Barbara Sonntag

Effects of the FSH receptor gene polymorphism p.N680S on cAMP and steroid production in cultured primary human granulosa cells

The study was designed to evaluate in vitro the cellular mechanisms of the single nucleotide polymorphism (SNP) p.N680S of the FSH receptor gene (FSHR) in human granulosa cells (GC) and included patients homozygous for the FSHR SNP (NN/SS) undergoing ovarian stimulation. GC were isolated during oocyte retrieval and cultured for 1–7 days. Basal oestradiol and progesterone concentrations were measured after short-term culture. The kinetics of cAMP, oestradiol and progesterone concentrations in response to various amounts of FSH were analysed in a 6–7 day culture. Basal oestradiol, but not progesterone, concentrations on day 1 of GC culture, were significantly higher in NN compared with SS (P = 0.045), but non-responsive to FSH stimulation. Immunofluorescence microscopy demonstrated the re-appearance of FSHR expression with increasing days in culture. Upon stimulation with FSH, GC cultured for 6–7 days displayed a dose-dependent increase of cAMP, oestradiol and progesterone but no difference in the EC50 values between both variants. Primary long-term GC cultures are a suitable system to study the effects of FSH in vitro. However, the experiments suggest that factors down-stream of progesterone production or external to GC might be involved in the clinically observed differences in an FSHR variant-mediated response to FSH.

Preterm birth but not mode of delivery is associated with an increased risk of developing inflammatory bowel disease later in life

Background: Exposure to bacterial antigens and other environmental factors in combination with a genetic susceptibility have been implicated in the etiology of inflammatory bowel disease (IBD). As certain perinatal circumstances, e.g., delivery by cesarean section, predispose to a different intestinal colonizations the aim of this analysis was to define a potential influence on the development of IBD in later life. Methods: In a case‐control study design, birth data were recorded from patients diagnosed with IBD (Crohns disease [CD], n = 1096; ulcerative colitis [UC], n = 763) and healthy controls ([C], n = 878) by a self‐administered questionnaire. Results: Preterm birth (CD: odds ratio [OR] 1.5 [95% confidence interval 1.1–2.0], UC: OR 1.3 [0.9–1.9]), mothers disease during pregnancy (CD: OR 1.9 [1.3–2.9], UC: OR 1.6 [1.0–2.4]), and disease in the first year of life (CD: OR 2.2 [1.6–2.9], UC: OR 1.7 [1.3–2.3]) are associated with the development of IBD in later life. No significant associations were found for the mode of delivery and breast feeding. In a logistic regression analysis female sex, smoking, appendectomy, maternal IBD, and disease in the first year of life were independently associated with CD. Female sex, appendectomy, and disease in the first year of life were independently associated with UC. Conclusions: Preterm birth and other perinatal circumstances are associated with the development of IBD, of which disease in the first year of life is an independent risk factor in multivariate analysis.

Androgens and Insulin – Two Key Players in Polycystic Ovary Syndrome

Androgens and insulin are endocrine key players in the pathophysiology of polycystic ovary syndrome (PCOS), a heterogenic condition of unexplained etiology and a suspected genetic background. Androgens mediate the clinical phenotype of the disease. Therefore,all criteria of the recent PCOS consensus definition are based on their biological effects. Insulin resistance, followed by compensatory hyperinsulinemia, is frequently found in patients with PCOS. Insulin resistance is correlated with a risk of metabolic complications of PCOS, and recent research has focused on possible long-term health consequences of the syndrome. Newest molecular genetic findings at the receptor level of both androgens and insulin support their pivotal role in PCOS. These results could help to better characterize the heterogenic disorder, enabling a refinement of existing individualized therapeutic strategies.

Lipoprotein (a) and other prothrombotic risk factors in Caucasian women with unexplained recurrent miscarriage Results of a multicentre case-control study

From 1998 to 2003, 133 Caucasian women aged 17-40 years (median 29 years) suffering from unexplained recurrent miscarriage (uRM) were consecutively enrolled. In patients and 133 age-matched healthy controls prothrombotic risk factors (factor V (FV) G1691A, factor II (FII) G20210A, MTHFR T677T, 4G/5G plasminogen activator inhibitor (PAI)-1, lipoprotein (Lp) (a), protein C (PC), protein S (PS), antithrombin (AT), antiphospholipid/anticardiolipin (APA/ACA) antibodies) as well as associated environmental conditions (smoking and obesity) were investigated. 70 (52.6%) of the patients had at least one prothrombotic risk factor compared with 26 control women (19.5%; p<0.0001). Body mass index (BMI; p=0.78) and smoking habits (p=0.44) did not differ significantly between the groups investigated. Upon univariate analysis the heterozygous FV mutation, Lp(a) > 30 mg/dL, increased APA/ACA and BMI > 25 kg/m(2) in combination with a prothrombotic risk factor were found to be significantly associated with uRM. In multivariate analysis, increased Lp(a) (odds ratio (OR): 4.7/95% confidence interval (CI): 2.0-10.7), the FV mutation (OR:3.8/CI:1.4-10.7), and increased APA/ACA (OR: 4.5/CI: 1.1-17.7) had independent associations with uRM.

Advanced follicle development in xenografted prepubertal ovarian tissue: the common marmoset as a nonhuman primate model for ovarian tissue transplantation

OBJECTIVE To establish a nonhuman primate model addressing follicular development in cryopreserved prepubertal ovarian tissue after xenografting. DESIGN Experimental study. SETTING Academic research center. ANIMAL(S) Ovarian tissue from female prepubertal common marmoset (Callytrix jacchus jacchus) grafted into immunodeficient nude mice (Crl:NU-FoxnI(nu)). INTERVENTION(S) Removal and subsequent cryopreservation of ovarian tissues with dimethyl sulfoxide, followed by grafting to subcutaneous sites of ovariectomized and intact nude mice. MAIN OUTCOME MEASURE(S) Histologic evaluation for the mean number of total and morphologically normal follicles in each class. RESULT(S) The mean number of unadvanced follicles in frozen-thawed grafted ovarian tissues was reduced compared with pregraft controls, but the prevalence of normal follicular morphology was either slightly increased (primordial follicles) or unchanged (primary follicles). Previous ovariectomy in graft recipients increased total follicle numbers without effect on normal follicular morphology and shifted the ratio of primordial to primary follicles toward an increase in primary follicles, indicating activation of follicular maturation. CONCLUSION(S) The marmoset is a suitable primate model for studies on the subsequent use of cryopreserved ovarian tissue, demonstrating graft sustainment and the development of follicles from prepubertal ovarian tissue in immunodeficient hosts up to secondary and preantral stages.

FSH prevents depletion of the resting follicle pool by promoting follicular number and morphology in fresh and cryopreserved primate ovarian tissues following xenografting

BackgroundCryopreservation and transplantation of ovarian tissue is one option for re-establishing ovarian function, but optimal conditions for graft sustainment and follicular survival are still considered experimental. The present study aims to analyze the effect of FSH treatment on the resting follicle pool in fresh and cryopreserved primate ovarian tissues following xenografting.MethodsOvarian tissues from adult marmosets were grafted freshly or following cryopreservation to ovarectomized nude mice treated with FSH 25 IU twice daily post transplantation or left untreated as controls. Grafts were retrieved 2 or 4 weeks after transplantation to evaluate the number and morphological appearance of follicles.ResultsEarly start of FSH treatment within 1 week following transplantation partly prevents primordial follicle loss in fresh and frozen-thawed tissues, whereas after a 3 weeks time interval this effect is present only in fresh tissues. A similar positive effect of early, but not later FSH treatment on primary follicles is seen in fresh tissues compared to only marginal effects in frozen-thawed tissues. The percentage of morphologically normal follicles is generally increased in FSH treated tissues, whereas the percentage of primary follicles over all primordial and primary follicles is increased by FSH only in freshly-grafted tissues.ConclusionsFSH treatment alleviates depletion of the resting follicle pool and promotes normal follicular morphology both in freshly and frozen-thawed grafted tissues. In previously cryopreserved tissues, applying to most of the tissues intended for clinical use in fertility preservation attempts, its positive effect on primordial follicle numbers and potential graft sustainment is dependent on an early start of treatment within one week of transplantation.

Assessment of follicular development in cryopreserved primate ovarian tissue by xenografting: prepubertal tissues are less sensitive to the choice of cryoprotectant

Improvements in cancer survival rates have renewed interest in the cryopreservation of ovarian tissue for fertility preservation. We used the marmoset as a non-human primate model to assess the effect of different cryoprotectives on follicular viability of prepubertal compared to adult ovarian tissue following xenografting. Cryopreservation was performed with dimethylsulfoxide (DMSO), 1,2-propanediol (PrOH), or ethylene glycol (EG) using a slow freezing protocol. Subsequently, nude mice received eight grafts per animal from the DMSO and the PrOH groups for a 4-week grafting period. Fresh, cryopreserved-thawed, and xenografted tissues were serially sectioned and evaluated for the number and morphology of follicles. In adult tissue, the percentage of morphologically normal primordial follicles significantly decreased from 41.2 ± 4.5% (fresh) to 13.6 ± 1.8 (DMSO), 9.5 ± 1.7 (PrOH), or 6.8 ± 1.0 (EG) following cryopreservation. After xenografting, the percentage of morphologically normal primordial (26.2 ± 2.5%) and primary follicles (28.1 ± 5.4%) in the DMSO group was significantly higher than that in the PrOH group (12.2 ± 3 and 5.4 ± 2.1% respectively). Proliferating cell nuclear antigen (PCNA) staining suggests the resumption of proliferative activity in all cellular compartments. In prepubertal tissues, primordial but not primary follicles display a similar sensitivity to cryopreservation, and no significant differences between DMSO and PrOH following xenografting were observed. In conclusion, DMSO shows a superior protective effect on follicular morphology compared with PrOH and EG in cryopreserved tissues. Xenografting has confirmed better efficacy of DMSO versus PrOH in adult but not in prepubertal tissues, probably owing to a greater capacity of younger animals to compensate for cryoinjury.

Differences in serum LH and FSH levels using depot or daily GnRH agonists in controlled ovarian stimulation: influence on ovarian response and outcome of ART

AbstractPurpose: To study effects of endogenous LH levels on ovarian response and outcome in ART cycles a controlled study was performed with two patient groups differing in the intensity of pituitary downregulation. Methods: Group I (n = 27) received 3.75 mg of the GnRH agonist triptorelin acetate depot, group II (n = 54) was given 0.1 mg triptorelin acetate daily, followed by ovarian stimulation with recombinant FSH. Results: After downregulation serum LH and FSH levels were significantly lower in group I. Patients of group I needed significantly higher FSH doses to achieve comparable levels of serum estradiol and preovulatory follicles. The number of retrieved oocytes and transferable embryos was lower in group I. Conclusion: Patients with profound endogenous LH suppression by depot GnRH agonists show higher FSH stimulation dose requirements and lower oocyte number and fertilization rate, indicating a need for minimal LH activity in folliculogenesis and oocyte development.

Association of Inhibin B Serum Levels with Parameters of Follicular Response in a Randomized Controlled Trial Comparing GnRH Agonist Versus Antagonist Protocols for Ovarian Hyperstimulation

AbstractPurpose: To study the association of inhibin B with ovarian response to FSH stimulation, applying either GnRH agonist or antagonist. Methods: In a prospective randomized controlled trial, 46 patients undergoing COH received either triptorelin (group I, n = 15) or ganirelix (group II, n = 31). Parameters of follicular response and inhibin B serum levels were assessed Results: Inhibin B before FSH stimulation was significantly lower in group I than group II. The FSH stimulation phase was significantly longer in group I than group II, and the total FSH dose was significantly higher with a comparable number of retrieved oocytes. Day 1 inhibin B in group I, but not group II, was significantly correlated with the number of large ovarian follicles and retrieved oocytes. In group II, but not group I, inhibin B on day 1 was inversely correlated with the daily and total FSH dose as well as FSH stimulation duration. Conclusions: The association of inhibin B serum levels with parameters of follicular response in COH is different in patients assigned to GnRH agonist vs. antagonist treatment protocols.

Reproduktionsmedizin. Ovarielle Stimulation – was gibt es Neues?

Unter Beibehaltung des klassischen Therapiekonzepts der Gonadotropinstimulation ermoglicht die Einfuhrung langwirksamer FSH-Praparate und der sogenannten FSH-Biosimilars in den vergangenen Jahren patientenbezogene Modifikationen bei der ovariellen Stimulation vor einer IVF/ICSI-Therapie. Zusatzlich tragt eine verbesserte Vorhersage des ovariellen Ansprechens zu dem Trend einer zunehmenden Individualisierung der Stimulation mit dem Ziel einer geringeren Patientenbelastung durch die Vermeidung ovarieller Uberstimulationssyndrome bei gleichzeitig hoherer Effektivitat bei.