistically associated with risk of any NCDs.
Cytochrome P450 (P450) 17A enzymes play a critical role in the oxidation of the steroids progesterone (Prog) and pregnenolone (Preg) to glucocorticoids and androgens. In mammals, a single enzyme, P450 17A1, catalyzes both 17α-hydroxylation and a subsequent 17α,20-lyase reaction with both Prog and Preg. Teleost fish contain two 17A P450s; zebrafish P450 17A1 catalyzes both 17α-hydroxylation and lyase reactions with Prog and Preg, and P450 17A2 is more efficient in pregnenolone 17α-hydroxylation but does not catalyze the lyase reaction, even in the presence of cytochrome b5. P450 17A2 binds all substrates and products, although more loosely than P450 17A1. Pulse-chase and kinetic spectral experiments and modeling established that the two-step P450 17A1 Prog oxidation is more distributive than the Preg reaction, i.e. 17α-OH product dissociates more prior to the lyase step. The drug orteronel selectively blocked the lyase reaction of P450 17A1 but only in the case of Prog. X-ray crystal structures of zebrafish P450 17A1 and 17A2 were obtained with the ligand abiraterone and with Prog for P450 17A2. Comparison of the two fish P450 17A-abiraterone structures with human P450 17A1 (DeVore, N. M., and Scott, E. E. (2013) Nature 482, 116-119) showed only a few differences near the active site, despite only ∼50% identity among the three proteins. The P450 17A2 structure differed in four residues near the heme periphery. These residues may allow the proposed alternative ferric peroxide mechanism for the lyase reaction, or residues removed from the active site may allow conformations that lead to the lyase activity.
© 2015 by The American Society for Biochemistry and Molecular Biology, Inc.
BACKGROUND - Causes of racial disparities in breast cancer incidence and mortality between white and African American women remain unclear. This study evaluated associations of menstrual and reproductive factors with breast cancer risk by race and cancer subtypes.
PATIENTS AND METHODS - Included in the study were 1866 breast cancer cases and 2306 controls recruited in the Nashville Breast Health Study, a population-based case-control study. Multivariable logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs).
RESULTS - African American women were more likely to have estrogen receptor-negative (ER(-)), progesterone receptor-negative (PR(-)), and triple-negative (ER(-)PR(-)HER2(-)) breast cancer than white women. Age at menarche (≥ 14 years) and multiparity (≥ 3 live births) were inversely associated with ER(+) tumors only, whereas late age at first live birth (> 30 years) and nulliparity were associated with elevated risk; such associations were predominantly seen in white women (OR = 0.70, 95% CI = 0.55-0.88; OR = 0.72, 95% CI = 0.56-0.92; OR = 1.42, 95% CI = 1.13-1.79; OR = 1.32, 95% CI = 1.06-1.63, respectively). Age at menopause between 47 and 51 years was associated with elevated risk of ER(-) tumors in both white and African American women. Among women who had natural menopause, positive association between ever-use of hormone replacement therapy and breast cancer risk was seen in white women only (OR = 1.39, 95% CI = 1.03-1.87).
CONCLUSION - This study suggests that certain hormone-related factors are differentially associated with risk of breast cancer subtypes, and these associations also differ by race.
Copyright © 2014 Elsevier Inc. All rights reserved.