Inge Hoffmann

Live birth after ovarian tissue autotransplantation following overnight transportation before cryopreservation

OBJECTIVE To describe the first live birth after transplantation of ovarian tissue following overnight transportation of the tissue before freezing. DESIGN Technical note. SETTING University department of obstetrics and gynecology. PATIENT(S) A 25-year-old cancer survivor with previous Hodgkin disease and relapse. INTERVENTION(S) The ovarian tissue was kept cool for >20 hours in a special transport medium and a special cooling device before it was cryopreserved. After premature ovarian failure due to preconditioning chemotherapy for bone marrow transplantation, the cryopreserved ovarian tissue was transplanted orthotopically. MAIN OUTCOME MEASURE(S) Resumption of ovarian function after transplantation, recovery of fertility, and pregnancy. RESULT(S) Ovarian function returned in the patient 3 months after transplantation, as shown by follicle development and estrogen production. During the fifth menstrual cycle, mild stimulation with FSH was initiated in accordance with a low-dose protocol. When ultrasonography revealed a follicle 18-20 mm in size in the ovarian graft, hCG was added and the patient had sexual intercourse at the optimal time point. On day 14 of the luteal phase, hCG concentration and vaginal echography confirmed a viable intrauterine pregnancy, which resulted in a healthy live birth. CONCLUSION(S) Overnight transportation of ovarian tissue appears to be possible in combination with appropriate transportation logistics. However, further investigations are needed before this procedure can be offered as a chance for women to preserve fertility independently of direct access to a tissue-processing bank.

Pregnancies and live births after 20 transplantations of cryopreserved ovarian tissue in a single center.

OBJECTIVE To report the results of 20 orthotopic retransplantations of cryopreserved ovarian tissue after cancer treatment. DESIGN Retrospective analysis. SETTING Tertiary gynecology department. PATIENT(S) Twenty patients with malignant disease: 11 with hematological malignancies (55%), four with breast cancer (20%), three with anal cancer (15%), and two with ovarian cancer (10%); the mean age before oncological treatment was 30.5 years. INTERVENTION(S) Ovarian tissue was removed from patients in various centers in Germany in 2005-2009. All patients received chemotherapy and/or radiotherapy. Afterward, 17 patients had complete premature ovarian insufficiency, while three still showed some ovarian activity. Overnight transportation of tissue before freezing was necessary in eight cases. Cryopreservation followed slow freezing protocols in all cases. Retransplantation was performed at Erlangen University Hospital 3.75 years after extraction, on average. Thawed tissue was transplanted into a peritoneal pouch in the broad ligament region, below the tube, in 16 cases. Fragments were sutured both onto the remaining ovary and into a peritoneal pouch in four cases. MAIN OUTCOME MEASURE(S) Restoration of ovarian activity, pregnancy, birth. RESULT(S) Ovarian activity resumed in all patients except one. Seven patients conceived, with one miscarriage and four ongoing pregnancies. Four patients delivered healthy babies. One pregnancy and live birth after oocyte donation need to be considered separately. CONCLUSION(S) These data clearly demonstrate that preserving fertility by cryopreserving ovarian tissue is a successful and safe clinical option that can be considered for selected cancer patients.

Thyroid hormone receptors and reproduction.

Thyroid disorders have a great impact on fertility in both sexes. Hyperthyroidism and hypothyroidism cause changes in sex hormone-binding globulin (SHBG), prolactin, gonadotropin-releasing hormone, and sex steroid serum levels. In females, thyroid hormones may also have a direct effect on oocytes, because it is known that specific binding sites for thyroxin are found on mouse and human oocytes. There is also an association between thyroid dysfunction in women and morbidity and outcome in pregnancy. In males, hyperthyroidism causes a reduction in sperm motility. The numbers of morphologically abnormal sperm are increased by hypothyroidism. When euthyroidism is restored, both abnormalities improve or normalize. In women, the alterations in fertility caused by thyroid disorders are more complex. Hyper- and hypothyroidism are the main thyroid diseases that have an adverse effect on female reproduction and cause menstrual disturbances--mainly hypomenorrhea and polymenorrhea in hyperthyroidism, and oligomenorrhea in hypothyroidism. In recent studies, it has become evident that it is not only changes in serum levels of SHBG and sex steroids that are responsible for these disorders, but also alterations in the metabolic pathway. Adequate levels of circulating thyroid hormones are of primary importance for normal reproductive function. This review presents an overview of the impact of thyroid disorders on reproduction.

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.

Xenotransplantation of cryopreserved ovarian tissue from patients with ovarian tumors into SCID mice—no evidence of malignant cell contamination

This study examined the possible presence of malignant cells in ovarian cortex from patients with ovarian tumors after xenografting of the ovarian tissue into severe combined immunodeficiency mice. None of the mice presented symptoms of reintroduced malignancy nor did microscopic and immunohistochemical evaluation of the grafts raise any suspicion of residual malignant disease.

Akt and p53 are potential mediators of reduced mammary tumor growth by Chloroquine and the mTOR inhibitor RAD001

PI3K/Akt/mTOR and p53 signaling pathways are frequently deregulated in tumors. The anticancer drug RAD001 (everolimus) is a known mTOR-inhibitor, but mTOR-inhibition leads to phosphorylation of Akt inducing resistance against RAD001 treatment. There is growing evidence that conflicting signals transduced by the oncogene Akt and the tumorsuppressor p53 are integrated via negative feedback between the two pathways. We previously showed that the anti-malarial Chloroquine, a 4-alkylamino substituted quinoline, is a p53 activator and reduced the incidence of breast tumors in animal models. Additionally, Chloroquine is an effective chemosensitizer when used in combination with PI3K/Akt inhibitors but the mechanism is unknown. Therefore, our aim was to test, if Chloroquine could inhibit tumor growth and prevent RAD001-induced Akt activation. Chloroquine and RAD001 caused G1 cell cycle arrest in luminal MCF7 but not in mesenchymal MDA-MB-231 breast cancer cells, they significantly reduced MCF7 cell proliferation on a collagen matrix and mammospheroid formation. In a murine MCF7 xenograft model, combined treatment of Chloroquine and RAD001 significantly reduced mammary tumor growth by 4.6-fold (p = 0.0002) compared to controls. Chloroquine and RAD001 inhibited phosphorylation of mTOR and its downstream target, S6K1. Furthermore, Chloroquine was able to block the RAD001-induced phosphorylation of Akt serine 473. The Chloroquine effect of overcoming the RAD001-induced activation of the oncogene Akt, as well as the promising antitumor activity in our mammary tumor animal model present Chloroquine as an interesting combination partner for the mTOR-inhibitor RAD001.

Differences in muscarinic-receptor agonist-, oxytocin-, and prostaglandin-induced uterine contractions.

OBJECTIVE To investigate the contractile response of the perfused swine uterus to various receptor pathways (oxytocin, prostaglandins, and muscarine). DESIGN An extracorporeal perfusion model of the swine uterus was used that keeps the uterus in a functional condition and is appropriate for the study of physiologic questions. INTERVENTION(S) Oxytocin-, prostaglandin-, and carbachol-induced uterine contractility and peristalsis were assessed using an intrauterine double-chip microcatheter. SETTING University hospital. MAIN OUTCOME MEASURE(S) Intrauterine pressure profiles. RESULT(S) A dose-dependent increase in intrauterine pressure (IUP) in the isthmus uteri and corpus uteri was observed after the administration of prostaglandin F(2alpha) (PGF(2alpha)) and oxytocin, which reached a plateau after further stimulation. A dose-dependent increase in IUP in the isthmus uteri and corpus uteri was also observed after the administration of prostaglandin E(1) (PGE(1)) and prostaglandin E(2) (PGE(2)), with a plateau in IUP in the middle-concentration range and a decrease during the further course of stimulation. After administration of PGE(1), PGE(2), and PGF(2alpha), different directions of contraction waves were also observed. Carbachol also showed a unique contractility pattern, with isolated, very powerful, dose-dependent contractions with an IUP gradient, suggesting directed transport from the upper region to the lower region. CONCLUSION(S) This study demonstrated that oxytocin, the prostaglandins, and carbachol modulate contractility in nonpregnant swine uteri in a characteristic way, resulting in different contractility patterns.

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.

Association of thyroid-stimulating hormone with insulin resistance and androgen parameters in women with PCOS

There is a relationship between thyroid function and insulin sensitivity and alterations in lipids and metabolic parameters. Little information is available regarding this relationship in women with polycystic ovary syndrome. However all those pathologies are also described as often affecting women with polycystic ovary syndrome. The association between thyroid-stimulating hormone <2.5 mIU/l and > or =2.5 mIU/l with insulin resistance and endocrine parameters in 103 women with polycystic ovary syndrome was studied. Clinical, metabolic and endocrine parameters were obtained and an oral glucose tolerance test was performed with calculation of insulin resistance indices. Women with thyroid-stimulating hormone > or =2.5 mIU/l had a significantly higher body mass index (P = 0.003), higher fasting insulin concentrations (P = 0.02) and altered insulin resistance indices (P = 0.007), higher total testosterone (P = 0.009) and free androgen indices (P = 0.001) and decreased sex hormone-binding globulin concentrations (P = 0.01) in comparison with women with thyroid-stimulating hormone <2.5 mIU/l. Generally, all of these parameters correlated significantly (P < 0.05) with thyroid-stimulating hormone only in women with thyroid-stimulating hormone > or =2.5 mIU/l. Women with polycystic ovary syndrome and with thyroid-stimulating hormone > or =2.5 mIU/l had significantly altered endocrine and metabolic changes.

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.