A composite menstrual cycle of captive orangutans, with associated hormonal and behavioral variability.

Knowledge of species-typical reproductive endocrinology profiles is crucial for testing hypotheses pertaining to the evolutionary history, reproductive parameters, and life history of a species, and for managing the well-being of individual animals in human care. Large-scale empirical measurements of ovarian hormones, however, are rare for most primate species, including orangutans. In this study, we used enzyme immunoassays (EIA) to quantify estrogen (estrone conjugates; E1 C) and progesterone (pregnanediol-3-glucuronide; PdG) levels for 98 cycles in 7 cycling zoo-housed female orangutans (10-43 years old). We use a subset of these cycles (N = 44) to create the first composite menstrual cycle for orangutans, which serves as a valuable baseline for future comparative analyses and veterinary considerations. Similar to previous studies, we determined the mean ovarian cycle length of orangutans to be 29.7 days (N = 98 cycles), although we illustrate evidence of both intra- and interindividual variation in ovarian steroid production. Given that this study took place in captivity, we consider how energetic and psychosocial aspects of the zoo environment, such as greater food availability and potential stress, may affect the reproductive physiology and sexual behavior of these females. Furthermore, we discuss the role that age and genetic background may play in producing variability. Finally, we test whether ovarian hormone levels correlate with the reproductive behaviors of these female orangutans using associated behavioral data. Our results suggest that matings are more common during the periovulatory period than outside of it, but do not support a consistent link between hormonal indices of fecundability and mating behaviors in these individuals.

Distinct urinary progesterone metabolite profiles during the luteal phase.

OBJECTIVES: During normal menstrual cycles, serum levels of progesterone vary widely between cycles of same woman and between women. This study investigated the profiles of pregnanediol during the luteal phase. METHODS: Data stemmed from a previous multicenter prospective observational study and concerned 107 women (who contributed 326 menstrual cycles). The study analyzed changes in observed cervical mucus discharge, various hormones in first morning urine, and serum progesterone. Transvaginal ultrasonography and cervical mucus helped identifying the day of ovulation. Changes in pregnanediol glucuronide levels during the luteal phase were examined and classified according to the length of that phase, a location parameter, and a scale parameter. Associations between nine pregnanediol glucuronide profiles and other hormone profiles were examined. RESULTS: Low periovulatory pregnanediol glucuronide levels and low periovulatory luteinizing hormone levels were associated with delayed increases in pregnanediol glucuronide after ovulation. That 'delayed increase profile' was more frequently associated with cycles with prolonged high LH levels than in cycles with rapid pregnanediol glucuronide increases. A 'plateau-like profile' during the luteal phase was associated with longer cycles, cycles with higher estrone-3-glucuronide and pregnanediol glucuronide during the preovulatory phase, and cycles with higher periovulatory pregnanediol glucuronide levels. CONCLUSIONS: Distinct profiles of urinary progesterone levels are displayed during the luteal phase. These profiles relate to early hormone changes during the menstrual cycle. In everyday clinical practice, these findings provide further evidence for recommending progesterone test seven days after the mucus peak day. The search for other correlations and associations is underway.

Short-term serum and urinary changes in sex hormones of healthy pre-pubertal children after the consumption of commercially available whole milk powder: a randomized, two-level, controlled-intervention trial in China.

Currently, commercial milk may contain abundant pregnancy-related hormones, the regular consumption of which puts children at a risk of precocious puberty and sex-hormone-associated tumors in adulthood. In this intervention trial, 51 healthy prepubescent children were randomly assigned to the intervention or control arms at a ratio of 3 : 1 to receive 250 or 600 mL m-2 (body surface area) of milk intervention or matching equienergetic sugar water as the control. On testing cow's milk, progesterone was detected, while estrone, estradiol (E2), and testosterone (T2) were not. Cow's milk ingestion did not significantly influence the serum FSH, E2, PRL, LH, and T2 levels (P > 0.05) of pre-pubertal children 3 h after the intervention, while it increased their serum progesterone levels (P < 0.05) when compared with that in the control arm. Regarding the urinary hormone levels, cow's milk ingestion increased the urinary pregnanediol level within 4 h (P < 0.05), but not significantly when compared with that of the control (P > 0.05). The level of pregnanediol and E2 in the morning urine for three consecutive days showed no significant difference between the two arms (P > 0.05). Drinking commercial milk with progesterone influenced the progesterone levels of pre-pubertal children in hours but not days and did not affect other sex hormone levels of pre-pubertal children.

Descriptive analysis of the relationship between progesterone and basal body temperature across the menstrual cycle.

OBJECTIVE: Describe the relationship between basal body temperature (BBT) and pregnanediol-3 alpha-glucuronide (PDG, the urine metabolite of progesterone) across the menstrual cycle. DESIGN: Observational study. SETTING: Study carried out from 1996 to 1997 in eight European family planning clinics. PARTICIPANT(S): One hundred and seven normally fertile and cycling women. MAIN OUTCOME MEASURE(S): BBT and PDG level on each day of 283 cycles and ultrasound determination of the day of ovulation. RESULT: (s): In comparison with previous end-of-cycle levels, decreases in PDG and BBT on the first day of menses were seen in nearly 90% and 80% of cycles, respectively. In a non-negligible percentage of cycles, luteolysis would continue during menses: between the second and the third day after menses, small but significant decreases in PDG and BBT were seen in 76% and 48% of cycles, respectively. During the peri-ovulatory phase, between the third and the second day before ovulation, PDG and BBT began to rise in 56% and 41% of cycles, respectively. There was a medium degree of correlation between PDG levels and BBT (r = 0.53; 7,279 days with available measurements). The relationship between PDG levels and BBT was linear at low PDG levels but BBT increased no longer when PDG levels continued to rise above a threshold of nearly 10 mcg/mg Cr. CONCLUSION: (s): PDG and BBT had parallel increases at low PDG rates but diverged at higher rates.

Algorithms with Area under the Curve for Daily Urinary Estrone-3-Glucuronide and Pregnanediol-3-Glucuronide to Signal the Transition to the Luteal Phase.

Background and Objectives: Home fertility assessment methods (FAMs) for natural family planning (NFP) have technically evolved with the objective metrics of urinary luteinizing hormone (LH), estrone-3-glucuronide (E3G) and pregnanediol-3-glucuronide (PDG). Practical and reliable algorithms for timing the phase of cycle based upon E3G and PDG levels are mostly unpublished and still lacking. Materials and Methods: A novel formulation to signal the transition to the luteal phase was discovered, tested, and developed with a data set of daily E3G and PDG levels from 25 women, 78 cycles, indexed to putative ovulation (day after the urinary LH surge), Day 0. The algorithm is based upon a daily relative progressive change in the ratio, E3G-AUC/PDG-AUC, where E3G-AUC and PDG-AUC are the area under the curve for E3G and PDG, respectively. To improve accuracy the algorithm incorporated a three-fold cycle-specific increase of PDG. Results: An extended negative change in E3G-AUC/PDG-AUC of at least nine consecutive days provided a strong signal for timing the luteal phase. The algorithm correctly identified the luteal transition interval in 78/78 cycles and predicted the start day of the safe period as: Day + 2 in 10/78 cycles, Day + 3 in 21/78 cycles, Day + 4 in 28/78 cycles, Day + 5 in 15/78 cycles, and Day + 6 in 4/78 cycles. The mean number of safe luteal days with this algorithm was 10.3 ± 1.3 (SD). Conclusions: An algorithm based upon the ratio of the area under the curve for daily E3G and PDG levels along with a relative PDG increase offers another approach to time the phase of cycle. This may have applications for NFP/FAMs and clinical evaluation of ovarian function.

Direct measurement of pregnanediol 3-glucuronide (PDG) in dried urine spots by liquid chromatography-mass spectrometry to detect ovulation.

BACKGROUND: Non-invasive self-testing using an objective chemical method to detect ovulation is valuable for women planning conception, practising contraception or undergoing infertility investigations or treatment. METHODS: Based on luteal phase secretion of progesterone (P4) and excretion of its major metabolite, pregnanediol glucuronide (PDG), we developed a novel direct liquid chromatography-mass spectrometry (LCMS) method to measure PDG and other steroid glucuronides in urine and in dried urine spots (DUS) without deconjugation or derivatization. Urine PDG by LCMS and immunoassay (P3G) and P4 by immunoassay with and without adjustment for creatinine were evaluated in daily first void urine samples from 10 women through a single menstrual cycle in which ovulation was confirmed by serial transvaginal ultrasound. RESULTS: Urine PDG with and without creatinine adjustment was stable during the follicular phase with the expected striking rise in the luteal phase peaking at 5 days after ovulation. Using a single spot urine sample (100 µL) or a DUS (<20 µL urine) and an optimal threshold to distinguish pre- from post-ovulatory samples, in ROC analysis urine PDG adjusted for creatinine accurately identified ovulation in 92 % of samples was comparable with P3G immunoassay and superior to urine P4 with or without adjustment for creatinine. Extending the analysis to two or three consecutive daily samples reduced the false negative rate from 8% to 2.6 % for two and 1.9 % for three urine samples. CONCLUSIONS: This method holds promise as a non-invasive self-test method for women to determine by an objective chemical method their ovulatory status.

Lowered progesterone metabolite excretion and a variable LH excretion pattern are associated with vasomotor symptoms but not negative mood in the early perimenopausal transition: Study of Women's Health Across the Nation.

OBJECTIVE: The menopausal transition is characterized by progressive changes in ovarian function and increasing circulating levels of gonadotropins, with some women having irregular menstrual cycles well before their final menstrual period. These observations indicate a progressive breakdown of the hypothalamic-pituitary-ovarian axis often associated with an increase in menopausal symptoms. Relationships between vasomotor symptoms (VMS) and depressed mood and sleep as well as a bidirectional association between VMS and depressed mood in mid-life women have been reported, but the endocrine foundations and hormone profiles associated with these symptoms have not been well described. Our objective was to determine the relationship between daily urinary hormone profiles and daily logs of affect and VMS during the early perimenopausal transition. STUDY DESIGN: SWAN, the Study of Women's Health Across the Nation, is a large, mutli-ethnic, multisite cohort study of 3302 women aged 42-52 at baseline, designed to examine predictors of health and disease in women as they traversed the menopause. Inclusion criteria were: an intact uterus and at least one ovary present, at least one menstrual period in the previous three months, no use of sex steroid hormones in the previous three months, and not pregnant or lactating. A subset (n = 849) of women aged 43-53 years from all study sites in the first Daily Hormone Study collection were evaluated for this substudy. OUTCOME MEASURES: We measured daily VMS, and urinary hormones: follicle stimulating hormone (FSH), luteinizing hormone (LH), pregnanediol glucuronide (PdG) and estradiol (estrone conjugate, E1C). RESULTS: A variable pattern of LH and negative LH feedback were the hormone patterns most strongly associated with increased VMS. In contrast, no hormone pattern was significantly related to negative mood. CONCLUSION: Fluctuations of LH associated with low progesterone production were associated with VMS but not negative mood, suggesting different endocrine patterns may be related to increased negative mood than to the occurrence of VMS.

The Fertility Indicator Equation Using Serum Progesterone and Urinary Pregnanediol-3-Glucuronide for Assessment of Ovulatory to Luteal Phase Transition.

Background and Objectives: The Fertility Indicator Equation (FIE) has been shown to signal the fertile phase during the ovulatory menstrual cycle. It was hypothesized that this formulation, a product of two sequential normalized changes with a sign indicating direction of change, could be used to identify the transition from ovulatory to luteal phase with daily serum progesterone (P) and urinary pregnanediol-3-glucuronide (PDG) levels. Materials and Methods: Day-specific serum P levels from two different laboratories and day-specific urinary PDG levels from an additional two different laboratories were submitted for FIE analysis. These day-specific levels included mean or median, 5th, 10th, 90th and 95th percentile data. They were indexed to the day of ovulation, day 0, by ultrasonography, serum or urinary luteinizing hormone (LH). Results: All data sets showed a clear "cluster"-a periovulatory sequence of positive FIE values with a maximum. All clusters of +FIE signaled the transition from the ovulatory to luteal phase and were at least four days in length. The start day for the serum P and urinary PDG FIE clusters ranged from -3 to -1 and -3 to +2, respectively. The end day for serum P and PDG clusters went from +2 to +7 and +4 to +8, respectively. Outside these periovulatory FIE-P and FIE-PDG clusters, there were no consecutive positive FIE values. In addition, the maximum FIE-P and FIE-PDG values throughout the entire cycles were found in the clusters. Conclusions: FIE analysis with either daily serum P or urinary PDG levels provided a distinctive signature to recognize the periovulatory interval. The Fertility Indicator Equation served to robustly signal the transition from the ovulatory phase to the luteal phase. This may have applications in natural family planning especially with the recent emergence of home PDG tests.

CYP3A7*1C allele: linking premenopausal oestrone and progesterone levels with risk of hormone receptor-positive breast cancers.

BACKGROUND: Epidemiological studies provide strong evidence for a role of endogenous sex hormones in the aetiology of breast cancer. The aim of this analysis was to identify genetic variants that are associated with urinary sex-hormone levels and breast cancer risk. METHODS: We carried out a genome-wide association study of urinary oestrone-3-glucuronide and pregnanediol-3-glucuronide levels in 560 premenopausal women, with additional analysis of progesterone levels in 298 premenopausal women. To test for the association with breast cancer risk, we carried out follow-up genotyping in 90,916 cases and 89,893 controls from the Breast Cancer Association Consortium. All women were of European ancestry. RESULTS: For pregnanediol-3-glucuronide, there were no genome-wide significant associations; for oestrone-3-glucuronide, we identified a single peak mapping to the CYP3A locus, annotated by rs45446698. The minor rs45446698-C allele was associated with lower oestrone-3-glucuronide (-49.2%, 95% CI -56.1% to -41.1%, P = 3.1 × 10-18); in follow-up analyses, rs45446698-C was also associated with lower progesterone (-26.7%, 95% CI -39.4% to -11.6%, P = 0.001) and reduced risk of oestrogen and progesterone receptor-positive breast cancer (OR = 0.86, 95% CI 0.82-0.91, P = 6.9 × 10-8). CONCLUSIONS: The CYP3A7*1C allele is associated with reduced risk of hormone receptor-positive breast cancer possibly mediated via an effect on the metabolism of endogenous sex hormones in premenopausal women.

Detecting the abuse of 19-norsteroids in doping controls: A new gas chromatography coupled to isotope ratio mass spectrometry method for the analysis of 19-norandrosterone and 19-noretiocholanolone.

The detection of 19-norsteroids abuse in doping controls currently relies on the determination of 19-norandrosterone (19-NA) by gas chromatography-tandem mass spectrometry (GC-MS/MS). An additional confirmatory analysis by gas chromatography coupled to isotope ratio mass spectrometry (GC-C-IRMS) is performed on samples showing 19-NA concentrations between 2.5 and 15 ng/ml and not originated from pregnant female athletes or female treated with 19-norethisterone. 19-Noretiocholanolone (19-NE) is typically produced to a lesser extent as a secondary metabolite. The aim of this work was to improve the GC-C-IRMS confirmation procedure for the detection of 19-norsteroids misuse. Both 19-NA and 19-NE were analyzed as target compounds (TCs), whereas androsterone (A), pregnanediol (PD), and pregnanetriol (PT) were selected as endogenous reference compounds (ERCs). The method was validated and applied to urine samples collected by three male volunteers after the administration of nandrolone-based formulations. Before the instrumental analysis, urine samples (<25 ml) were hydrolyzed with ß-glucuronidase from Escherichia coli and extracted with n-pentane. Compounds of interest were purified through a single (for PT) or double (for 19-NE, 19-NA, A, and PD) liquid chromatographic step, to reduce the background noise and eliminate interferences that could have affect the accuracy of δ13 C values. The limit of quantification (LOQ) of 2 ng/ml was ensured for both 19-NA and 19-NE. The 19-NE determination could be helpful in case of "unstable" urine samples, in late excretion phases or when coadministration with 5α-reductase inhibitors occur.

Stability of chimpanzee (Pan troglodytes) urinary reproductive hormones during long-term preservation on filter paper.

Urine contains multiple water-soluble hormones, which are valuable non-invasive biomarkers for the monitoring of reproductive status and health. An effective method for drying urine on filter paper was previously developed to preserve wildlife urine samples where electrical equipment was not available for this; however, the stability of samples preserved in this way remains to be verified. Here, we developed and validated a method to elute multiple water-soluble reproductive hormones from filter paper that had been stored for an extended period of time. Aliquots of urine from chimpanzees were adsorbed on filter papers, air dried and stored for 1 year at room temperature. Estrone-3-conjugate (E1C), pregnanediol-3-glucuronide (PdG), estriol-3-glucuronide (E3G), and chorionic gonadotropin (CG) were eluted into deionized water from the filter papers and measured using enzyme immunoassays (EIAs). The mean recoveries of E1C, PdG, and creatinine from filter papers stored for 1 year were 69.5%, 128.7%, and 83.8%, respectively. The profiles of E1C and PdG from preserved filter papers significantly correlated with those derived from a direct analysis of the frozen urine of menstruating chimpanzees. We detected E3G and CG from 1-year-old filter papers for urine collected during early pregnancy, but the recovery of E3G was low and CG profiles did not correlate with those of the original frozen urine samples. The method proposed here for the elution and measurement of reproductive hormones in urine preserved for a long period of time on filter paper provides a practical and simple way to monitor the reproductive status of chimpanzees. We propose that this method can also be utilized in field studies of other wild nonhuman primates.

Non-targeted metabolomics analyses by mass spectrometry to explore metabolic stress after six training weeks in high level swimmers.

The objective was to compare the metabolic responses of high-level national swimmers to threshold or polarised training. 22 swimmers (n = 12 males and 10 females) participated in a 28-week cross-over intervention study consisting of 2 × 6 period weeks of training. Swimmers were assigned randomly to either training group for the first period: polarised (POL) (81% in energetic zone 1: blood lactate [La]b ≤ 2 mmol.L-1; 4% in zone 2: 2 mmol.L-1 <[La]b ≤ 4 mmol.L-1; 15% in zone 3: [La]b > 4 mmol.L-1) or threshold (THR) (65%/25%/10%). Before and after each training period, urine samples were collected for non-targeted metabolomics analysis. Mixed model analysis was performed on metabolomics data including fatigue class factors and/or training and/or interaction. Ion intensities of 6-keto-decanoylcarnitine (+31%), pregnanediol-3-glucuronide (+81%), P-cresol sulphate (+18%) were higher in the threshold group (P < 0.05) indicating higher glycogenic depletion and inflammation without alteration of the neuroendocrine stress axis. 4-phenylbutanic acid sulphate was 200% higher in less fatigued swimmers (P < 0.01) linking the anti-inflammatory activity at the cell membrane level to the subjective perception of fatigue. This research suggests the importance of replenishing glycogen stores and reducing inflammation during high thresholds training loads.

Hormonal Profiles of Menstrual Bleeding Patterns During the Luteal-Follicular Transition.

CONTEXT: Menstrual cycle function is determined by a complex endocrine axis that controls the ovaries and endometrium. While the late luteal phase is characterized by declining progesterone and estrogen, how these hormonal profiles relate to menstrual bleeding patterns is not well understood. OBJECTIVE: Characterize associations between luteal phase hormonal profiles and subsequent menstrual bleeding patterns, specifically spotting before bleeding. DESIGN, SETTING, AND PARTICIPANTS: We examined creatinine-adjusted urinary estrone 3-glucuronide (E13G) and pregnanediol 3-glucuronide (Pd3G) levels in relation to spotting in 116 premenopausal women (ages 20-47) who kept daily menstrual diaries and collected first morning urine samples for ≥ 2 consecutive cycles or 1 luteal-follicular transition (n = 283 transitions). We used linear mixed models to estimate associations between luteal phase hormone levels and spotting before bleeding. MAIN OUTCOME MEASURE(S) AND RESULTS: Transitions with ≥ 1 days of spotting before menstrual bleeding (n = 118) had greater luteal phase Pd3G levels vs nonspotting transitions (n = 165). Differences in Pd3G between spotting and nonspotting transitions were largest at menses onset (34.8%, 95% confidence interval, 18.9%, 52.7%). Pd3G levels for spotting transitions dropped to similar levels as nonspotting transitions an average of 1 day later, which aligned with the first day of bleeding for transitions with contiguous spotting. Spotting transitions were preceded by slower rates of Pd3G decline than nonspotting transitions, whereas E13G declines were similar. CONCLUSIONS: Self-reported bleeding patterns may provide insight into luteal phase Pd3G levels. First bleed appears to be the best choice for defining the end of the luteal phase and achieving hormonal consistency across transitions.

Quantitation of serum pregnanediol-3-glucuronide level in different endocrine statuses by LC-MS/MS.

Pregnanediol-3-glucuronide (PdG) is the major terminal metabolite of progesterone, playing an important role in physiological processes, such as the female menstrual cycle, pregnancy (supports gestation), embryogenesis and maternal immune response of humans and other species. Hence, accurate measurement of PdG in serum/plasma is needed for the evaluation of progesterone production. However, such high-specificity determination of PdG is lacking in clinical sample detection. In this study, a highly sensitive and accurate LC-MS/MS method was firstly established for subsequent measurement of PdG in serum of three different female groups: thyroid cancer patients (TCs), healthy controls (HCs) and pregnant women. The factors affecting the sample preparation, MS/MS method, gradient elution program, selection of chromatographic column and internal standard (IS) have been optimized in this study. Compared with enzyme immunoassay (EIA) method, we used LC-MS/MS to shorten analysis time, increase sensitivity, raise specificity, simplify sample preparation, and reduce costs. As a result, the linear range of the method was from 0.38 to 100 ng/mL with a limit of quantification (LOD) of 0.01 ng/mL. Precision assays showed that relative standard deviation (RSD) was less than 10.6, accuracy was between 90.6 % and 110.4 %, and mean recovery was 103.4 %. In addition, the serum PdG/creatinine levels were significantly down-regulated in TCs and up-regulated in pregnant women versus HCs. Receive operating characteristic curve (ROC) analysis enabled to identify TC with a sensitivity of 83.3 %, specificity of 68.0 % and area under curve (AUC) of 0.781 (95 % CI: 0.684 to 0.879), and it enabled to identify pregnant women with a sensitivity of 94.7 %, specificity of 68.5 % and AUC of 0.811 (95 % CI: 0.732 to 0.890). Our results implied that an increase in female serum PdG/creatinine level might be associated with a risk of pregnancy, but serum PdG/creatinine decreasing might be related to a risk of TC.

The effect of testosterone on ovulatory function in transmasculine individuals.

BACKGROUND: An estimated 1.4 million persons in the United States identify as transgender or nonbinary, signifying that their gender identity does not correspond with their assigned sex at birth. Individuals assigned female at birth may seek gender-affirming hormone therapy with testosterone. No studies have directly examined ovulatory function in transmasculine individuals using injectable testosterone. OBJECTIVES: Our primary objective was to determine the effect of testosterone on ovulatory suppression in transmasculine individuals. Secondary objectives were to determine predictors of ovulation in transmasculine individuals on testosterone, and to assess the effect of testosterone on antimüllerian hormone. MATERIALS AND METHODS: This prospective observational study recruited participants from a community clinic that provides gender-affirming hormone therapy. Enrolled individuals were assigned female at birth and were currently using or seeking to initiate masculinizing therapy with injectable testosterone esters (transmasculine individuals). Over a 12-week study period, participants collected daily urine samples for pregnanediol-3-glucoronide testing and completed daily electronic bleeding diaries. We assessed monthly serum mid-dosing interval testosterone, estradiol and sex hormone binding globulin, and antimüllerian hormone values at baseline and study end. Ovulation was defined as pregnanediol-3-glucoronide greater than 5 µg/mL for 3 consecutive days. The primary outcome was the proportion of participants who ovulated during the study period. We examined predictors of ovulation such as age, length of time on testosterone, serum testosterone levels, body mass index, and bleeding pattern. RESULTS: From July to November 2018, we enrolled 32 individuals; 20 completed the study (14 continuing testosterone users, 6 new users). Median age was 23 years (range 18-37 years). Bleeding or spotting during the study period was noted by 41% of participants (13/32). Among continuing users, median testosterone therapy duration was 11 months (range 1-60 months). A single ovulation was observed out of a total of 61 combined months of testosterone use; however, several transient rises in pregnanediol-3-glucoronide followed by bleeding episodes were suggestive of 7 dysfunctional ovulatory cycles among 7 individuals. There was no difference in antimüllerian hormone from baseline to 12 weeks between participants initiating testosterone and continuing users of testosterone. We did not have the power to examine our intended predictors given the low numbers of ovulatory events, but found that longer time on testosterone and presence of vaginal bleeding over 12 weeks were associated with transient rises in pregnanediol-3-glucoronide. CONCLUSION: This study suggests that testosterone rapidly induces hypothalamic-pituitary-gonadal suppression, resulting in anovulation in a proportion of new users. Importantly, these data also suggest that some long-term testosterone users break through the hormonal suppression and experience an ovulatory event, thereby raising concerns pertaining to the need for contraception in transmasculine individuals engaged in sexual intercourse with sperm-producing partners. Given the small number of overall participants, this work is hypothesis generating. Larger studies are needed to confirm and to clarify these findings.

Multiplex Surface-Enhanced Raman Scattering Identification and Quantification of Urine Metabolites in Patient Samples within 30 min.

Successful translation of laboratory-based surface-enhanced Raman scattering (SERS) platforms to clinical applications requires multiplex and ultratrace detection of small biomarker molecules from a complex biofluid. However, these biomarker molecules generally exhibit low Raman scattering cross sections and do not possess specific affinity to plasmonic nanoparticle surfaces, significantly increasing the challenge of detecting them at low concentrations. Herein, we demonstrate a "confine-and-capture" approach for multiplex detection of two families of urine metabolites correlated with miscarriage risks, 5ß-pregnane-3α,20α-diol-3α-glucuronide and tetrahydrocortisone. To enhance SERS signals by 1012-fold, we use specific nanoscale surface chemistry for targeted metabolite capture from a complex urine matrix prior to confining them on a superhydrophobic SERS platform. We then apply chemometrics, including principal component analysis and partial least-squares regression, to convert molecular fingerprint information into quantifiable readouts. The whole screening procedure requires only 30 min, including urine pretreatment, sample drying on the SERS platform, SERS measurements, and chemometric analyses. These readouts correlate well with the pregnancy outcomes in a case-control study of 40 patients presenting threatened miscarriage symptoms.

Daily luteal serum and urinary hormone profiles in the menopause transition: Study of Women's Health Across the Nation.

OBJECTIVE: To further characterize the endocrinology of the menopause transition, we sought to determine: whether relationships between urine and serum hormones are maintained as women enter their sixth decade; whether a single luteal phase serum progesterone (P) is reflective of integrated-luteal urinary pregnanediol glucuronide (uPdg); and whether serum P, like luteal uPdg, declines as women approach their final menses (FMP). METHODS: The Study of Women's Health Across the Nation (SWAN) Daily Hormone Study's (DHS) is a community-based observational study. A subset of participants underwent a timed, luteal blood draw planned for cycle days 16 to 24 during the same month of DHS collection. Serum-luteinizing hormone (LH), follicle-stimulating hormone (FSH), estradiol and P, and urine LH, FSH, estrone conjugates (E1c), and daily and integrated luteal uPdg were measured in 268 samples from 170 women. Serum/urine hormone associations were determined using Pearson's correlation and linear regression, adjusted for concurrent age, body mass index, smoking status, and race/ethnicity. RESULTS: Pearson's r ranged from 0.573 (for LH) to 0.843 (for FSH) for serum/urine correlations. Integrated luteal uPdg weakly correlated with serum P (Pearson's r = 0.26, P = 0.004) and explained 7% of the variability in serum P in adjusted linear regression (total R 0.09, P = 0.002). Serum P demonstrated a marginally significant decline with approaching FMP in adjusted analysis (P = 0.04). CONCLUSIONS: Urine and serum hormones maintain a close relationship in women into their sixth decade of life. Serum luteal P was weakly reflective of luteal Pdg excretion.

Preconception Perceived Stress Is Associated with Reproductive Hormone Levels and Longer Time to Pregnancy.

BACKGROUND: Women who experience pregnancy loss are especially prone to high stress, though the effects of stress on reproductive outcomes in this vulnerable population are unknown. We assessed relationships between perceived stress and hormones, anovulation, and fecundability among women with prior loss. METHODS: One thousand two hundred fourteen women with 1-2 prior losses were followed for ≤6 cycles while attempting pregnancy and completed end-of-cycle stress assessments. For cycles 1 and 2, women also collected daily urine and completed daily perceived stress assessments. We assessed anovulation via. an algorithm based on human chorionic gonadotropin (hCG), pregnanediol-3-glucuronide (PdG), luteinizing hormone (LH), and fertility monitor readings. Pregnancy was determined via. hCG. Adjusted weighted linear mixed models estimated the effect of prospective phase-varying (menses, follicular, periovulatory, and luteal) perceived stress quartiles on estrone-1-glucuronide (E1G), PdG, and LH concentrations. Marginal structural models accounted for time-varying confounding by hormones and lifestyle factors affected by prior stress. Poisson and Cox regression estimated risk ratios and fecundability odds ratios of cycle-varying stress quartiles on anovulation and fecundability. Models were adjusted for age, race, body mass index (BMI), parity, and time-varying caffeine, alcohol, smoking, intercourse, and pelvic pain. RESULTS: Women in the highest versus lowest stress quartile had lower E1G and PdG concentrations, a marginally higher risk of anovulation [1.28; 95% confidence interval (CI) = 1.00, 1.63], and lower fecundability (0.71; 95% CI = 0.55, 0.90). CONCLUSION: Preconception perceived stress appears to adversely affect sex steroid synthesis and time to pregnancy. Mechanisms likely include the effects of stress on ovulatory function, but additional mechanisms, potentially during implantation, may also exist.

Pilot observational prospective cohort study on the use of a novel home-based urinary pregnanediol 3-glucuronide (PDG) test to confirm ovulation when used as adjunct to fertility awareness methods (FAMs) stage 1.

RATIONALE: Ovulation confirmation is a fundamental component of the evaluation of infertility. PURPOSE: To inform the design of a larger clinical trial to determine the effectiveness of a new home-based pregnanediol glucuronide (PDG) urine test to confirm ovulation when compared with the standard of serum progesterone. METHODS: In this observational prospective cohort study (single group assignment) in an urban setting (stage 1), a convenience sample of 25 women (aged 18-42 years) collected daily first morning urine for luteinisinghormone (LH), PDG and kept a daily record of their cervical mucus for one menstrual cycle. Serum progesterone levels were measured to confirm ovulation. Sensitivity and specificity were used as the main outcome measures. Estimation of number of ultrasound (US)-monitored cycles needed for a future study was done using an exact binomial CI approach. RESULTS: Recruitment over 3 months was achieved (n=28) primarily via natural fertility regulation social groups. With an attrition rate of 22%, specificity of the test was 100% for confirming ovulation. Sensitivity varied depending on whether a peak-fertility mucus day or a positive LH test was observed during the cycle (85%-88%). Fifty per cent of participants found the test results easy to determine. A total of 73 US-monitored cycles would be needed to offer a narrow CI between 95% and 100%. CONCLUSION: This is first study to clinically evaluate this test when used as adjunct to the fertility awareness methods. While this pilot study was not powered to validate or test efficacy, it helped to provide information on power, recruitment and retention, acceptability of the procedures and ease of its use by the participants. Given this test had a preliminary result of 100% specificity, further research with a larger clinical trial (stage 2) is recommended to both improve this technology and incorporate additional approaches to confirm ovulation. TRIAL REGISTRATION NUMBER: NCT03230084.

Urinary excretion of sex steroid hormone metabolites after consumption of cow milk: a randomized crossover intervention trial.

Background: Current cow milk production practices introduce considerable levels of pregnancy hormones into the milk. Humans are exposed to these hormones when cow milk is consumed, and this may explain the observed association between cow milk consumption and several hormone-sensitive cancers. Objectives: The aim of the study was to evaluate whether cow milk consumption is associated with an increase in urinary excretion of sex steroid hormones and their metabolites in humans. Methods: We conducted a randomized crossover intervention feeding experiment. A total of 109 postmenopausal women consumed 1 L of semiskimmed milk (1.5% fat) per day for 4 d and 1 L of whole milk (3.5% fat) per day for 4 d, intersected by 4-d wash-out periods. Sex steroid hormone levels were measured in 24-h urine samples collected at the end of each intervention and wash-out period. Results: Estrogens, androgens, and progesterone were detected in the examined milk samples used for our intervention. Although a very high proportion of the estrogens were conjugated, only small proportions of the androgens and progesterone were conjugated. Milk consumption resulted in a significant increase in urinary estrone (E1) excretion, whereas estradiol (E2), estriol (E3), and 16ketoE2 excretion only increased after semiskimmed milk consumption. Urinary pregnanediol glucuronide excretion was not significantly affected. Conclusion: Cow milk consumption increases urinary excretion of E1 in humans. Ingestion of semiskimmed milk appears also to raise E2, E3, and 16ketoE2 excretion, but future studies need to confirm these associations. This trial was registered at https://www.drks.de as DRKS00003377.