Oocyte diameter predicts the maturation rate of human immature oocytes collected ex vivo.

PURPOSE: To study the impact of oocyte diameter and cumulus cell mass on the potential for final maturation of immature human oocytes in vitro. METHODS: Immature oocytes (n = 1563) from 75 women undergoing fertility preservation by ovarian tissue cryopreservation (14-41 years) were collected. After preparation of the ovarian cortex for freezing, immature oocytes were collected from the surplus medulla. After collection, IVM was performed according to standard published methods. The mass of cumulus cell surrounding the immature oocyte was grouped according to size. After IVM, each oocyte was photographed, measured, and the diameter was calculated as a mean of two perpendicular measurements. RESULTS: The diameter of the oocytes ranged from 60 to 171 µm with a mean of 115 µm (SD:12.1) and an interquartile range from 107 to 124 µm. The oocyte diameter was positively associated with a higher incidence of MII (p < 0.001). MII oocytes had a significantly larger mean diameter than MI, GV, and degenerated oocytes. The size of the cumulus cell mass was significantly associated with the MII stage (p < 0.001) and larger oocyte diameter (p < 0.001). The results further confirm that the diameter of the fully grown oocyte is reached relatively early in human follicular development and that the factors governing oocyte maturation in vitro are connected to the surrounding cell mass and the oocyte. CONCLUSION: The diameter of the oocyte is a highly determining factor in the nuclear maturation of the human oocyte during in vitro maturation, and the size of the cumulus cell mass is closely positively associated with a larger diameter.

Parenthood among men diagnosed with cancer in childhood and early adulthood: trends over time in a Danish national cohort.

STUDY QUESTION: Is the rate of fatherhood among men diagnosed with cancer in childhood and early adulthood different from men without cancer, and, if so, have the differences changed over time? SUMMARY ANSWER: Men diagnosed with cancer have had significantly reduced rates of fatherhood compared with undiagnosed men; however, the rates of fatherhood among the cancer survivors have increased markedly over time. WHAT IS KNOWN ALREADY: The number of children and young adolescents who survive cancer has steadily increased over recent decades, with a current 5-year survival rate of approximately 80%. Consequently, life circumstances after cancer have gained increasing importance, including the desire among survivors to have children and a family. ARTs to aid reproduction among cancer survivors have been developed, and fertility preservation is increasingly a topic being discussed before undergoing cancer treatment. But the potential for fertility preservation differs dependent on age at diagnosis and type of cancer. Earlier studies have shown a decreased fertility rate among survivors of child and adolescent cancer compared to those diagnosed in early adulthood. STUDY DESIGN, SIZE, DURATION: This study is a national, register-based cohort study. Men diagnosed with cancer in childhood and early adulthood (<30 years of age) were registered in the Danish Cancer Register in 1978-2016 (n = 9353). According to the time of diagnosis, each cancer-diagnosed man was randomly matched with 150 undiagnosed men from the background population within the same birth year. The men were followed until having their first child, death, migration or the end of the study (31 December 2017) in medical registers and socio-demographic population registers. PARTICIPANTS/MATERIALS, SETTING, METHODS: Fatherhood among the boys and young men diagnosed with cancer were compared with the age-matched comparison group in all statistical analyses. Cancer diagnoses were categorised as central nervous system (CNS) cancers, haematological cancers or solid cancers. Analyses were stratified by age at diagnosis (0-9, 10-19, 20-29 years) and time of diagnosis (1978-1989, 1990-1999, 2000-2009, 2010-2016). Death was incorporated as a competing risk in all analyses. MAIN RESULTS AND THE ROLE OF CHANCE: The study population consisted of 9353 boys and young men diagnosed with cancer between 1978 and 2016 and 1 386 493 men in the age-matched comparison group. Those surviving CNS cancer as young men had the lowest hazard ratio (HR) of fatherhood compared with the age-matched comparison group (HR 0.67, 95% CI 0.57-0.79), followed by survivors of haematological cancers (HR 0.90, 95% CI 0.81-1.01), while the highest chance of fatherhood was among survivors of solid cancers (HR 1.16, 95% CI 1.12-1.20) with a slightly increased HR compared with undiagnosed males. The HR of becoming a father increased over time. From the first decade to the last decade 30 years later, the HR of becoming a father increased for solid tumours (HR 0.78, 95% CI 0.73-0.83 to HR 1.08, 95% CI 0.95-1.22), haematological cancers (HR 0.64, 95% CI 0.53-0.79 to HR 0.97, 95% CI 0.73-1.30) and CNS cancers (HR 0.44, 95% CI 0.34-0.57 to HR 0.98, 95% CI 0.49-1.95) compared to the age-matched comparison group. Also, when compared with the age-matched comparison group, men diagnosed with cancer when aged 20-29 years were more likely became fathers over the time of the study (HR 0.80, 95% CI 0.74-0.86 to HR 1.08, 95% CI 0.96-1.22). LIMITATIONS, REASONS FOR CAUTION: The study was based on register data, and information was not available about the men's fertility potential, whether they had a desire to have children and whether it was possible for them to find a partner. Information about fertility preservation, e.g. sperm freezing, could also have provided additional insights. Furthermore, information about diagnosis and ART treatment would have been beneficial. WIDER IMPLICATIONS OF THE FINDINGS: Information and education of male patients diagnosed with cancer about fertility preservation options and their chances to create their own family is crucial. Reassuringly, time trends showed more men with a previous cancer diagnosis becoming fathers in recent years than in earlier years, reflecting that survival and fertility preservation have improved over time. STUDY FUNDING/COMPETING INTEREST(S): R.S. received a PhD grant from the Rosa Ebba Hansen Foundation and from the Health Foundation (J.nr. 15-B-0095). The funding for the establishment of the DANAC II Cohort was obtained from the Rosa Ebba Hansen Foundation. The authors have no conflicts of interest to declare. TRIAL REGISTRATION NUMBER: N/A.

A simple method to quantify follicle survival in cryopreserved human ovarian tissue.

STUDY QUESTION: Can follicle survival in frozen-thawed human ovarian tissue be quantified in situ using the dye Neutral Red (NR) to stain viable follicles specifically? SUMMARY ANSWER: A follicle survival rate within ovarian tissue can be calculated using NR followed by histological evaluation and evidence for a consistently high follicle survival in a series of ovarian tissue from 25 Danish girls and women undergoing ovarian tissue cryopreservation (OTC) was obtained. WHAT IS KNOWN ALREADY: Securing follicle survival in cryopreserved ovarian tissue is crucial for proper quality control when centers wish to implement OTC. The only established technique for validation of follicle survival is xenografting of thawed ovarian tissue to immunodeficient mice. However, this functional test is expensive, time consuming, requires animal facilities and only provides a qualitative-not quantitative-measure for follicle survival. STUDY DESIGN SIZE, DURATION: Quantification of follicle survival in human ovarian tissue donated from 30 girls and women having tissue cryopreserved for fertility preservation from 2000 to 2015 at the Laboratory of Reproductive Biology in Copenhagen, Denmark. PARTICIPANTS/MATERIALS, SETTING, METHODS: Cryopreserved ovarian cortex was donated from 25 girls and young women aged 10-36 years (mean age: 25 years) and the average storage time in liquid nitrogen was 9.1 ± 5.6 years, ranging from 1.6 to 17.9 years. In 12 of the cases, the ovarian tissue was collected from the local hospital and in the other 13 cases the ovarian tissue was transported on ice up to 6 h prior to freezing. Donated fresh ovarian surplus tissue was obtained from five women aged 23-34 years (mean age: 27 years). Ovarian tissues were chopped into small fragments and incubated in culture medium containing 50 mg/ml NR for 3-4 h. Fragments of ovarian tissue containing clearly NR-stained follicles were selected for counting, encapsulated in 4% agar and were processed for histology to calculate a follicular survival rate. MAIN RESULTS AND THE ROLE OF CHANCE: The mean follicle survival rate in the 25 patients after freezing and thawing was 84% ± 11 (mean ±SD), ranging from 50% to 98%. The high follicle survival rate in this clinical series of patients reflects a constant high-quality service performed in our center and confirms the robustness of the slow freezing protocol. No significant association between follicle survival rates and storage time was found using linear regression analysis, suggesting that storage in liquid nitrogen does not affect viability of the tissue. No significant association in follicle survival rates was found between ovarian tissues collected at the local hospital compared to tissues transported on ice prior to freezing, supporting that prolonged cooling does not seem to greatly affect the follicle survival. For the fresh ovarian tissue, the average follicle survival rate was 91% ± 5 (mean ± SD) in five patients, ranging from 81% to 95%. LIMITATIONS, REASONS FOR CAUTION: Even though the NR staining requires active incorporation of the dye, the test is merely a short in situ test that cannot completely replace the functional value of xenografting studies in which the integrity and developmental potential of the ovarian follicles are assessed. WIDER IMPLICATIONS OF THE FINDINGS: OTC is now being employed around the world but to date it has been difficult for centers to evaluate the effectiveness of their program and perform proper quality control. NR staining combined with histological evaluation is the first quantitative method to provide a survival rate for follicles in frozen-thawed human ovarian tissue and offer a valuable and easily applicable tool to validate the cryopreservation procedure when implementing OTC or as routine quality control for the overall freezing performance within tissue banking facilities. STUDY FUNDING/COMPETING INTEREST(S): The Research Pools of Rigshospitalet, the Danish Cancer Foundation, Dagmar Marshalls Foundation, and the Novo Nordic Foundation are thanked for having funded this study. The authors have no conflicts of interest.

The polymorphic insertion of the luteinizing hormone receptor "insLQ" show a negative association to LHR gene expression and to the follicular fluid hormonal profile in human small antral follicles.

The luteinizing hormone receptor (LHCGR) has a little studied polymorphic 6 bp insertion (rs4539842/insLQ). This study has evaluated the insLQ polymorphism in relation to potential associations with hormonal characteristics of human small antral follicles (hSAFs). In total, 310 hSAFs were collected from 86 women undergoing fertility preservation. Analysis included hormonal profile of 297 follicular fluid (FF) samples and 148 corresponding granulosa cells samples were evaluated by qPCR for selected genes. Significantly reduced and non-detectable mRNA levels of anti-Müllerian hormone receptor II (AMHR2) and LHCGR, respectively, were observed for insLQ/insLQ compared to -/insLQ and the -/- genotypes. Moreover, LHCGR and CYP19a1 together with oestradiol and inhibin-B were significantly increased in -/insLQ compared to the -/- genotype. The homozygous insLQ genotype showed strong significant associations to GC specific genes LHCGR and CYP19a1, which may translate into significant changes in FF hormone profiles and an altered LH signaling.

Fertility preservation issues in pediatric hematopoietic stem cell transplantation: practical approaches from the consensus of the Pediatric Diseases Working Party of the EBMT and the International BFM Study Group.

Fertility preservation is an urgent challenge in the transplant setting. A panel of transplanters and fertility specialists within the Pediatric Diseases Working Party of the European Society for Blood and Marrow Transplantation (EBMT) and the International BFM Study Group provides specific guidelines. Patients and families should be informed of possible gender- and age-specific cryopreservation strategies that should be tailored according to the underlying disease, clinical condition and previous exposure to chemotherapy. Semen collection should be routinely offered to all postpubertal boys at the diagnosis of any disease requiring therapy that could potentially impair fertility. Testicular tissue collection might be offered to postpubertal boys; nevertheless, its use has been unsuccessful to date. Oocyte collection after hormonal hyperstimulation should be offered to postpubertal girls facing gonadotoxic therapies that could be delayed for the 2 weeks required for the procedure. Ovarian tissue collection could be offered to pre-/post-pubertal girls. Pregnancies have been reported after postpubertal ovarian tissue reimplantation; however, to date, no pregnancy has been reported after the reimplantation of prepubertal ovarian tissue or in vitro maturation of pre-/post-pubertal ovarian tissue. Possible future advances in reproductive medicine could change this scenario. Health authorities should prioritize fertility preservation projects in pediatric transplantation to improve patient care and quality of life.

State-of-the-art fertility preservation in children and adolescents undergoing haematopoietic stem cell transplantation: a report on the expert meeting of the Paediatric Diseases Working Party (PDWP) of the European Society for Blood and Marrow Transplantation (EBMT) in Baden, Austria, 29-30 September 2015.

Nowadays, allogeneic haematopoietic stem cell transplantation (allo-HSCT) is a well-established treatment procedure and often the only cure for many patients with malignant and non-malignant diseases. Decrease in short-term complications has substantially contributed to increased survival. Therefore long-term sequelae are reaching the focus of patient care. One of the most important risks of stem cell transplant survivors is infertility. As well as in the field of allo-HSCT also the field of reproductive medicine has achieved substantial advances to offer potential options for fertility preservation in both boys and girls. Access to these procedures as well as their financing differs significantly throughout Europe. As all European children and adolescents should have the same possibility, the Paediatric Diseases Working Party of the European Society for Blood and Marrow Transplantation organised an expert meeting in September 2015. This manuscript describes the recommendations for the diagnosis and pre-emptive procedures that should be offered to all children and adolescents in Europe who have to undergo an allo-HSCT.

Cryopreservation of ovarian tissue for fertility preservation in a large cohort of young girls: focus on pubertal development.

STUDY QUESTION: Is there an association between the need for medical puberty induction and the diagnosis or treatment received in girls who have undergone cryopreservation of ovarian tissue for fertility preservation? SUMMARY ANSWER: There was a clear association between the intensity of treatment received and requirement for medical puberty induction but no association with the diagnosis. WHAT IS KNOWN ALREADY: Although it cannot be predicted which girls will become infertile or develop premature ovarian insufficiency (POI) following intensive chemotherapy or irradiation, patients who are at high risk of POI should be offered ovarian tissue cryopreservation (OTC). This includes girls who are planned to receive either high doses of alkylating agents, conditioning regimen before stem cell transplantation (SCT), total body irradiation (TBI) or high radiation doses to the craniospinal, abdominal or pelvic area. STUDY DESIGN, SIZE, DURATION: This is a retrospective cohort study. In total, 176 Danish girls under 18 years of age have had OTC performed over a period of 15 years. An overview of the girls' diagnoses and mean age at OTC as well as the number of deceased is presented. Of the 176 girls, 38 had died and 46 girls were still younger than 12 years so their pubertal development cannot be evaluated yet. For the 60 girls who had OTC performed after 12 years of age, the incidence of POI was evaluated and in the group of 32 girls who were younger than 12 years at OTC, the association between the diagnosis and received treatment and the requirement for medical puberty induction was examined. PARTICIPANTS/MATERIALS, SETTING, METHODS: The need for medical puberty induction was assessed in 32 girls who were prepubertal at the time of OTC. MAIN RESULTS AND THE ROLE OF CHANCE: Indications for OTC were allogeneic SCT for leukaemia, myelodysplastic syndrome or benign haematological disorders, autologous SCT for lymphoma or sarcoma, and irradiation to the pelvis or to the spinal axis. The mean age at OTC of the 176 girls were 11.3 years. The two most prevalent diagnoses of the 176 girls were malignant tumours and malignant haematological diseases. Among the 32 prepubertal girls, 12 received high dose chemotherapy and either TBI prior to SCT or irradiation to the pelvis, abdomen or the spinal axis, 13 received high dose alkylating agents but no irradiation prior to SCT, six received alkylating agents as part of conventional chemotherapy and one patient had a genetic metabolic disorder and did not receive gonadotoxic treatment. Among these 32 girls, 23 did not undergo puberty spontaneously and thus received medical puberty induction. Among the nine girls, who went through spontaneous puberty, four had received high dose alkylating agents and five had received conventional chemotherapy. LIMITATIONS REASONS FOR CAUTION: All information was retrieved retrospectively from patient records, and thus some information was not available. WIDER IMPLICATIONS OF THE FINDINGS: OTC should be recommended to all young girls, who present a high risk of developing ovarian insufficiency and/or infertility following high dose chemotherapy and/or irradiation. STUDY FUNDING/COMPETING INTERESTS: The Childhood Cancer Foundation (2012-2016) and the EU interregional project ReproHigh are thanked for having funded this study. They had no role in the study design, collection and analysis of the data or writing of the report. The authors have no conflict of interest to disclose.

Outcomes of transplantations of cryopreserved ovarian tissue to 41 women in Denmark.

STUDY QUESTION: What are the results of transplanting cryopreserved ovarian tissue? SUMMARY ANSWER: The transplanted ovarian tissue can last up to 10 years, with no relapses following the 53 transplantations, and the chance of a successful pregnancy is currently around one in three for those with a pregnancy-wish. WHAT IS KNOWN ALREADY: Cryopreservation of ovarian tissue is now gaining ground as a valid method for fertility preservation. More than 36 children worldwide have now been born following this procedure. STUDY DESIGN, SIZE, DURATION: This is a retrospective cohort study of 41 women who had thawed ovarian tissue transplanted 53 times over a period of 10 years, including 1 patient who was lost to follow-up. PARTICIPANTS/MATERIALS, SETTING, METHODS: The 41 Danish women, who had in total 53 transplantations, were followed for ovarian function and fertility outcome. Safety was assessed by monitoring relapse in cancer survivors. MAIN RESULTS, AND THE ROLE OF CHANCE: Among 32 women with a pregnancy-wish, 10 (31%) had a child/children (14 children in total); this included 1 woman with a third trimester on-going pregnancy. In addition, two legal abortions and one second trimester miscarriage occurred. A total of 24 clinical pregnancies were established in the 32 women with a pregnancy-wish. The tissue remained functional for close to 10 years in some cases and lasted only a short period in others. Three relapses occurred but were unlikely to be due to the transplanted tissue. LIMITATIONS, REASONS FOR CAUTION: Self-report through questionnaires with only in-one hospital formalised follow-up of transplanted patients could result in unreported miscarriages. The longevity of the tissue may vary by few months compared with those reported because some patients simply could not remember the date when the tissue became non-functional. WIDER IMPLICATIONS OF THE FINDINGS: Cryopreservation of ovarian tissue is likely to become integrated into the treatment of young women, with cancer, who run a risk of losing their fertility. The full functional lifespan of grafts is still being evaluated, because many of the transplanted women have continued to maintain ovarian activity. Some of our first cases have had tissue functioning for ∼ 10 years.

Family intentions and personal considerations on postponing childbearing in childless cohabiting and single women aged 35-43 seeking fertility assessment and counselling.

STUDY QUESTION: What characterizes childless women aged 35 years and above seeking fertility assessment and counselling in relation to their reproduction and are there significant differences between single and cohabiting women? SUMMARY ANSWER: Despite the women's advanced age and knowledge of the age-related decline in fecundity, 70% of the single women sought fertility assessment and counselling to gain knowledge regarding the possibility of postponing pregnancy. WHAT IS KNOWN ALREADY: Recent studies have indicated an increasing demand for ovarian reserve testing in women without any known fertility problem to obtain knowledge on their reproductive lifespan and pro-fertility advice. Women postpone their first pregnancy, and maternal age at first birth has increased in western societies over the past two to four decades. Postponed childbearing implies a higher rate of involuntary childlessness, smaller families than desired and declining fertility rates. STUDY DESIGN, SIZE, AND DURATION: Baseline data from a cross-sectional cohort study of 340 women aged 35-43 years examined at the Fertility Assessment and Counselling (FAC) Clinic at Copenhagen University Hospital from 2011 to 2014. The FAC Clinic was initiated to provide individual fertility assessment and counselling. PARTICIPANTS/MATERIALS, SETTING, METHODS: Eligible women were childless and at least 35 years of age. All completed a web-based questionnaire before and after the consultation including socio-demographic, reproductive, medical, lifestyle and behavioural factors. Consultation by a fertility specialist included transvaginal ultrasound, full reproductive history and AMH measurement. MAIN RESULTS AND THE ROLE OF CHANCE: The study comprised 140 cohabiting and 200 single women. The majority (82%) were well-educated and in employment. Their mean age was 37.4 years. Nonetheless, the main reasons for attending were to obtain knowledge regarding the possibility of postponing pregnancy (63%) and a concern about their fecundity (52%). The majority in both groups (60%) wished for two or more children. The women listed their ideal age at birth of first child and last child as 33 (±4.7) years and 39 (±3.5) years, respectively. Of the single women, 70% would accept use of sperm donation compared with 25% of the cohabiting women (P < 0.001). In general, 45% considered oocyte vitrification for social reasons, yet only 15% were positive towards oocyte donation. The two groups were comparable regarding lifestyle factors, number of previous sexual partners, pregnancies, and ovarian reserve parameters. LIMITATIONS, REASONS FOR CAUTION: The women in the present study were conscious of the risk of infertility with increasing age and attended the FAC Clinic due to a concern about their remaining reproductive lifespan, which in combination with their high educational level could impair the generalizability to the background population. WIDER IMPLICATIONS OF THE FINDINGS: The results indicate that in general women overestimate their own reproductive capacity and underestimate the risk of future childlessness with the continuous postponement of pregnancies.

Ovarian reserve assessment in users of oral contraception seeking fertility advice on their reproductive lifespan.

STUDY QUESTION: To what extent does oral contraception (OC) impair ovarian reserve parameters in women who seek fertility assessment and counselling to get advice on whether their remaining reproductive lifespan is reduced? SUMMARY ANSWER: Ovarian reserve parameters defined by anti-Müllerian hormone (AMH), antral follicle count (AFC) and ovarian volume were found to be significantly decreased by 19% (95% CI 9.1-29.3%), 18% (95% CI 11.2-24.8%) and 50% (95% CI 45.1-53.7%) among OC users compared with non-users. WHAT IS KNOWN ALREADY: AMH and AFC have proved to be reliable predictors of ovarian ageing. In women, AMH declines with age and data suggest a relationship with remaining reproductive lifespan and age at menopause. OC may alter parameters related to ovarian reserve assessment but the extent of the reduction is uncertain. STUDY DESIGN, SIZE, DURATION: A cross-sectional study of 887 women aged 19-46 attending the Fertility Assessment and Counselling Clinic (FACC) from 2011 to 2014 comparing ovarian reserve parameters in OC users with non-OC users. PARTICIPANTS/MATERIALS, SETTING, METHODS: The FAC Clinic was initiated to provide individual fertility assessment and counselling. All women were examined on a random cycle day by a fertility specialist. Consultation included; transvaginal ultrasound (AFC, ovarian volume, pathology), a full reproductive history and AMH measurement. Women were grouped into non-users and users of OC (all combinations of estrogen-progestin products and the contraceptive vaginal ring). Non-users included women with an intrauterine device (IUD) or no hormonal contraception. MAIN RESULTS AND THE ROLE OF CHANCE: Of the 887 women, 244 (27.5%) used OC. In a linear regression analyses adjusted for age, ovarian volume was 50% lower (95% CI 45.1-53.7%), AMH was 19% lower (95% CI 9.1-29.3%), and AFC was 18% lower (95% CI 11.2-24.8%) in OC users compared with non-users. Comparison of AMH at values of <10 pmol/l OC was found to have a significant negative influence on AMH (OR 1.6, 95% CI 1.1; 2.4, P = 0.03). Furthermore, we found a significant decrease in antral follicles sized 5-7 mm (P < 0.001) and antral follicles sized 8-10 mm (P < 0.001) but an increase in antral follicles sized 2-4 mm (P = 0.008) among OC users. The two groups (OC users versus non-users) were comparable regarding age, BMI, smoking and maternal age at menopause. LIMITATIONS, REASON FOR CAUTION: The study population comprised women attending the FAC Clinic. Recruitment was based on self-referral, which could imply a potential selection bias. Ovarian reserve was examined at a random cycle day. However, both AMH and AFC can be assessed independently of the menstrual cycle. The accuracy in predicting residual reproductive lifespan is still needed in both users and non-users of OC. WIDER IMPLICATIONS OF THE FINDINGS: OC has a major impact on the ovarian volume, and a moderate impact on AFC and AMH with a shift towards the smaller sized antral follicle subclasses. The most evident reduction occurs in the antral follicles of 5-7 and 8-10 mm with the highest number of AMH secreting granulosa cells. It is essential to be aware of the impact of OC use on ovarian reserve parameters when guiding OC users on their fertility status and reproductive lifespan. STUDY FUNDING/COMPETING INTERESTS: The FAC Clinic was established in 2011 as part of the ReproHigh collaboration. This study received funding through the Capital Region Research Fund and by EU-regional funding. There are no competing interests. TRIAL REGISTRATION NUMBER: The biobank connected to FAC Clinic is approved by the Scientific Ethical Committee (H-1-2011-081).

Distribution and function of 3',5'-Cyclic-AMP phosphodiesterases in the human ovary.

The concentration of the important second messenger cAMP is regulated by phosphodiesterases (PDEs) and hence an attractive drug target. However, limited human data are available about the PDEs in the ovary. The aim of the present study was to describe and characterise the PDEs in the human ovary. Results were obtained by analysis of mRNA microarray data from follicles and granulosa cells (GCs), combined RT-PCR and enzymatic activity analysis in GCs, immunohistochemical analysis of ovarian sections and by studying the effect of PDE inhibitors on progesterone production from cultured GCs. We found that PDE3, PDE4, PDE7 and PDE8 are the major families present while PDE11A was not detected. PDE8B was differentially expressed during folliculogenesis. In cultured GCs, inhibition of PDE7 and PDE8 increased basal progesterone secretion while PDE4 inhibition increased forskolin-stimulated progesterone secretion. In conclusion, we identified PDE3, PDE4, PDE7 and PDE8 as the major PDEs in the human ovary.