Helge Binder

Association of FSH receptor and CYP19A1 gene variations with sterility and ovarian hyperstimulation syndrome

Severe ovarian hyperstimulation syndrome (OHSS) is a potentially life-threatening complication during assisted reproduction technology (ART). The aetiology of this condition is still not fully understood. Several gene variations in the FSH receptor (FSHR) gene have been identified for the very rare cases of spontaneous OHSS. There are only few published data on gene variations in sterility and iatrogenic OHSS and no data regarding aromatase (cytochrome P450 19A1; CYP19A1). Ninety-one ART patients with OHSS, eighty-eight ART patients without OHSS and ninety-seven women with assumed normal fecundity were analysed for the FSHR single nucleotide polymorphism (SNP) gene variations Asn680Ser (rs6166), Ala189Val, Ile160Thr, Thr449Ile (rs28928870) and the CYP19A1 rs10046 locus using real-time PCR. In addition, exon 10 of FSHR of two patients with spontaneous hyperreactio luteinalis (HL) was sequenced. Significantly lower frequencies of homozygous Ser680/Ser680 (P=0.035) and heterozygous Thr160/Ile160 (P=0.039) were found in patients with normal fecundity than those undergoing ART. The Ile160Thr SNP with a frequency of 6.7 and 6.1% in ART patients with and without OHSS respectively does not represent a rare mutation as previously published. There were no differences in the frequencies of all other gene variations. Of two patients with HL, both had homozygous point mutations for Ser680/Ser680 and one was heterozygous for Ile160Thr and CYP19A1 rs10046. The FSHR gene variations Asn680Ser as well as Ile160Thr may be contributing factors in unexplained sterility. The other FSHR coding gene variations and CYP19A1 rs10046 investigated are most likely not involved in the aetiology of iatrogenic OHSS or sterility.

The effect of a GnRH agonist on cryopreserved human ovarian grafts in severe combined immunodeficient mice

This prospective study compares the effect of a GnRH agonist on the number of follicles in different developmental stages in cryopreserved human ovarian grafts transplanted into gonadotropin-stimulated or not stimulated severe combined immunodeficient mice (SCID mice). Human ovarian tissue from seven patients was cryopreserved with an open-freezing system and xenotransplanted in SCID mice. The SCID mice were then treated according to different stimulation protocols. The survival of the tissue after cryopreservation was examined by LIVE/DEAD viability staining or transplanted in the neck muscle of 41 SCID mice. Development of follicles, estradiol production, vaginal cytology, and uterus weight were assessed after 15 weeks with or without gonadotropin stimulation. Viable follicles were detected in all frozen/thawed specimens using the LIVE/DEAD assay. Triptorelin, a GnRH agonist, caused a significant reduction of follicles in all developmental stages in the non-gonadotropin-stimulated animals (P<0.001). In gonadotropin-stimulated animals, GnRH agonist treatment has no significant effect on primordial, primary and preantral follicle count, whereas the antral follicles were significantly fewer (P = 0.03). The GnRH agonist treatment is not able to prevent the primordial follicle depletion after the xenografting of ovarian tissue in SCID mice with or without gonadotropin stimulation. Furthermore, it causes an additional loss of follicles if administered during the critical neovascularization period after the transplantation.

Low sex hormone-binding globulin as a predictive marker for insulin resistance in women with hyperandrogenic syndrome.

BACKGROUND The aim of the present study is to assess insulin resistance (IR) in women with hyperandrogenic syndrome, which was suggested to replace the term polycystic ovary syndrome by the Androgen Excess Society, and to evaluate whether sex hormone-binding globulin (SHBG) can be used as a predictive marker of IR in hyperandrogenic women. METHODS Clinical, metabolic, and endocrine parameters were measured, and an oral glucose tolerance test was carried out. The women were classified as IR group or non-IR group, in accordance with defined cutoff points for the homeostatic model assessment of IR (HOMA-IR) at > or =2.5, the quantitative insulin sensitivity check index at < or = 0.33, and the Matsuda insulin sensitivity index (ISI) at < or = 5. RESULTS The women classified as having IR had a significantly higher body mass index (BMI) and free androgen index (FAI) and showed significantly lower SHBG and high-density lipoprotein (HDL) levels, regardless of the indices used. However, with the Matsuda ISI, generally more women were diagnosed as having IR, and this group had significantly higher total testosterone and triglyceride values, as well as a higher incidence of hirsutism. CONCLUSIONS Women who were classified as being insulin resistant using insulin sensitivity indices showed significantly higher BMI and FAI values and lower SHBG and HDL levels. However, the Matsuda ISI may be more favorable for identifying IR in hyperandrogenic women. SHBG may serve as a predictive marker of IR in these women, particularly in those who are obese.

Blood group A: an overseen risk factor for early-onset ovarian hyperstimulation syndrome?

Ovarian hyperstimulation syndrome (OHSS), a potentially life-threatening complication, is classified into two distinct forms, early-onset and late-onset OHSS. Few risk factors have been established, but no association with ABO blood group antigens was known. From January 2000 to October 2007, 122 patients with known blood groups and the diagnosis of OHSS were hospitalized. OHSS classification, pregnancies, age and time of in-patient treatment were collated. Two control groups were established. One group comprised 177 patients treated for infertility without developing OHSS (treatment/no OHSS) and known blood groups. A second one consisted of 2289 obstetric and gynaecological patients (O/G). OHSS grade I, II or III was found in 20, 47 and 55 patients, respectively. The pregnancy rate was 50.8% and did not differ among the different OHSS grades. Blood group A was significantly more frequent and blood group O less frequent in patients with early-onset OHSS compared with the two control cohorts (P = 0.009 versus treatment/no OHSS; P = 0.001 versus O/G). The odds ratio for patients with blood group A versus O to develop early-onset OHSS was 2.169 and 2.262, respectively. No increased risk for late-onset OHSS was found. Blood group A may be associated with early-onset OHSS in Caucasians.

Endocrinological Markers for Assessment of Hyperandrogenemia in Hirsute Women

Background: Measured endocrinological parameters (total testosterone [TT], free testosterone [FT], dihydrotestosterone [DHT], dehydroepiandrosterone sulfate [DHEAS], and sex hormone binding globulin [SHBG]) and calculated parameters (calculated FT (cFT), calculated bioavailable testosterone (cBT), and the free androgen index [FAI]) in women with hirsutism were compared to the values of a control group. The question remains if cFT or cBT are more appropriate markers for assessment of hyperandrogenemia in clinical situations such as hirsutism in women. Methods: Sixty-six women showed an modified Ferriman-Gallwey (mF-G) score of ≧6 and were classified as hirsutism group and 58 women showed mF-G scores of ≤5 and were classified as control group. TT, FT, DHT, DHEAS, and SHBG were measured with enzyme immunoassays in one serum sample. Albumin was measured in the same sample. Afterwards, cFT, cBT, and the FAI were calculated. Results: The hirsutism group revealed significantly increased cFT, cBT, and FAI, while SHBG was significantly decreased compared to the controls. All values of the hirsutism group showed significant, but poor correlation with the mF-G score. SHBG correlated negatively with the mF-G score and BMI. Conclusion: Despite the limited correlation of all endocrinological parameters to severity of hirsutism described by the mF-G score, cFT, cBT, and FAI may be more appropriate markers for assessment of hyperandrogenemia in women with hirsutism in comparison with measured values of simple enzyme immunoassays.

Increased uterine arterial pressure and contractility of perfused swine uterus after treatment with serum from pre-eclamptic women and endothelin-1

The present study was designed to examine the effects of ET-1 (endothelin-1) and serum from PE (pre-eclamptic), HP (healthy pregnant) and HNP (healthy non-pregnant) women on uterine arterial perfusion pressure and uterine contractility. Swine uteri (n = 25) were perfused for a period of up to 11 h, with the aim being to preserve a viable organ. Various concentrations of ET-1 as well as serum from PE, HP and HNP women (n = 10 per group) were administered to the perfused swine uteri and IUP (intrauterine pressure) and IAP (intra-arterial pressure) were recorded. ET-1 produced dose-dependent increases in IUP and IAP. The ET-1 concentration in serum was higher in serum from PE women than in HP and HNP women (P > 0.05). Administration of all serum samples had a contractile effect on the swine uterus, with the greatest effect being seen in HNP women (12.8 +/- 5.3 mmHg), followed by PE (9.06 +/- 4.2 mmHg) and HP (6.1 +/- 4.1 mmHg) women. Statistically significant differences were observed between HNP and PE women (P = 0.048), and PE and HP women (P = 0.021). Increases in IAP following administration of serum from PE women (48.8 +/- 20.0 mmHg) were significantly higher (P = 0.024) compared with the effect of serum from HP women (28.4 +/- 13.7 mmHg). In conclusion, the findings show that serum from PE women has significant vasoconstrictive and oxytocic effects compared with serum from HP women. In pre-eclampsia, the balance between vasorelaxing and vasoactive substances is disturbed.

Assessment of FSHR variants and antimüllerian hormone in infertility patients with a reduced ovarian response to gonadotropin stimulation

OBJECTIVE To study women with a poor response to ovarian hormone stimulation, known as low responders. Genetic defects in the FSH receptor gene (FSHR) were analyzed as well as antimüllerian hormone (AMH) for ovarian reserve. DESIGN Retrospective cohort study. SETTING University hospital. PATIENT(S) Two hundred fifty-nine patients total: 74 low responders; 88 patients receiving assisted reproduction therapy (ART) with a normal ovarian response; and 97 women with a normal fertility status. INTERVENTION(S) DNA from patients was analyzed using real-time polymerase chain reaction. Serum concentrations of AMH were assessed using ELISA. MAIN OUTCOME MEASURE(S) The FSHR variants Asn680Ser (rs6166), Ala189Val (rs121909658), Ile160Thr (rs121909659), Thr449Ile (rs28928870) and the serum AMH concentrations were assessed. RESULT(S) With the exception of the frequent Asn680Ser polymorphism, no homozygotic SNPs of FSHR were found. In the group of ART patients, Thr160/Ile160 variants were more frequent in comparison with women with normal fertility. The Ser680/Ser680 was more frequent in ART patients than in women with normal reproductive function. The rate of live births was markedly reduced, particularly in the low responder group. No difference was noted in the distribution of the Ala189Val and Thr449Ile variant. Low serum AMH values were observed in 75% of the low responder group. CONCLUSION(S) FSHR gene variations such as Asn680Ser, Ala189Val, Thr449Ile, and Ile160Thr did not seem to be a decisive factor of poor response to fertility treatment, whereas the low ovarian reserve determined by AMH is considered more crucial.

First retransplantation of cryopreserved ovarian tissue following cancer therapy in Germany.

INTRODUCTION New anticancer treatments have increased survival rates for cancer patients but often at the cost of sterility. One way of preserving fertility in these patients is the use of cryopreservation of ovarian tissue with subsequent retransplantation following a period of recurrence-free survival. We report the follow-up of the first case of retransplantation of ovarian tissue in Germany. METHODS Immediately following the diagnosis of anal cancer, ovarian tissue was removed laparoscopically, and cryopreserved. The patient was then treated with combined radiochemotherapy, which resulted in iatrogenic premature ovarian failure, and was associated with inhibin B serum levels lower than 10 ng/L. After the 2.5 year period of cancer remission, the cryopreserved ovarian tissue was retransplanted orthotopically. RESULTS Five months later estradiol serum levels had risen from lower than 20 pg/mL to 436 pg/mL. Three ovarian follicles were detected ultrasonographically in the pelvic side wall. Finally the patient reported her first menstruation after the intervention. The endocrine activity of the transplanted cryopreserved tissue has demonstrated viability, and the ability to develop. DISCUSSION Cycle monitoring and timed intercourse should now help to achieve conception. These first results from Germany for retransplantation of cryopreserved ovarian tissue clearly show its potential for preserving fertility.

Uterus Cryopreservation: Maintenance of Uterine Contractility by the Use of Different Cryoprotocols

Cryopreservation of cells and even tissue is feasible. New exciting findings arise in the promising field of cryobiology, e.g. the cryopreservation of whole ovaries. Uterus cryopreservation would be advantageous not only for experimental biology, but also for transplantation surgery. The objective of this study was to evaluate various cryopreservation protocols as well as various storage temperatures in cryopreservation of whole swine uteri. The used freezing protocol was slow (0.2 degrees C/min) after arterial perfusion with 1%, 5% or 10% dimethyl sulfoxide (DMSO) solution for 10 min and equilibration in this solution for 30 min. Viability of the organs was tested by histological examination, biochemical parameters and by the capability of rhythmical contractions in a perfusion system. Eighty swine uteri were cryopreserved. All uteri that were frozen with 10% and 5% DMSO were viable after thawing for at least 1 h, whereas only 40% survived with the use of 1% DMSO and 0% with the use of 0.5% DMSO, respectively. There was no difference regarding the survival rates after various cryostorage periods for up to 16 weeks or after cryostorage for 2 weeks in -70 degrees C or -130 degrees C. The cryopreservation of a whole organ such as the swine uterus is a valuable method for the study of cryoprotective agents and freezing protocols. This study demonstrates clearly that the perfusion of the organ with cryoprotectants is the only factor which allows the uterus to contract.

Course of Ovarian Hyperstimulation Syndrome in 19 Intact Twin Pregnancies after Assisted Reproduction Techniques, with a Case Report of Severe Thromboembolism

Ovarian hyperstimulation syndrome (OHSS) is a serious complication of assisted reproduction techniques using in vitro fertilization and intracytoplasmic sperm injection (IVF/ICSI). Its etiology has still not yet been fully resolved. Human chorionic gonadotrophin, administered exogenously as well as produced endogenously during pregnancy, is responsible for the onset of OHSS, and high levels of estradiol appear to worsen the condition. In this case series, the course of mild to severe OHSS was evaluated in 19 intact twin pregnancies after IVF/ICSI. Another serious complication associated with OHSS is thromboembolic events. In these cases, the pregnancy can be protected through anticoagulation treatment, but there may be exceptions to this. This series includes a case of bilateral thrombosis of the internal and external jugular veins in the 7th gestational week in a twin pregnancy after OHSS and ICSI, with termination of the pregnancy in the 9th gestational week due to progressive thrombosis during anticoagulation therapy.