No open license is recorded for this paper. Reuse terms are set by the publisher.
Open Access
Free full text on PubMed Central (PMC9698449)
Author affiliations
1 institution
Unit of Andrology and Reproductive Medicine, Department of Medicine, University of Padova, Padova, Italy.
RRM Academy Synopsis
Cervical mucus protein levels differed by cycle phase in six women
Cervical mucus protein levels differed by cycle phase in six healthy women. In this small lab study, 17 mucus samples were checked under a microscope and by mass spectrometry. Of 48 proteins found, 25 were proposed as possible ovulation markers.
Key Findings
Six fertile women aged 25 to 30 with regular cycles gave cervical mucus samples on days 7, 12 and 18 of one cycle. Seventeen samples were analyzed.
Light microscopy identified four mucus types, L, S, P and G, by their crystal patterns. The patterns matched earlier descriptions in the literature.
Mass spectrometry identified 48 proteins in total across the three cycle-day groups.
Seventeen proteins significantly decreased and eight significantly increased on day 18 compared with days 7 and 12. The authors propose these 25 as potential ovulation biomarkers.
Seven proteins were absent on day 18, when most samples were in the ovulatory phase. All seven take part in the immune response.
Interpretation
The study is a small exploratory laboratory analysis. Six women each gave samples once on three days of a single cycle, and the proteins were measured in pooled samples grouped by cycle day. The paper lists which proteins change and proposes candidates. The study did not test whether any protein predicts ovulation in advance. The authors acknowledge the relatively small sample. Samples were assigned to a cycle phase by their microscope pattern and the date of the last period. The authors suggest ovulation came later in the cycle than clinical guidelines assume, on the strength of these six women.
RRM Context
Cervical mucus is a working sign in fertility awareness-based methods. The authors say physicians and women using them treat mucus as an indirect guide to when ovulation happens. This paper adds a laboratory view of what changes inside the mucus. The authors cite a study of 612,613 cycles in which only 13% of cycles were 28 days long. They call the fertile window highly unpredictable even when cycles are regular.
Our editorial summary of this paper, not the article's abstract.
Abstract
There is an increasing number of couples interested in identifying the fertile window for the purpose of conceiving. From what has been published so far, it can be concluded that there are no reliable methods to predict ovulation, and, therefore, to predict the fertile window. Proteins of the cervical mucus (CM) could behave as biomarkers to allow the early and precise identification of ovulation. CM samples were collected from the lumen of the cervical canal from women of reproductive age, on three different days of the same menstrual cycle. Samples were first analyzed and classified by light microscopy. High-resolution mass spectrometry and bioinformatic analysis were performed afterwards to determine the in vivo changes of CM protein composition. CM underwent cyclical changes in its biophysical composition, which were evidenced by changes in the crystallographic patterns observed under the light microscope. The proteomic analysis revealed changes in the protein composition of CM along the cycle. Twenty-five out of the forty-eight total proteins identified could become potential biomarkers of ovulation. The coordinated changes in the composition of the CM around the time of ovulation could be happening to specifically grant access to a foreign body, such as the sperm might be.
Genetic variability within the follicle-stimulating hormone (FSH)-related genes might contribute to phenotypic heterogeneity in patients with Klinefelter syndrome (KS), yet its clinical impact on sperm retrieval remains unclear.
To investigate the association between FSHB c.211 G > T and FSHR polymorphisms (c.2039 A > G and c.29 G > A) and hormonal parameters, as well as their potential role in predicting sperm retrieval rate (SRR) in patients with KS undergoing testicular sperm extraction (TESE).
A retrospective cohort of patients with KS was analyzed. Only subjects not receiving testosterone replacement therapy were included (n = 417). Hormonal, anthropometric, and clinical variables were compared across genotypes using nonparametric tests. Additive genetic models were applied to evaluate allele-dose effects. Multivariable logistic regression was performed to identify independent predictors of SRR. Predictive performance of clinical and combined clinical-genetic models was assessed using ROC curve analysis.
FSHB and FSHR genes polymorphisms were associated with variations in serum FSH concentrations, confirming a modulatory role of the FSH-related genes. In additive modeling, the FSHR c.29 G > A polymorphism emerged as an independent predictor of SRR (adjusted OR: 0.29; 95% CI, 0.09-0.97), whereas FSHB c.211 G > T showed a nonsignificant trend. The inclusion of genetic variants modestly improved predictive accuracy compared with clinical variables alone, as demonstrated by ROC analysis.
Genetic variants within the genes involved in FSH action might contribute incremental information for the prediction of sperm retrieval in patients with KS. While the overall predictive gain remains moderate, additive genetic modeling highlights a potential allele-dose effect of FSHR c.29 G > A on reproductive outcome, supporting a role for integrated clinical-genetic assessment in this population, although external validation remains required.
Klinefelter syndrome (KS) is characterized by marked phenotypic heterogeneity that might be influenced by genetic modifiers, including androgen receptor (AR) repeat length (CAGn and GGCn). The clinical relevance of these repeat lengths in patients with KS before testosterone replacement therapy (TRT) remains unclear.
To investigate the association between AR repeat length and anthropometric, hormonal, metabolic, and reproductive parameters in a well-characterized cohort of untreated adult patients with KS.
In this cross-sectional single-center study, 214 men with classical 47, XXY karyotype were evaluated prior to TRT. Clinical, biochemical, and reproductive parameters were analyzed according to AR CAGn and GGCn repeat length. Nonparametric tests, multivariable linear and logistic regression models, and interaction terms between CAGn and GGCn were tested. Standardized beta coefficients were used to compare the relative contribution of AR repeat length with major clinical determinants.
In unadjusted analyses, CAG repeat length was associated with estradiol concentrations, whereas GGC repeat length showed associations with hematocrit, platelet count, and total cholesterol. However, most associations were characterized by small effect sizes and did not persist after multivariable adjustment for possible confounders (age, BMI, and total testosterone levels). Moreover, AR repeat length was not associated with sperm retrieval rate. Standardized beta analyses demonstrated that testosterone levels, BMI, and age accounted for the largest proportion of phenotypic variability, whereas CAGn and GGCn repeat length had minimal roles.
In untreated patients with KS, AR repeat length (CAGn and GGCn) appears to have a limited clinical impact compared with classical endocrine and metabolic determinants. These findings suggest that phenotypic variability in KS might be primarily driven by chromosomal aneuploidy and primary testicular dysfunction rather than AR repeat length.
Graziani A et al., 2026·Journal of clinical medicine·Free full text on PubMed Central
Erectile dysfunction (ED) is increasingly recognized as an early manifestation of systemic vascular disease and might represent a window for cardiovascular risk assessment. Dynamic penile colour Doppler ultrasound (PCDU) provides quantitative arterial and venous parameters that could reflect subclinical vascular impairment. We investigated the association between PCDU parameters and estimated cardiovascular risk in men with ED. In this single-center retrospective observational study, 275 men undergoing PCDU for ED were evaluated. Clinical characteristics, biochemical data, and QRISK3 10-year cardiovascular risk scores were collected. Mean peak systolic velocity (PSV), end-diastolic velocity (EDV), and resistive index (RI) were analyzed. Correlation analyses, logistic regression using a QRISK3 ≥ 10% threshold, linear regression models, age-stratified analyses, and receiver operating characteristic (ROC) curve analyses were performed. Patients with impaired PSV (<35 cm/s) were older and exhibited higher QRISK3 scores and a greater prevalence of diabetes mellitus and previous cardiovascular events. Mean PSV was inversely correlated with QRISK3 (r = -0.203, p < 0.01) and was associated with higher cardiovascular risk categories in unadjusted logistic regression (β = -0.016, p = 0.048), but not after adjustment for age and diabetes mellitus. ROC analysis showed modest discrimination of increased cardiovascular risk (AUC = 0.60). The addition of PSV to a model including age and diabetes resulted in minimal improvement in discrimination (AUC 0.966 vs. 0.968). Age-stratified analysis demonstrated a significant association between lower PSV and higher cardiovascular risk only in patients ≤60 years. A progressive increase in QRISK3 was observed according to the number of abnormal Doppler parameters (p = 0.013). PCDU parameters reflect the overall cardiovascular risk burden in men with ED. Although not independent predictors beyond traditional risk factors, penile Doppler abnormalities might identify a vascular phenotype associated with higher estimated cardiovascular risk, particularly in younger individuals. These findings support the role of comprehensive vascular assessment in selected patients with ED.
Barbato M et al., 2026·Frontiers in Reproductive Health·Free full text on PubMed Central
Couple infertility is a common clinical condition that is too often treated with assisted reproductive techniques (ARTs) without a proper evaluation of both male and female factors. To improve the likelihood of natural conception, fertility awareness methods (FAMs) are widely used. We performed a multicenter prospective study enrolling couples with primary idiopathic infertility who were seeking natural conception. Participants were followed for 12 months using FAMs, and their outcomes were compared with those of couples who only used ARTs. The aim of our study was to evaluate the pregnancy rate after 12 months among couples with idiopathic infertility using FAMs compared with those who immediately pursued ARTs. We evaluated 41 couples in the FAM group and 56 couples in the ART group. In the FAM group, we reported a pregnancy rate (PR) of 51.22%. Among women aged <34 years, we reported a PR of 90.9%, while it decreased to 36.7% among women aged 35-39 years. In the ART group, 10 couples achieved pregnancy (PR 17.8%). Within this group, we reported a PR of 30% among women aged <34 years and 17.4% among women aged 35-39 years. After 12 months of unprotected intercourse without spontaneous conception in women younger than 35 years or after 6 months in women aged 35-39 years, couples should undergo a complete multidisciplinary diagnostic evaluation involving both the male and female partners. If a diagnosis of idiopathic infertility is established at the end of this process, couples (especially younger ones) may be advised to wait an additional 12 months while using FAMs, as no advantage has been observed with direct access to ARTs. They could then be referred to ART if a spontaneous pregnancy is not achieved during this period.
27 healthy young Italian women were studied to evaluate their ability to identify symptomatically the potentially fertile phase of the menstrual cycle by self observation of their cervical mucus pattern as described in the Ovulation Method Billings. The women's observations were correlated with daily plasma levels of FSH, LH, estradiol-17 beta and progesterone. Ovulation was considered to occur on the day following the LH peak. The hormonal assays revealed that 2 of the 34 cycles studied were anovulatory. 24 of the 27 subjects in the study group demonstrated their ability to recognize the onset of the mucus discharge and the peak symptom from the first cycle after teaching, another two from the second cycle. The only other subject contributed an anovulatory cycle in which the hormonal assay confirmed the accuracy of her mucus observations. In the study, the mean interval between the time of ovulation as assessed and the peak symptom recorded by the subjects was 0.0 days, with a range from -2 to +1 days. The mean time interval from the first recorded symptom to the estimated day of ovulation was 6.0 days, with a range from 3 to 10 days. The study shows that young Italian women can be taught to recognize their cervical mucus pattern as described in the Ovulation Method Billings. The accuracy of their observations is demonstrated by the hormonal assays. The study also confirms the conclusion reached in earlier similar studies that there is a direct correlation between the cervical mucus symptom and the potentially fertile phase of the cycle. Research is currently being conducted on a larger number of couples employing the Ovulation Method Billings to actually regulate their fertility.
Vigil P et al., 2012·Linacre Q·Free full text on PubMed Central
Fertility awareness constitutes fundamental knowledge for every woman and is an important tool for health professionals. The objective of this review is to show how fertility awareness can be useful in the assessment of a woman's health. The main techniques for detecting ovulation are explained, and then the events that characterize a normal menstrual cycle are discussed. The relevance of cervical mucus from the perspective of female fertility is highlighted. Finally, the usefulness of fertility awareness 1) to identify fertile and infertile periods, 2) to help to detect several pathologies, and 3) in regards to how it exerts an important role in the success of programs in education for affectivity and sexuality are discussed.
Fernandez-Hermida Y et al., 2018·Protein & Peptide Letters
Cervical Mucus (CM) is a viscous fluid produced by the secretory cells of the cervical crypts. The CM undergoes modifications throughout the cycle that make it have different biochemical and biophysical characteristics, becoming a crucial element for the identification of ovulation. Since CM is rich in secreted proteins, it may represent moreover a source of biomarkers for female reproductive tract diseases. This review is an attempt to collect relevant knowledge about the physicochemical properties and functions of the cervical mucus, including its important role as a clinical marker of female fertility, and draws attention to CM as a source of potential proteomic biomarkers. All the assessed studies evidenced that the observation of the CM allows the identification of the days with the highest probability of pregnancy. CM proteome changes throughout the menstrual cycle have been revealed. Few proteomic studies on the constitutive protein composition of CM of fertile women have been conducted to date. In the CM of patients affected by endometriosis have been identified some proteins that could represent potential biomarkers of the disease. There is still limited knowledge about the physicochemical properties and functions of the CM and how these undergo to changes during menstrual cycle. CM is a reliable predictor of fertility. Further characterization of CM proteins would contribute to a better understanding of the key role they have on fertility, reproduction and biological regulation. CM may represent moreover a source of biomarkers for gynecological diseases.
Najmabadi S et al., 2021·Hum Reprod·Free full text on PubMed Central
What is the normal range of cervical mucus patterns and number of days with high or moderate day-specific probability of pregnancy (if intercourse occurs on a specific day) based on cervical mucus secretion, in women without known subfertility, and how are these patterns related to parity and age? The mean days of peak type (estrogenic) mucus per cycle was 6.4, the mean number of potentially fertile days was 12.1; parous versus nulliparous, and younger nulliparous (<30 years) versus older nulliparous women had more days of peak type mucus, and more potentially fertile days in each cycle. The rise in estrogen prior to ovulation supports the secretion of increasing quantity and estrogenic quality of cervical mucus, and the subsequent rise in progesterone after ovulation causes an abrupt decrease in mucus secretion. Cervical mucus secretion on each day correlates highly with the probability of pregnancy if intercourse occurs on that day, and overall cervical mucus quality for the cycle correlates with cycle fecundability. No prior studies have described parity and age jointly in relation to cervical mucus patterns. STUDY DESIGN, SIZE, This study is a secondary data analysis, combining data from three cohorts of women: 'Creighton Model MultiCenter Fecundability Study' (CMFS: retrospective cohort, 1990-1996), 'Time to Pregnancy in Normal Fertility' (TTP: randomized trial, 2003-2006), and 'Creighton Model Effectiveness, Intentions, and Behaviors Assessment' (CEIBA: prospective cohort, 2009-2013). We evaluated cervical mucus patterns and estimated fertile window in 2488 ovulatory cycles of 528 women, followed for up to 1 year. PARTICIPANTS/MATERIALS, SETTING, Participants were US or Canadian women age 18-40 years, not pregnant, and without any known subfertility. Women were trained to use a standardized protocol (the Creighton Model) for daily vulvar observation, description, and recording of cervical mucus. The mucus peak day (the last day of estrogenic quality mucus) was used as the estimated day of ovulation. We conducted dichotomous stratified analyses for cervical mucus patterns by age, parity, race, recent oral contraceptive use (within 60 days), partial breast feeding, alcohol, and smoking. Focusing on the clinical characteristics most correlated to cervical mucus patterns, linear mixed models were used to assess continuous cervical mucus parameters and generalized linear models using Poisson regression with robust variance were used to assess dichotomous outcomes, stratifying by women's parity and age, while adjusting for recent oral contraceptive use and breast feeding. MAIN The majority of women were <30 years of age (75.4%) (median 27; IQR 24-29), non-Hispanic white (88.1%), with high socioeconomic indicators, and nulliparous (70.8%). The mean (SD) days of estrogenic (peak type) mucus per cycle (a conservative indicator of the fertile window) was 6.4 (4.2) days (median 6; IQR 4-8). The mean (SD) number of any potentially fertile days (a broader clinical indicator of the fertile window) was 12.1 (5.4) days (median 11; IQR 9-14). Taking into account recent oral contraceptive use and breastfeeding, nulliparous women age ≥30 years compared to nulliparous women age <30 years had fewer mean days of peak type mucus per cycle (5.3 versus 6.4 days, P = 0.02), and fewer potentially fertile days (11.8 versus 13.9 days, P < 0.01). Compared to nulliparous women age <30 years, the likelihood of cycles with peak type mucus ≤2 days, potentially fertile days ≤9, and cervical mucus cycle score (for estrogenic quality of mucus) ≤5.0 were significantly higher among nulliparous women age ≥30 years, 1.90 (95% confidence interval (CI) 1.18, 3.06); 1.46 (95% CI 1.12, 1.91); and 1.45 (95% CI 1.03, 2.05), respectively. Between parous women, there was little difference in mucus parameters by age. Thresholds set a priori for within-woman variability of cervical mucus parameters by cycle were examined as follows: most minus fewest days of peak type mucus >3 days (exceeded by 72% of women), most minus fewest days of non-peak type mucus >4 days (exceeded by 54% of women), greatest minus least cervical mucus cycle score >4.0 (exceeded by 73% of women), and most minus fewest potentially fertile days >8 days (found in 50% of women). Race did not have any association with cervical mucus parameters. Recent oral contraceptive use was associated with reduced cervical mucus cycle score and partial breast feeding was associated with a higher number of days of mucus (both peak type and non-peak type), consistent with prior research. Among the women for whom data were available (CEIBA and TTP), alcohol and tobacco use had minimal impact on cervical mucus parameters. LIMITATIONS, We did not have data on some factors that may impact ovulation, hormone levels, and mucus secretion, such as physical activity and body mass index. We cannot exclude the possibility that some women had unknown subfertility or undiagnosed gynecologic disorders. Only 27 women were age 35 or older. Our study participants were geographically dispersed but relatively homogeneous with regard to race, ethnicity, income, and educational level, which may limit the generalizability of the findings. Patterns of cervical mucus secretion observed by women are an indicator of fecundity and the fertile window that are consistent with the known associations of age and parity with fecundity. The number of potentially fertile days (12 days) is likely greater than commonly assumed, while the number of days of highly estrogenic mucus (and higher probability of pregnancy) correlates with prior identifications of the fertile window (6 days). There may be substantial variability in fecundability between cycles for the same woman. Future work can use cervical mucus secretion as an indicator of fecundity and should investigate the distribution of similar cycle parameters in women with various reproductive or gynecologic pathologies. STUDY FUNDING/COMPETING INTEREST(S): Funding for the three cohorts analyzed was provided by the Robert Wood Johnson Foundation (CMFS), the Eunice Kennedy Shriver National Institute of Child Health and Human Development (TTP), and the Office of Family Planning, Office of Population Affairs, Health and Human Services (CEIBA). The authors declare that they have no conflict of interest. N/A.