Good practice recommendations on add-ons in reproductive medicine†.

STUDY QUESTION: Which add-ons are safe and effective to be used in ART treatment? SUMMARY ANSWER: Forty-two recommendations were formulated on the use of add-ons in the diagnosis of fertility problems, the IVF laboratory and clinical management of IVF treatment. WHAT IS KNOWN ALREADY: The innovative nature of ART combined with the extremely high motivation of the patients has opened the door to the wide application of what has become known as 'add-ons' in reproductive medicine. These supplementary options are available to patients in addition to standard fertility procedures, typically incurring an additional cost. A diverse array of supplementary options is made available, encompassing tests, drugs, equipment, complementary or alternative therapies, laboratory procedures, and surgical interventions. These options share the common aim of stating to enhance pregnancy or live birth rates, mitigate the risk of miscarriage, or expedite the time to achieving pregnancy. STUDY DESIGN, SIZE, DURATION: ESHRE aimed to develop clinically relevant and evidence-based recommendations focusing on the safety and efficacy of add-ons currently used in fertility procedures in order to improve the quality of care for patients with infertility. PARTICIPANTS/MATERIALS, SETTING, METHODS: ESHRE appointed a European multidisciplinary working group consisting of practising clinicians, embryologists, and researchers who have demonstrated leadership and expertise in the care and research of infertility. Patient representatives were included in the working group. To ensure that the guidelines are evidence-based, the literature identified from a systematic search was reviewed and critically appraised. In the absence of any clear scientific evidence, recommendations were based on the professional experience and consensus of the working group. The guidelines are thus based on the best available evidence and expert agreement. Prior to publication, the guidelines were reviewed by 46 independent international reviewers. A total of 272 comments were received and incorporated where relevant. MAIN RESULTS AND THE ROLE OF CHANCE: The multidisciplinary working group formulated 42 recommendations in three sections; diagnosis and diagnostic tests, laboratory tests and interventions, and clinical management. LIMITATIONS, REASONS FOR CAUTION: Of the 42 recommendations, none could be based on high-quality evidence and only four could be based on moderate-quality evidence, implicating that 95% of the recommendations are supported only by low-quality randomized controlled trials, observational data, professional experience, or consensus of the development group. WIDER IMPLICATIONS OF THE FINDINGS: These guidelines offer valuable direction for healthcare professionals who are responsible for the care of patients undergoing ART treatment for infertility. Their purpose is to promote safe and effective ART treatment, enabling patients to make informed decisions based on realistic expectations. The guidelines aim to ensure that patients are fully informed about the various treatment options available to them and the likelihood of any additional treatment or test to improve the chance of achieving a live birth. STUDY FUNDING/COMPETING INTEREST(S): All costs relating to the development process were covered from ESHRE funds. There was no external funding of the development process or manuscript production. K.L. reports speakers fees from Merck and was part of a research study by Vitrolife (unpaid). T.E. reports consulting fees from Gynemed, speakers fees from Gynemed and is part of the scientific advisory board of Hamilton Thorne. N.P.P. reports grants from Merck Serono, Ferring Pharmaceutical, Theramex, Gedeon Richter, Organon, Roche, IBSA and Besins Healthcare, speakers fees from Merck Serono, Ferring Pharmaceutical, Theramex, Gedeon Richter, Organon, Roche, IBSA and Besins Healthcare. S.R.H. declares being managing director of Fertility Europe, a not-for-profit organization receiving financial support from ESHRE. I.S. is a scientific advisor for and has stock options from Alife Health, is co-founder of IVFvision LTD (unpaid) and received speakers' fee from the 2023 ART Young Leader Prestige workshop in China. A.P. reports grants from Gedeon Richter, Ferring Pharmaceuticals and Merck A/S, consulting fees from Preglem, Novo Nordisk, Ferring Pharmaceuticals, Gedeon Richter, Cryos and Merck A/S, speakers fees from Gedeon Richter, Ferring Pharmaceuticals, Merck A/S, Theramex and Organon, travel fees from Gedeon Richter. The other authors disclosed no conflicts of interest. DISCLAIMER: This Good Practice Recommendations (GPRs) document represents the views of ESHRE, which are the result of consensus between the relevant ESHRE stakeholders and are based on the scientific evidence available at the time of preparation.ESHRE GPRs should be used for information and educational purposes. They should not be interpreted as setting a standard of care or bedeemedinclusive of all proper methods of care, or be exclusive of other methods of care reasonably directed to obtaining the same results.Theydo not replace the need for application of clinical judgement to each individual presentation, or variations based on locality and facility type.Furthermore, ESHRE GPRs do not constitute or imply the endorsement, or favouring, of any of the included technologies by ESHRE.

Reliability and consistency of a validated sun exposure questionnaire in a population-based Danish sample.

An important feature of questionnaire validation is reliability. To be able to measure a given concept by questionnaire validly, the reliability needs to be high. The objectives of this study were to examine reliability of attitude and knowledge and behavioral consistency of sunburn in a developed questionnaire for monitoring and evaluating population sun-related behavior. Sun related behavior, attitude and knowledge was measured weekly by a questionnaire in the summer of 2013 among 664 Danes. Reliability was tested in a test-retest design. Consistency of behavioral information was tested similarly in a questionnaire adapted to measure behavior throughout the summer. The response rates for questionnaire 1, 2 and 3 were high and the drop out was not dependent on demographic characteristic. There was at least 73% agreement between sunburns in the measurement week and the entire summer, and a possible sunburn underestimation in questionnaires summarizing the entire summer. The participants underestimated their outdoor exposure in the evaluation covering the entire summer as compared to the measurement week. The reliability of scales measuring attitude and knowledge was high for majority of scales, while consistency in protection behavior was low. To our knowledge, this is the first study to report reliability for a completely validated questionnaire on sun-related behavior in a national random population based sample. Further, we show that attitude and knowledge questions confirmed their validity with good reliability, while consistency of protection behavior in general and in a week's measurement was low.

Symptom load and individual symptoms before and after repair of parastomal hernia: a prospective single centre study.

AIM: The symptom load and individual symptoms before and after repair of parastomal hernia were investigated. METHOD: Stoma-related symptoms were prospectively recorded before repair of a parastomal hernia and at 10 days and 6 months postoperatively: leakage, skin problems, difficulty with the appliance, limitation of activity, difficulty with clothing, cosmetic complaints, social restriction, erratic action of the stoma, a bearing-down sensation at the site of the stoma and pain. Episodes of intermittent bowel obstruction and difficulty with irrigation were also recorded. Patients were seen at 1, 2 and 3 years and were examined for recurrent parastomal herniation. RESULTS: Of 131 consecutive patients referred to a specialized centre for treatment of parastomal bulging, 61 underwent parastomal hernia repair. Forty-eight patients were treated with the Sugarbaker technique. Six different symptoms were present in more than half the patients before surgery. The overall symptom load decreased significantly from a median of 4 [interquartile range (IQR) 2.5-6] preoperatively to 2 (IQR 1-3) on postoperative day 10 and 1 (IQR 0-2) at 6 months, P < 0.001. The number of symptoms decreased in 93% of patients; in 5% there was no change and in 2% symptoms increased. Skin problems and leakage were the only symptoms that were not significantly reduced. The overall recurrence rate of herniation was 5/48 (10%) at a median of 12 (IQR 6-24) months. CONCLUSION: The preoperative symptom load was high and this fell after repair in over 90% of patients. Recurrence occurred in 10% of patients within 2 years of repair. The study emphasizes the importance of detailed knowledge of the symptoms of parastomal hernia when addressing and managing patients' problems and complaints.

The validated sun exposure questionnaire: association of objective and subjective measures of sun exposure in a Danish population-based sample.

BACKGROUND: Few questionnaires used in monitoring sun-related behaviour have been tested for validity. OBJECTIVES: We established the criteria validity of a questionnaire developed for monitoring population sun-related behaviour. METHODS: During May-August 2013, 664 Danes wore a personal electronic ultraviolet radiation (UVR) dosimeter for 1 week that measured their outdoor time and dose of erythemal UVR exposure. In the following week, they answered a questionnaire on their sun-related behaviour in the measurement week. RESULTS: Outdoor time measured by dosimetry correlated strongly with both outdoor time and the developed exposure scale measured in the questionnaire. Exposure measured in standard erythema dose (SED) by dosimetry correlated strongly with the exposure scale. In a linear regression model of UVR (SED) received, 41% of the variation was explained by skin type, age, week of participation and exposure scale, with exposure scale as the main contributor. The weekly sunburn fraction correlated strongly with the number of ambient sun hours (r = 0·73, P < 0·001). CONCLUSIONS: This criteria-validated questionnaire provides evidence of the exposure that the questionnaire aimed to measure. The evidence provided showed a strong link between the objectively measured behaviour and the behaviour measured by this survey construct. The questionnaire is the first validated tool to measure the UVR exposure in a national population-based sample.

Infertile women below the age of 40 have similar anti-Müllerian hormone levels and antral follicle count compared with women of the same age with no history of infertility.

STUDY QUESTION: Do infertile patients below the age of 40 years have a lower ovarian reserve, estimated by anti-Müllerian hormone (AMH) and total antral follicle count (AFC), than women of the same age with no history of infertility? SUMMARY ANSWER: Serum AMH and AFC were not lower in infertile patients aged 20-39 years compared with a control group of the same age with no history of infertility. WHAT IS KNOWN ALREADY?: The management of patients with a low ovarian reserve and a poor response to controlled ovarian stimulation (COS) remains a challenge in assisted reproductive technologies (ART). Both AMH levels and AFC reflect the ovarian reserve and are valuable predictors of the ovarian response to exogenous gonadotrophins. However, there is a large inter-individual variation in the age-related depletion of the ovarian reserve and a broad variability in the levels of AMH and AFC compatible with conception. Women with an early depletion of the ovarian reserve may experience infertility as a consequence of postponement of childbearing. Thus, low ovarian reserve is considered to be overrepresented among infertile patients. STUDY DESIGN, SIZE, DURATION: A prospective cohort study including 382 women with a male partner referred to fertility treatment at Rigshospitalet, Copenhagen, Denmark during 2011-2013 compared with a control group of 350 non-users of hormonal contraception with no history of infertility recruited during 2008-2010. PARTICIPANTS/MATERIALS, SETTING, METHODS: Included patients and controls were aged 20-39 years. Women with polycystic ovary syndrome were excluded. On Cycle Days 2-5, AFC and ovarian volume were measured by transvaginal sonography, and serum levels of AMH, FSH and LH were assessed. MAIN RESULTS AND THE ROLE OF CHANCE: Infertile patients had similar AMH levels (11%, 95% confidence interval (CI): -1;24%) and AFC (1%, 95% CI: -7;8%) compared with controls with no history of infertility in an age-adjusted linear regression analysis. The prevalence of very low AMH levels (<5 pmol/l) was similar in the two cohorts (age-adjusted odds ratio: 0.9, 95% CI: 0.5;1.7). The findings persisted after adjustment for smoking status, body mass index, gestational age at birth, previous conception and chronic disease in addition to age. LIMITATIONS, REASON FOR CAUTION: The comparison of ovarian reserve parameters in women recruited at different time intervals could be a reason for caution. However, all women were examined at the same centre using the same sonographic algorithm and AMH immunoassay. WIDER IMPLICATIONS OF THE FINDINGS: This study indicates that the frequent observation of patients with a poor response to COS in ART may not be due to an overrepresentation of women with an early depletion of the ovarian reserve but rather a result of the expected age-related decline in fertility. STUDY FUNDING/COMPETING INTERESTS: The study received funding from MSD and the Interregional European Union (EU) projects 'ReproSund' and 'ReproHigh'. The authors have no conflict of interest. TRIAL REGISTRATION NUMBER: Not applicable.

High-risk sun-tanning behaviour: a quantitative study in Denmark, 2008-2011.

OBJECTIVES: The incidences of melanoma and non-melanoma skin cancer have increased markedly over the past 30 years. The main risk factor is ultraviolet radiation from the sun and from sunbeds. The Danish Sun Safety campaign was launched in 2007 to curb this development by reducing the exposure of adolescents and young children. In this study, the characteristics of high-risk sun-tanning behaviour were assessed and the effect of the campaign was determined. STUDY DESIGN: Cross-sectional study. METHODS: Data from annual Internet surveys were compiled in 2008-2011 of 18, 685 15-64-year-old Danes. A tanning index based on sunbed use and intentional tanning in and outside Denmark was the outcome measure in a linear regression model, which included age, gender, skin type, education, income and survey year as exposure variables. RESULTS: High-risk tanning behaviour was associated with female gender, younger age, shorter education, skin type 3 or 4, higher income, smaller household and living in larger cities. The tanning index, where 100 represent high-risk behaviour, increased by 13.45 points for women as compared with men, dropped by 1.35 points for each 5-year increase in age, rose by 20.72 points for skin type 4 as compared with type 1 and increased by 10.33 points with an income >€105, 409 as compared with <€26, 352. High-risk behaviour decreased during the study period, especially among women and younger people. CONCLUSIONS: High-risk sun-tanning behaviour is linked to certain personal and social characteristics. After initiation of the Danish Sun Safety Campaign in 2007, this high-risk behaviour decreased, especially in the groups initially targeted by the campaign. The campaign may thus reduce the future incidence of melanoma and non-melanoma skin cancer.

The prevalence of polycystic ovary syndrome in a normal population according to the Rotterdam criteria versus revised criteria including anti-Mullerian hormone.

STUDY QUESTION: What is the prevalence in a normal population of polycystic ovary syndrome (PCOS) according to the Rotterdam criteria versus revised criteria including anti-Müllerian hormone (AMH)? SUMMARY ANSWER: The prevalence of PCOS was 16.6% according to the Rotterdam criteria. When replacing the criterion for polycystic ovaries by antral follicle count (AFC) > 19 or AMH > 35 pmol/l, the prevalence of PCOS was 6.3 and 8.5%, respectively. WHAT IS KNOWN ALREADY?: The Rotterdam criteria state that two out of the following three criteria should be present in the diagnosis of PCOS: oligo-anovulation, clinical and/or biochemical hyperandrogenism and polycystic ovaries (AFC ≥ 12 and/or ovarian volume >10 ml). However, with the advances in sonography, the relevance of the AFC threshold in the definition of polycystic ovaries has been challenged, and AMH has been proposed as a marker of polycystic ovaries in PCOS. STUDY DESIGN, SIZE, DURATION: From 2008 to 2010, a prospective, cross-sectional study was performed including 863 women aged 20-40 years and employed at Copenhagen University Hospital, Rigshospitalet, Denmark. PARTICIPANTS/MATERIAL, SETTING, METHODS: We studied a subgroup of 447 women with a mean (±SD) age of 33.5 (±4.0) years who were all non-users of hormonal contraception. Data on menstrual cycle disorder and the presence of hirsutism were obtained. On cycle Days 2-5, or on a random day in the case of oligo- or amenorrhoea, sonographic and endocrine parameters were measured. MAIN RESULTS AND THE ROLE OF CHANCE: The prevalence of PCOS was 16.6% according to the Rotterdam criteria. PCOS prevalence significantly decreased with age from 33.3% in women < 30 years to 14.7% in women aged 30-34 years, and 10.2% in women ≥ 35 years (P < 0.001). In total, 53.5% fulfilled the criterion for polycystic ovaries with a significant age-related decrease from 69.0% in women < 30 years to 55.8% in women aged 30-34 years, and 42.8% in women ≥ 35 years (P < 0.001). AMH or age-adjusted AMH Z-score was found to be a reliable marker of polycystic ovaries in women with PCOS according to the Rotterdam criteria [area under the curve (AUC) 0.994; 95% confidence interval (CI): 0.990-0.999] and AUC 0.992 (95% CI: 0.987-0.998), respectively], and an AMH cut-off value of 18 pmol/l and AMH Z-score of -0.2 showed the best compromise between sensitivity (91.8 and 90.4%, respectively) and specificity (98.1 and 97.9%, respectively). In total, AFC > 19 or AMH > 35 occurred in 17.7 and 23.0%, respectively. The occurrence of AFC > 19 or AMH > 35 in the age groups < 30, 30-34 and ≥ 35 years was 31.0 and 35.7%, 18.8 and 21.3%, and 9.6 and 18.7%, respectively. When replacing the Rotterdam criterion for polycystic ovaries by AFC > 19 or AMH > 35 pmol/l, the prevalence of PCOS was 6.3 or 8.5%, respectively, and in the age groups < 30, 30-34 and ≥ 35 years, the prevalences were 17.9 and 22.6%, 3.6 and 5.6%, and 3.6 and 4.8%, respectively. LIMITATIONS, REASON FOR CAUTION: The participants of the study were all health-care workers, which may be a source of selection bias. Furthermore, the exclusion of hormonal contraceptive users from the study population may have biased the results, potentially excluding women with symptoms of PCOS. WIDER IMPLICATIONS OF THE FINDINGS: AMH may be used as a marker of polycystic ovaries in PCOS. However, future studies are needed to validate AMH threshold levels, and AMH Z-score may be appropriate to adjust for the age-related decline in the AFC. STUDY FUNDING/COMPETING INTEREST(S): None. TRIAL REGISTRATION NUMBER: Not applicable.

A 10-year follow up of reproductive function in women treated for childhood cancer.

Previously, this study group found that female childhood cancer survivors could be at risk of early cessation of fertility. The aim of the present study was to evaluate reproductive function in the same group of survivors 10 years after the initial study. Of the original cohort of 100, 71 were re-examined. Thirty-six survivors reported regular menstrual cycles. When they were compared with 210 controls, they differed significantly in antral follicle count (AFC) (median 15 versus 18, P=0.047) but not in anti-Müllerian hormone (AMH) (median 13.0 versus 17.8 pmol/l). Survivors cured with minimal gonadotoxic treatment had significantly higher AMH and AFC compared with survivors cured with either potentially gonadotoxic treatment or treatment including alkylating chemotherapy and ovarian irradiation (20.0, 5.8 and <3 pmol/l, P<0.001; and 15, 9 and 2, P=0.03, respectively). Thirty-eight survivors had achieved at least one live birth. Complicated second-trimester abortions (n=4) were observed primarily in survivors cured with radiotherapy affecting pelvic organs. In conclusion, childhood cancer survivors have signs of diminished ovarian reserve. However, if the ovarian function is preserved in the early to mid-twenties, it is likely to persist until the mid-thirties, giving a good chance of childbearing.

Ovarian antral follicle subclasses and anti-mullerian hormone during normal reproductive aging.

CONTEXT: The interindividual variation in the age-related decline of ovarian follicles is wide. Hence, it is important to identify reliable, sensitive, and specific markers to assess the ovarian reserve of the individual woman. OBJECTIVE: The aim of this study was to characterize the relation between age and ovarian reserve parameters in a population of healthy women with regular menstrual cycle. DESIGN AND SETTING: We conducted a prospective, population-based, cross-sectional study. PARTICIPANTS: A total of 366 health care workers aged 21-41 years employed at a University Hospital were included. INTERVENTIONS: There were no interventions. MAIN OUTCOME MEASURES: Serum anti-Müllerian hormone (AMH) concentration, antral follicle count (AFC), antral follicle size categories (small: 2-4 mm; intermediate: 5-7 mm; and large: 8-10 mm), and ovarian volume were measured. RESULTS: Serum AMH level declined by 5.6% per year (95% confidence interval 3.7-7.4%, P < .001), AFC (2-10 mm) declined by 4.4% per year (3.2-5.7%, P < .001), and ovarian volume declined by 1.1% per year (0.2-2.0, P = .002), respectively. The mean proportion of small follicles decreased with age (P = .04), the proportion of intermediate follicles displayed no significant change with age (P = .58), and the mean proportion of large follicles increased with age (P < .001). The prevalence of large follicles increased with decreasing serum AMH concentration [odds ratio 1.04 per 1 pmol/L (1.02-1.06), P < .001, area under the curve 0.66], and with decreasing total AFC [odds ratio 1.04 per follicle (1.02-1.05), P < .001, area under the curve 0.62]. CONCLUSION: Chronological age was inversely related to serum AMH concentration, total AFC, and ovarian volume. Subclasses of AFC sized 2-4 and 5-7 mm decreased with increasing age, whereas AFC sized 8-10 mm increased with increasing age. Within AFC, a shift toward larger follicles with increasing age was observed. The occurrence of large follicles was more strongly related to biological age in terms of AMH and AFC than chronological age.

Transmission electron microscopy specimen preparation method for multiphase porous functional ceramics.

An optimum method is proposed to prepare thin foil transmission electron microscopy (TEM) lamellae of multiphase porous functional ceramics: prefilling the pore space of these materials with an epoxy resin prior to focused ion beam milling. Several advantages of epoxy impregnation are demonstrated by successful preparation of TEM specimens that maintain the structural integrity of the entire lamella. Feasibility of the TEM alignment procedure is demonstrated, and ideal TEM analyses are illustrated on solid oxide fuel cell and solid oxide electrolysis cell materials. Some potential drawbacks of the TEM specimen preparation method are listed for other samples.

Maternal menopause as a predictor of anti-Mullerian hormone level and antral follicle count in daughters during reproductive age.

STUDY QUESTION: Is the ovarian reserve in a woman at a given age associated with her mother's age at menopause? SUMMARY ANSWER: We demonstrated a significant, positive association between age at maternal menopause and serum anti-Müllerian hormone (AMH) levels and antral follicle count (AFC) in daughters. The rate of decline in serum-AMH level and AFC is also associated with age at maternal menopause. WHAT IS KNOWN AND WHAT THIS PAPER ADDS: The association between menopausal age in mothers and daughters has been established through several epidemiological studies. This paper shows that early maternal menopause is related to an advanced depletion of the ovarian reserve and that late maternal menopause is related to a delayed depletion. STUDY DESIGN AND SIZE: Cross-sectional data were obtained from a prospective cohort study of 863 women. The study comprised 527 participants from this prospective cohort whose mothers' age at natural menopause was known. PARTICIPANTS, SETTING AND METHODS: Participants were recruited from female health care workers aged 20-40 years employed at Copenhagen University Hospital, Rigshospitalet, and were enrolled in the study between September 2008 and February 2010. The response rate was 52.1%. Endocrine and ovarian parameters related to reproductive ageing (AMH and AFC) were assessed by serum AMH analyses and transvaginal ovarian sonography on cycle Day 2-5. Data on reproductive history, including age at natural maternal menopause, were obtained through an internet-based questionnaire. We used an analysis of covariance model with serum-AMH and AFC as outcomes, age as the quantitative predictor and onset of maternal menopause as the categorical predictor, with further adjustments for BMI, use of oral contraceptives, participants' smoking habits and prenatal smoking exposure. MAIN FINDINGS: We found a significant effect of age at maternal menopause on both serum AMH levels (P < 0.001) and AFC (P = 0.005). Median serum-AMH concentration declined by 8.6% per year [95% confidence interval (CI): 6.4-10.8%, P < 0.001] in the group with early maternal menopausal age (≤ 45 years), by 6.8% per year (95% CI: 5.0-8.6%, P < 0.001) in the group with normal maternal menopausal age (46-54 years) and by 4.2% per year (95% CI: 2.0-6.4%, P < 0.001) in the group with late maternal menopausal age (≥ 55 years). Median AFC declined by 5.8% per year (95% CI: 4.0-7.5%, P < 0.001) in the group with early maternal menopausal age (≤ 45 years), by 4.7% per year (95% CI: 3.3-6.1%, P < 0.001) in the group with normal maternal menopausal age (46-54 years) and by 3.2% per year (95% CI: 1.4-4.9%, P < 0.001) in the group with late maternal age (≥ 55 years) at menopause. BIAS, LIMITATIONS AND GENERALIZABILITY: Information on 'age at maternal menopause' was obtained retrospectively and may be prone to recall bias and digit preference. The study population consisted of health care workers, which implies a potential selection bias. Finally, the cross-sectional nature of the data limits the generalizability. STUDY FUNDING/POTENTIAL COMPETING INTERESTS: This study was co-financed by PhD scholarships where funding was covered by the Danish Agency for Science, Technology and Innovation, Copenhagen Graduate School of Health Science (CGSHS) and the Fertility Clinic at Copenhagen University Hospital, Rigshopitalet. No competing interests are declared.

Ovarian reserve parameters: a comparison between users and non-users of hormonal contraception.

It remains controversial whether anti-Müllerian hormone (AMH) concentration is influenced by hormonal contraception. This study quantified the effect of hormonal contraception on both endocrine and sonographic ovarian reserve markers in 228 users and 504 non-users of hormonal contraception. On day 2-5 of the menstrual cycle or during withdrawal bleeding, blood sampling and transvaginal sonography was performed. After adjusting for age, ovarian reserve parameters were lower among users than among non-users of hormonal contraception: serum AMH concentration by 29.8% (95% CI 19.9 to 38.5%), antral follicle count (AFC) by 30.4% (95% CI 23.6 to 36.7%) and ovarian volume by 42.2% (95% CI 37.8 to 46.3%). AFC in all follicle size categories (small, 2-4 mm; intermediate, 5-7 mm; large, 8-10 mm) was lower in users than in non-users of hormonal contraception. A negatively linear association was observed between duration of hormonal-contraception use and ovarian reserve parameters. No dose-response relation was found between the dose of ethinyloestradiol and AMH or AFC. This study indicates that ovarian reserve markers are lower in women using sex steroids for contraception. Thus, AMH concentration and AFC may not retain their accuracy as predictors of ovarian reserve in women using hormonal contraception. Serum anti-Müllerian hormone (AMH) concentration is an indirect marker of the number of small follicles in the ovary and thereby the ovarian reserve. The AMH concentration is now widely used as one of the markers of the ovarian reserve in ovarian hormonal stimulation regimens. Hence the AMH concentration in a patient is used to decide the dose of the ovarian hormonal stimulation prior to IVF treatment. In some infertile patients, hormonal contraception is used prior to ovarian hormonal stimulation and therefore it is important to clarify whether serum AMH concentration is influenced by the use of sex steroids. The aim of this study was to quantify the potential effect of hormonal contraception on the ovarian function by hormonal analyses and ovarian ultrasound examination. Examinations were performed in the early phase of the menstrual cycle or the hormone-free interval of hormonal contraception. We compared the AMH concentration, the antral follicle count (AFC) and the ovarian volume in 228 users versus 504 non-users of hormonal contraception. Users of hormonal contraception had 29.8% lower AMH concentration, 30.4% lower AFC and 42.2% lower ovarian volume than non-users. These findings were more pronounced with increasing duration of hormonal contraception. No dose-response relation was found between the dose of ethinylestradiol and the impact on serum AMH and AFC. The study indicates that ovarian reserve markers are lower in women using sex steroids for contraception. Thus, serum AMH concentration and AFC may not retain their accuracy as predictors of the ovarian reserve in women using hormonal contraception.

Demographic and medical consequences of the postponement of parenthood.

BACKGROUND Across the developed world couples are postponing parenthood. This review assesses the consequences of delayed family formation from a demographic and medical perspective. One main focus is on the quantitative importance of pregnancy postponement. METHODS Medical and social science databases were searched for publications on relevant subjects such as delayed parenthood, female and male age, fertility, infertility, time to pregnancy (TTP), fetal death, outcome of medically assisted reproduction (MAR) and mental well-being. RESULTS Postponement of parenthood is linked to a higher rate of involuntary childlessness and smaller families than desired due to increased infertility and fetal death with higher female and male age. For women, the increased risk of prolonged TTP, infertility, spontaneous abortions, ectopic pregnancies and trisomy 21 starts at around 30 years of age with a more pronounced effects >35 years, whereas the increasing risk of preterm births and stillbirths starts at around 35 years with a more pronounced effect >40 years. Advanced male age has an important but less pronounced effect on infertility and adverse outcomes. MAR treatment cannot overcome the age-related decline in fecundity. CONCLUSIONS In general, women have partners who are several years older than themselves and it is important to focus more on the combined effect of higher female and male age on infertility and reproductive outcome. Increasing public awareness of the impact of advanced female and male age on the reproductive outcome is essential for people to make well-informed decisions on when to start family formation.

Novel polymorphisms in the GCKR gene and their influence on glucose and insulin levels in a Danish twin population.

AIM: The glucokinase regulatory protein gene is a candidate gene for Type 2 diabetes. This study reveals three new polymorphisms and examines the impact of one new and one known polymorphism on insulin secretion and parameters associated with the insulin resistance syndrome in Danish twins with different degrees of glucose tolerance. METHODS: Single nucleotide polymorphism detection was performed in 20 healthy subjects and in 20 subjects with Type 2 diabetes. The effect of the polymorphisms on lipid, glucose and insulin measures was studied in 566 same-sex twins aged 55-74 years. RESULTS: The new nucleotide (nt) 363 polymorphism was found only in subjects with impaired glucose tolerance and Type 2 diabetes. The nt 11216 polymorphism influenced insulin measured at 120 min during an oral glucose tolerance test (OGTT). Subjects with genotype C11216C/T11216C had 21% higher insulin values (P<0.05) than subjects with genotype T11216T. In twins discordant for this genotype, the C-allele was associated with significantly higher plasma insulin levels at all time points during the OGTT, higher beta-cell function and lower plasma glucose levels during the OGTT. CONCLUSION: The C-allele of nt 11216 polymorphism was associated with increased insulin secretion, and may therefore exert a potentially protective effect against Type 2 diabetes. This remains to be shown in a larger study population.

Reduced calcium tolerance in rat cardiomyocytes after myocardial infarction.

During ischaemia and reperfusion the intracellular Na+ concentration is elevated in the cardiomyocytes and the cells are depolarized, both favouring reverse mode Na,Ca-exchange loading of the cell with Ca2+. We examined whether cardiomyocytes from rats with congestive heart failure (CHF) and younger rats (HINCX) which both have a high expression of the Na,Ca-exchanger protein (NCX) showed reduced tolerance to extracellular Ca2+. The CHF was induced in Isofluran anaesthetized rats by left coronary artery ligation. Isolated cardiomyocytes were loaded with Fura-2AM and 140 mm Na+ and exposed to 0.05 mm Ca2+. Expression of the Na,Ca-exchanger protein was analysed. Fura-2 340/380 ratio rose more rapidly in HINCX and CHF than in SHAM, and the rise was abolished by Ni2+. Hypercontracture developed more frequently in HINCX and CHF than in SHAM cells. The amount of NCX was 54% higher in HINCX and 76% higher in CHF compared with SHAM. Na+-loaded cardiomyocytes from CHF and HINCX rats are more susceptible to Ca2+ overload than SHAM cells because of the increased capacity for Na,Ca-exchange.

The effect of six polymorphisms in the Apolipoprotein B gene on parameters of lipid metabolism in a Danish population.

Lipoproteins are vehicles for the distribution of plasma lipids and polymorphisms in the genes for apolipoproteins could influence the amount of lipid in plasma. We examined the effect of six single nucleotide polymorphisms in codons 71, 591, 2488, 2712, 3611, and 4154 of the apolipoprotein B gene on fasting levels of triglyceride, VLDL-, LDL-, HDL- and total cholesterol and on body mass index (BMI) in a cohort of 2656 Danes aged 40-70 years using a linear model correcting for the effects of gender, age, BMI, smoking, alcohol consumption and physical activity. The codon 2488 polymorphism was the most influential of the tested polymorphisms, significantly influencing triglyceride (P = 0.002), LDL-cholesterol (P < or = 0.0004), VLDL-cholesterol (P = 0.006) and total cholesterol (P = 0.0001). The codon 2712 polymorphism had an impact on triglyceride (P = 0.007) and VLDL-cholesterol (P = 0.001), while the codon 71 polymorphism influenced LDL- and total cholesterol (P = 0.04 and P = 0.02, respectively). An interaction between smoking and codon 591 (P = 0.03) and smoking and codon 3611 (P = 0.02) on BMI was observed, as well as modest interactions between codon 3611 and codons 2488 and 2712 on lipid parameters. All polymorphisms were in close linkage disequilibrium. The population was not in Hardy-Weinberg equilibrium in four of the six polymorphisms but the lack of equilibrium was restricted mainly to the 60-year olds.

The influence of the polymorphism in apolipoprotein B codon 2488 on insulin and lipid levels in a Danish twin population.

AIMS: The apolipoprotein B codon 2488 polymorphism has been associated with the metabolism of lipoproteins in subjects with Type 2 diabetes. However, no data are available on the influence of the polymorphism on insulin or glucose metabolism. This study examines the impact of the polymorphism on parameters associated with the insulin resistance syndrome in Danish twins. METHODS: The effect of the polymorphism on lipid, glucose and insulin measures was studied in 548 same sex twins aged 55-74 years. RESULTS: The codon 2488 polymorphism influenced fasting triglyceride levels, as well as insulin, as measured at 120 min in an oral glucose tolerance test. Subjects with the genotype T2488T had 14% higher triglyceride levels (P = 0.02) and 31% higher insulin levels (P = 0.004) than subjects with genotype C2488C. In twins discordant for genotype, the T-allele was associated with higher levels of triglyceride (P = 0.04) and insulin (P = 0.02) and lower levels of HDL-cholesterol (P = 0.04). CONCLUSION: The T-allele of the codon 2488 polymorphism influenced parameters related to the insulin resistance syndrome, i.e. increased levels of insulin, increased levels of triglyceride and decreased levels of HDL. As the polymorphism is silent, these effects must be mediated through linkage to other polymorphisms in apolipoprotein B or other genes on chromosome 2.

The prognostic importance of volume-weighted mean nuclear volume, mitotic index, and other stereologically measured quantitative parameters in supraglottic laryngeal carcinoma.

BACKGROUND: Stereologically measured mean nuclear volume has been proven to have prognostic importance in several types of cancer, such as malignant melanoma and carcinomas of the breast, oral region, bladder, and uterine cervix. The main purpose of the current study was to investigate the possible prognostic importance of mean nuclear volume and mitotic index in carcinoma of the supraglottic larynx. METHODS: The study was performed with a stratified, random sample of 113 patients from a well-defined group of 386 patients with supraglottic laryngeal carcinoma treated with radiotherapy at the Finsen Institute in Copenhagen. Histologic sections from pretreatment biopsies were used to estimate the following parameters: mean nuclear volume (Vv(3)(0)), mitotic index (MI), number of nuclei per mm(2) (QA), mean nuclear profile area (Anuc), and the area fraction of nuclei in cancer tissue (AA). The geometric means of the parameters were used as cutoff points in a single factor and in a multivariate survival analysis with relapse free survival as the primary endpoint. RESULTS: The geometric means of the measured parameters were (Vv(3)(0)) = 480 micro(3), QA = 3630 nuclei/micro(2) cancer tissue, MI = 0.48 mitosis/100 nuclei, AA = 0.21, and Anuc = 57.9 micro(2). CONCLUSIONS: None of the stereologically estimated parameters proved to have prognostic importance, whereas tumor size, and lymph node status did. The method of adaptive, stratified, random sampling used in this study can save a great deal of work and is highly recommended by the authors.