Cortisol/glucocorticoid receptor: a critical mediator of the ovulatory process and luteinization in human periovulatory follicles.

STUDY QUESTION: Do cortisol/glucocorticoid receptors play an active role in the human ovary during ovulation and early luteinization? SUMMARY ANSWER: The ovulatory hCG stimulation-induced glucocorticoid receptor signaling plays a crucial role in regulating steroidogenesis and ovulatory cascade in human periovulatory follicles. WHAT IS KNOWN ALREADY: Previous studies reported an increase in cortisol levels in the human follicular fluid after the LH surge or ovulatory hCG administration. However, little is known about the role of cortisol/glucocorticoid receptors in the ovulatory process and luteinization in humans. STUDY DESIGN, SIZE, DURATION: This study was an experimental prospective clinical and laboratory-based study. An in vivo experimental study was accomplished utilizing the dominant ovarian follicles from 38 premenopausal women undergoing laparoscopic sterilization. An in vitro experimental study was completed using the primary human granulosa/lutein cells (hGLC) from 26 premenopausal women undergoing IVF. PARTICIPANTS/MATERIALS, SETTING, METHODS: This study was conducted in a private fertility clinic and academic medical centers. Dominant ovarian follicles were collected before the LH surge and at defined times after hCG administration from women undergoing laparoscopic sterilization. Primary hGLC were collected from women undergoing IVF. hGLC were treated without or with hCG in the absence or presence of RU486 (20 µM; dual antagonist for progesterone receptor and glucocorticoid receptor) or CORT125281 (50 µM; selective glucocorticoid receptor antagonist) for 12 or 36 h. The expression of genes involved in glucocorticoid receptor signaling, steroidogenesis, and ovulatory cascade was studied with RT-quantitative PCR and western blotting. The production of cortisol, corticosterone, and progesterone was assessed by hormone assay kits. MAIN RESULTS AND THE ROLE OF CHANCE: hCG administration upregulated the expression of hydroxysteroid 11-beta dehydrogenase 1 (HSD11B1), nuclear receptor subfamily 3 group C member 1 (NR3C1), FKBP prolyl isomerase 5 (FKBP5), and FKBP prolyl isomerase 4 (FKBP4) in human ovulatory follicles and in hGLC (P < 0.05). RU486 and CORT125281 reduced hCG-induced increases in progesterone and cortisol production in hGLC. The expression of genes involved in glucocorticoid receptor signaling, steroidogenesis, and the key ovulatory process was reduced by RU486 and/or CORT125281 in hGLC. LARGE SCALE DATA: N/A. LIMITATIONS, REASONS FOR CAUTION: The role of cortisol/glucocorticoid receptors demonstrated using the hGLC model may not fully reflect their physiological roles in vivo. WIDER IMPLICATIONS OF THE FINDINGS: Successful ovulation and luteinization are essential for female fertility. Women with dysregulated cortisol levels often suffer from anovulatory infertility. Deciphering the functional role of glucocorticoid receptor signaling in human periovulatory follicles enhances our knowledge of basic ovarian physiology and may provide therapeutic insights into treating infertility in women. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by P01HD71875 (to M.J., T.E.C., and M.B.) and R01HD096077 (to M.J.) from the Foundation for the National Institutes of Health and the BTPSRF of the University of Kentucky Markey Cancer Center (P30CA177558). The authors report no competing interests. TRIAL REGISTRATION NUMBER: N/A.

Unilateral inguinofemoral lymphadenectomy in patients with early-stage vulvar squamous cell carcinoma and a unilateral metastatic sentinel lymph node is safe.

OBJECTIVE: Optimal management of the contralateral groin in patients with early-stage vulvar squamous cell carcinoma (VSCC) and a metastatic unilateral inguinal sentinel lymph node (SN) is unclear. We analyzed patients who participated in GROINSS-V I or II to determine whether treatment of the contralateral groin can safely be omitted in patients with a unilateral metastatic SN. METHODS: We selected the patients with a unilateral metastatic SN from the GROINSS-V I and II databases. We determined the incidence of contralateral additional non-SN metastases in patients with unilateral SN-metastasis who underwent bilateral inguinofemoral lymphadenectomy (IFL). In those who underwent only ipsilateral groin treatment or no further treatment, we determined the incidence of contralateral groin recurrences during follow-up. RESULTS: Of 1912 patients with early-stage VSCC, 366 had a unilateral metastatic SN. Subsequently, 244 had an IFL or no treatment of the contralateral groin. In seven patients (7/244; 2.9% [95% CI: 1.4%-5.8%]) disease was diagnosed in the contralateral groin: five had contralateral non-SN metastasis at IFL and two developed an isolated contralateral groin recurrence after no further treatment. Five of them had a primary tumor ≥30 mm. Bilateral radiotherapy was administered in 122 patients, of whom one (1/122; 0.8% [95% CI: 0.1%-4.5%]) had a contralateral groin recurrence. CONCLUSION: The risk of contralateral lymph node metastases in patients with early-stage VSCC and a unilateral metastatic SN is low. It appears safe to limit groin treatment to unilateral IFL or inguinofemoral radiotherapy in these cases.

Differential effects of the immunosuppressive calcineurin inhibitors cyclosporine-A and tacrolimus on ovulation in a murine model.

STUDY QUESTION: Do therapeutic levels of cyclosporine-A and tacrolimus affect ovulation in a rat gonadotrophin-induced ovulation model? SUMMARY ANSWER: Cyclosporine-A, but not tacrolimus, decreases ovulation rate when administered for 5 days before induced ovulation. WHAT IS KNOWN ALREADY: The mainstays of immunosuppression in solid organ transplantation, to prevent rejection, are the calcineurin inhibitors cyclosporine-A or tacrolimus. These drugs could potentially affect fertility in transplanted patients. Since ovulation is an inflammation-like process with pivotal roles for several immune cells and modulators, it is possible that the calcineurin inhibitors, with broad effects on the immune system, could interfere with this sensitive, biological process. STUDY DESIGN SIZE DURATION: Experimental design at university-based animal facilities. A total of 45 immature Sprague-Dawley rats were used. The study was carried out over 3 months. PARTICIPANTS/MATERIALS SETTING METHODS: Immature Sprague-Dawley rats (n = 45) were randomly assigned to receive equivalent doses of tacrolimus (0.5 mg/kg/day; TAC), cyclosporine-A (10 mg/kg/day; CyA) or vehicle (Control). Ovarian hyperstimulation was induced with 10 IU of equine chorionic gonadotrophin, and ovulation was triggered with 10 IU of hCG. Oocytes were retrieved from the oviducts and ovulation rates were calculated. Various subpopulations of white blood cells were counted in peripheral blood and ovarian tissue samples. MAIN RESULTS AND THE ROLE OF CHANCE: Animals in the CyA group showed a lower ovulation rate when compared to the TAC and Control groups (CyA: mean 9 oocytes (range 0-22); TAC: 21 oocytes (8-41); Control: 22 oocytes (6-39); P = 0.03). Regarding counts of the white blood cell subpopulations and resident neutrophils in the ovary, no significant differences were observed between the groups. LIMITATIONS REASONS FOR CAUTION: Although the ovulation process is highly conserved within species, the differences between rodents and humans may limit the external translatability of the study. WIDER IMPLICATIONS OF THE FINDINGS: These findings suggest that tacrolimus should be the preferred calcineurin inhibitor of choice in transplanted patients who are aiming for pregnancy. STUDY FUNDING/COMPETING INTERESTS: Swedish Research Council and ALF of Sahlgrenska Academy, Sweden. Rio Hortega Grant from the Instituto de Salud Carlos III, Spain (CM09/00063). There are no conflicts of interest.

Type 2 diabetes mellitus in women with polycystic ovary syndrome during a 24-year period: importance of obesity and abdominal fat distribution.

STUDY QUESTION: What are the predictive factors for later development of type 2 diabetes (T2DM) in women with polycystic ovary syndrome (PCOS)? SUMMARY ANSWER: Obesity and abdominal fat distribution in women with PCOS in the mid-fertile years were the major risk factors for T2DM development 24 years later when lifestyle factors were similar to controls. WHAT IS KNOWN ALREADY: Women with PCOS have an increased prevalence of T2DM. STUDY DESIGN SIZE DURATION: A longitudinal and cross-sectional study was performed. Women with PCOS were examined in 1992 and in 2016. Randomly selected, age-matched women from the general population served as controls. PARTICIPANTS/MATERIALS SETTING METHODS: Women with PCOS (n = 27), attending an outpatient clinical at a tertiary care centre for infertility or hirsutism were diagnosed in 1992 (mean age 30 years) and re-examined in 2016 (mean age 52 years). Women from the World Health Organization MONItoring of trends and determinants for CArdiovascular disease (WHO MONICA-GOT) 2008, aged 38-68 years, served as controls (n = 94), and they were previously examined in 1995. At both at baseline and at follow-up, women had blood samples taken, underwent a clinical examination and completed structured questionnaires, and the women with PCOS also underwent a glucose clamp test at baseline. MAIN RESULTS AND THE ROLE OF CHANCE: None of women with PCOS had T2DM at baseline. At the 24-year follow-up, 19% of women with PCOS had T2DM versus 1% of controls (P < 0.01). All women with PCOS who developed T2DM were obese and had waist-hip ratio (WHR) >0.85 at baseline. No difference was seen between women with PCOS and controls regarding use of high-fat diet, Mediterranean diet or amount of physical activity at follow-up at peri/postmenopausal age. However, women with PCOS had a lower usage of a high-sugar diet as compared to controls (P = 0.01). The mean increases in BMI and WHR per year were similar in women with PCOS and controls during the follow-up period. LIMITATIONS REASONS FOR CAUTION: The small sample size of women with PCOS and the fact that they were recruited due to infertility or hirsutism make generalization to women with milder forms of PCOS uncertain. WIDER IMPLICATIONS OF THE FINDINGS: Obesity and abdominal fat distribution, but not hyperandrogenism per se, in women with PCOS in the mid-fertile years were the major risk factors for T2DM development 24 years later when peri/postmenopausal. Lifestyle factors were similar to controls at that time. STUDY FUNDING/COMPETING INTERESTS: The study was financed by grants from the Swedish state under the agreement between the Swedish government and the country councils, the ALF-agreement (ALFGBG-718611), the Gothenburg Medical Association GLS 694291 and 780821, the Swedish Heart Lung Foundation and Hjalmar Svensson Foundation. The authors have no conflict of interest.

Towards uterus tissue engineering: a comparative study of sheep uterus decellularisation.

Uterus tissue engineering may dismantle limitations in current uterus transplantation protocols. A uterine biomaterial populated with patient-derived cells could potentially serve as a graft to circumvent complicated surgery of live donors, immunosuppressive medication and rejection episodes. Repeated uterine bioengineering studies on rodents have shown promising results using decellularised scaffolds to restore fertility in a partially impaired uterus and now mandate experiments on larger and more human-like animal models. The aim of the presented studies was therefore to establish adequate protocols for scaffold generation and prepare for future in vivo sheep uterus bioengineering experiments. Three decellularisation protocols were developed using vascular perfusion through the uterine artery of whole sheep uteri obtained from slaughterhouse material. Decellularisation solutions used were based on 0.5% sodium dodecyl sulphate (Protocol 1) or 2% sodium deoxycholate (Protocol 2) or with a sequential perfusion of 2% sodium deoxycholate and 1% Triton X-100 (Protocol 3). The scaffolds were examined by histology, extracellular matrix quantification, evaluation of mechanical properties and the ability to support foetal sheep stem cells after recellularisation. We showed that a sheep uterus can successfully be decellularised while maintaining a high integrity of the extracellular components. Uteri perfused with sodium deoxycholate (Protocol 2) were the most favourable treatment in our study based on quantifications. However, all scaffolds supported stem cells for 2 weeks in vitro and showed no cytotoxicity signs. Cells continued to express markers for proliferation and maintained their undifferentiated phenotype. Hence, this study reports three valuable decellularisation protocols for future in vivo sheep uterus bioengineering experiments.

Experiences of a multistep process with medical and psychological interventions for patients with congenital uterine aplasia to achieve motherhood: the Gothenburg-Tübingen collaboration.

Congenital uterine aplasia, also known as Mayer-Rokitansky-Küster-Hauser syndrome (MRKHS) is a condition associated to a non-functional uterus in the presence of functional ovaries. In a setting where surrogacy is illegal (or not accepted) and adoption is the only alternative, neovaginoplasty and subsequent uterus transplantation (UTx) can provide a route to motherhood for women with MRKHS. This review article describes a multistep process by which patients with MRKHS can achieve motherhood with their own biological child. This process involving a careful clinical diagnosis, psychological counselling, assessment of eligibility for neovagina creation and UTx, the surgical treatment, fertility treatment, and long-term follow-up was developed at the Tübingen University Hospital and in close collaboration with Sahlgrenska Academy, University of Gothenburg, Sweden, where the basic experimental and clinical groundwork for UTx was laid and the first-ever UTx procedure was performed.

Monitoring of Human Uterus Transplantation With Cervical Biopsies: A Provisional Scoring System for Rejection.

Until now, absolute uterine factor infertility has been the major untreatable form of female infertility. Uterus transplantation has recently proven to be the first successful treatment for absolute uterine factor infertility, with demonstration of live births. In this study, live donation uterus transplantation was performed in nine women. In total, 163 cervical biopsies (149 protocol, 14 follow-up) were taken to detect histopathological signs of rejection. Based on experience from animal experiments, we used a three-grade scoring system to evaluate biopsies systematically. Nine episodes of rejection were diagnosed in five patients: grade 1 in six episodes, grade 2 in two episodes, and grade 3 in one episode. Treatment decisions were based on histopathology, and all rejection episodes were reversed after treatment. The biopsies were reviewed retrospectively, and immunohistochemistry was performed to characterize the inflammatory infiltrates. A borderline category was introduced to avoid overtreatment of patients. Based on our review of all biopsies, we put forward a simple grading system for monitoring of rejection and to guide immunosuppressive treatment in uterus transplantation.

Obstetric and neonatal outcome of pregnancies fathered by males on immunosuppression after solid organ transplantation.

Immunosuppressive drugs may influence spermatogenesis, but little is known about outcome of pregnancies fathered by transplanted males. We estimated risk of adverse outcomes in pregnancies (with data after the first trimester) fathered by males that had undergone organ transplantation and were treated with immunosuppression. A population-based study, linking data from the Norwegian transplant registry and the Medical Birth Registry of Norway during 1967-2009 was designed. All Norwegian men undergoing solid organ transplantation were included. Odds ratios for major malformations, preeclampsia, preterm delivery (<37 weeks) and small-for-gestational-age were obtained using logistic regression. A total of 2463 transplanted males, fathering babies of 4614 deliveries before and 474 deliveries after transplantation were identified. The risk of preeclampsia was increased (AOR: 7.4, 95% CI: 1.1-51.4,) after transplantation compared to prior to transplantation. No increased risk was found for congenital malformations or other outcomes when compared with pregnancies before transplantation or with the general population (2 511 506 births). Our results indicate an increased risk of preeclampsia mediated through the transplanted and immunosuppressed father. Importantly, no increased risk was found for other adverse obstetric outcomes or malformations, which may reassure male transplant recipients planning to father children.

Towards the development of a bioengineered uterus: comparison of different protocols for rat uterus decellularization.

Uterus transplantation (UTx) may be the only possible curative treatment for absolute uterine factor infertility, which affects 1 in every 500 females of fertile age. We recently presented the 6-month results from the first clinical UTx trial, describing nine live-donor procedures. This routine involves complicated surgery and requires potentially harmful immune suppression to prevent rejection. However, tissue engineering applications using biomaterials and stem cells may replace the need for a live donor, and could prevent the required immunosuppressive treatment. To investigate the basic aspects of this, we developed a novel whole-uterus scaffold design for uterus tissue engineering experiments in the rat. Decellularization was achieved by perfusion of detergents and ionic solutions. The remaining matrix and its biochemical and mechanical properties were quantitatively compared from using three different protocols. The constructs were further compared with native uterus tissue composition. Perfusion with Triton X-100/dimethyl sulfoxide/H2O led to a compact, weaker scaffold that showed evidence of a compromised matrix organization. Sodium deoxycholate/dH2O perfusion gave rise to a porous scaffold that structurally and mechanically resembled native uterus better. An innovative combination of two proteomic analyses revealed higher fibronectin and versican content in these porous scaffolds, which may explain the improved scaffold organization. Together with other important protocol-dependent differences, our results can contribute to the development of improved decellularization protocols for assorted organs. Furthermore, our study shows the first available data on decellularized whole uterus, and creates new opportunities for numerous in vitro and in vivo whole-uterus tissue engineering applications.

Preclinical report on allogeneic uterus transplantation in non-human primates.

STUDY QUESTION: Is it possible to perform allogeneic uterus transplantation (UTx) with a donation from a live donor in a non-human primate species and what immunosuppression is needed to prevent rejection? SUMMARY ANSWER: Allogeneic UTx in the baboon is a donor- and recipient-safe surgical procedure; immunosuppression with induction therapy and a triple protocol should be used. WHAT IS KNOWN ALREADY: UTx may become a treatment for absolute uterine factor infertility. Autologous UTx models have been developed in non-human primates with reports on long-term survival of the uterine grafts. STUDY DESIGN, SIZEAND DURATION: This experimental study included 18 female baboons as uterus donors and 18 female baboons as uterus recipients. The follow-up time was 5-8 weeks. PARTICIPANTS/MATERIALS, SETTING AND METHODS: Uterus retrieval was performed with extended hysterectomy including bilateral uterine and internal iliac arteries and ovarian veins. After UTx, with vascular anastomoses unilateral to the internal iliac artery and the external iliac vein, the uterus recipients received one of the following: no immunosuppression (n = 4); monotherapy (oral slow release tacrolimus) (n = 4) or induction therapy (antithymocyte globulin) followed by triple therapy (tacrolimus, mycophenolate, corticosteroids; n = 10). Surgical parameters, survival, immunosuppression and rejection patterns were evaluated. MAIN RESULTS AND THE ROLE OF CHANCE: The durations of uterus retrieval and recipient surgery were around 3 and 3.5 h, respectively. The total ischemic time was around 3 h. All the recipients and the donors survived the surgery. All the recipients presented rejection to some extent within the first weeks following UTx. In one recipient, the uterus was of normal appearance at the end of the study period. In spite of occasional high (>60 ng/ml) blood levels of tacrolimus, there was no evidence of nephrotoxicity. LIMITATIONS AND REASONS FOR CAUTION: This initial non-human primate allogeneic UTx study indicates that further research is needed to optimize immunosuppression protocols in order to avoid uterine rejection. WIDER IMPLICATIONS OF THE FINDINGS: The findings suggest that allogeneic UTx in primate species is feasible but continued work on this issue is needed. STUDY FUNDING/COMPETING INTEREST(S): The study was funded by the Swedish Research Council, ALF University of Gothenburg, Hjalmar Svensson Foundation and by Jane and Dan Olsson Research Foundation. The authors do not have any competing interest.

Uterus transplantation in a non-human primate: long-term follow-up after autologous transplantation.

BACKGROUND: Uterus transplantation (UTx) may provide the first available treatment for women affected by uterine infertility. The present study aimed to further develop a surgical technique for autologous UTx in a non-human primate species and to assess long-term function. METHODS: Female baboons (n= 16) underwent autologous transplantation of the uterus with the Fallopian tubes and ovaries, performed with a previously published surgical technique (n= 6, Group 1) or using a modified technique (n= 10; Group 2). The uterine arteries were dissected to the proximal end of the anterior branch (Group 1) or the entire (Group 2) internal iliac artery, and the ovarian veins were dissected to the crossing over the ureter (Group 1) or further cranially to include greater lengths and patches of the cava/renal vein (Group 2). Back-table preparation created common venous and arterial ends with arterial anastomosis either end-to-side to the left external iliac artery (Group 1) or end-to-end to the left internal iliac artery (Group 2). RESULTS: Overall short-time survival of the animals was 88% (66% in Group 1 and 100% in Group 2). Of all the operated animals, 75% (66% in Group 1 and 80% in Group 2) resumed ovarian cyclicity. Regular menstruation after UTx was demonstrated only in Group 2 (60%). Menstruating animals (n= 6) were each exposed to timed mating for ≥5 menstrual cycles, but pregnancy did not occur. Adhesions and tubal blockage were seen in post-mortem analysis. CONCLUSIONS: The modified UTx model of Group 2 is a safe procedure and shows resumed long-term uterine function in a majority of the animals, although pregnancy could not be demonstrated.

Uterine transplantation research: laboratory protocols for clinical application.

The aim of this review is to summarize the state-of the-art methods that are used in clinical organ transplantation today, as well as the major findings of recent experimental uterus transplantation (UTx) research regarding organ donation/retrieval, ischemic preservation, surgical techniques for anastomosis, immunosuppression and pregnancy. Absolute uterine factor infertility lacks treatment despite the major developments in infertility treatment and assisted reproduction. Concerning uterine factor infertile patients, genetic motherhood is only possible through gestational surrogacy. The latter can pose medical, ethical and legal concerns such as lack of control of life habits during surrogate pregnancy, economic motives for women to become surrogate mothers, medical/psychological pregnancy-related risks of the surrogate mother and uncertainties regarding the mother definition. Thus, surrogacy is non-approved in large parts of the world. Recent advances in the field of solid organ transplantation and experimental UTx provide a favourable and safe background in a scenario in which a human clinical UTx trial can take place. Protocols based on animal research over the last decade are described with a view to providing a scientifically guided approach to human UTx as an experimental procedure in the future.

Ovarian cortex transplantation in the baboon: comparison of four different intra-abdominal transplantation sites.

BACKGROUND: Several sites have been used for ovarian cortex transplantation (OCT) in humans. The present study was designed to evaluate different intra-abdominal transplantation sites in the baboon to gain further knowledge about alternative transplantation sites in a human setting. METHODS: Autologous fresh OCTs were performed in 12 baboons (Papio anubis). Four different sites were tested: the free portion of the omentum (OMF), the portion of the omentum adjacent to the spleen (OMS), the pouch of Douglas (D) and the pelvic wall on the psoas muscle (PW). Cortex survival, follicle density, cyclicity and hormonal levels were compared between the different sites, 3 and 6 months after transplantation. RESULTS: Macroscopically, antral follicles were only found in the OMS and OMF locations, which also showed a higher proportion of follicle-containing cortex at light microscopy (OMF 71.4%, OMS 83.3% versus PW 58.8% and D 40%, P< 0.05). Higher densities of primordial [OMF: 3.54 (0-13.18) follicles/grid, OMS: 3.85 (0-8.53), PW: 0 (0-13.25), D 0 (0-1.33), P< 0.05] and primary follicles [OMF: 3.54 (0-18.52), OMS: 3.85 (0-1), PW: 0 (0-4.58), D 0 (0-0.25), P< 0.05] was also found in the omental locations. CONCLUSIONS: Omental locations provide a better site, in terms of follicle survival, for intra-abdominal OCT in the baboon compared with the pelvic wall and the D.

Fertility after autologous ovine uterine-tubal-ovarian transplantation by vascular anastomosis to the external iliac vessels.

BACKGROUND: Transplantation of the uterus has been suggested as a treatment of uterine factor infertility. This study investigates whether the sheep uterus can resume its capacity to harbour normal pregnancies after autotransplantation by vascular anastomosis. METHODS: From 14 ewes, the uterus, excluding one uterine horn, was isolated along with its oviduct and ovary and preserved ex vivo and then transplanted back with end-to-side anastomosis of the vessels of the graft to the external iliac vessels. After recovery, the ewes underwent surgical examination and serum progesterone measurements to ascertain healing and ovarian activity. Afterwards, five autotransplanted and five control ewes were placed with a ram for mating. Caesarean sections were performed before the estimated term of pregnancy and data on fetal measures were compared. RESULTS: Of the 14 ewes, seven survived surgery with ovarian activity intact and grafts showing normal appearance. Mating occurred in four of five transplanted ewes and in five out of five controls, and three transplanted animals and five control animals conceived. In one transplanted ewe, torsion of the uterus was observed after spontaneous initiation of labour. Foeti from transplanted mothers were comparable in size to those of controls. CONCLUSIONS: Despite the encountered complications, this is the first report to demonstrate fertility and pregnancies going to term after autotransplantation of the uterus in an animal of a comparable size to the human.

Uterus transplantation in the baboon: methodology and long-term function after auto-transplantation.

BACKGROUND: Techniques for uterus transplantation (UTx) have been developed in rodent/domestic animals towards future clinical introduction of UTx to treat uterine factor infertility. The aim of this study was to extend the UTx research into a non-human primate species by developing surgical techniques for uterus retrieval and transplantation in the baboon. METHODS: Female baboons (n = 15) underwent surgery, with the initial five animals used for studies of pelvic vascular anatomy. Retrieval surgery included isolation of the ovarian veins and the uterine arteries together with the anterior branches of the internal iliacs. The utero-tubal-ovarian specimen was removed, flushed and kept ex vivo for 2 h when the two arterial ends and two venous ends were anastomosed side-to-side to construct one arterial and one venous end. These were, at auto-transplantation, anastomosed end-to-side to the external iliacs and the animals (n = 10) were evaluated concerning cyclicity and later by laparoscopy/laparotomy. RESULTS: The total duration of organ retrieval, backtable preparation and transplantation was around 6 h with an overall ischaemic time of the specimen of about 3 h. One animal died due to cardiomyopathy. Five out of the nine surviving animals resumed cyclicity, as a sign of re-established ovarian function. Only two out of these five animals exhibited resumed menstruation, indicating re-established ovarian and uterine function. Laparoscopy confirmed normal-sized uteri in these two animals. CONCLUSIONS: This study demonstrates the feasibility of UTx by vascular anastomosis in a non-human primate species. The low success rate demonstrates the complexity involved in UTx surgery and the need for further methodological developments.

Cyclosporine A exposure during pregnancy in mice: effects on reproductive performance in mothers and offspring.

BACKGROUND: Pregnancies after organ transplantation and under immunosuppressive treatment are associated with slightly elevated risks for obstetric and post-natal complications but can usually be managed well. However, little is known about the effects of intrauterine exposure (IUE) to immunosuppressants in the growing and adult offspring. One major issue is the potentially negative effects of immunosuppressive medication on reproduction. This study investigates the effect of exposure during pregnancy to the most commonly used immunosuppressant in organ transplantation, cyclosporine A (CsA), on the reproductive outcome in mothers and offspring. METHODS: Female C57CBA-F1 mice received 0, 10, 20 or 30 mg/kg bodyweight of CsA daily by subcutaneous mini-osmotic pumps during mating and pregnancy. Blood concentrations of CsA, implantation rates, resorption rates and fetal weights were analysed. In addition, female and male mice exposed to CsA in utero were mated to unexposed partners and pregnancy outcomes were analysed. RESULTS: Direct maternal exposure to CsA at high doses reduced implantation rates and fetal survival. IUE to CsA reduced adolescent growth but did not affect fertility, although a reduction in birthweight was seen in offspring of females exposed to CsA in utero. CONCLUSIONS: CsA exposure during pregnancy correlates with impaired reproductive outcome, but offspring fertility is not affected. The cause of reduction in adolescent weight gain and low birthweight in offspring of females exposed to CsA in utero need further investigation.

Rejection of allogenic uterus transplant in the mouse: time-dependent and site-specific infiltration of leukocyte subtypes.

BACKGROUND: Animal models of uterus transplantation are being developed ahead of a possible treatment for absolute uterus infertility in women. Our knowledge of inflammatory cell involvement in acute rejection of a uterus transplant is limited; therefore, we examined the pattern of invasion of leukocyte subtypes into an allogeneic uterus transplant. METHODS: The uterus and its vasculature were removed from BALB/c mice and transplanted into C57Bl/6 recipient mice at a heterotopic position, with the native uterus left in situ. Both uteri were removed on post-operative day 2 (D2, n = 5), D5 (n = 5) and D10 (n = 6). Immunohistochemistry for neutrophilic granulocytes, macrophages, cytotoxic CD8(+) T-cells, CD4(+) T-helper cells and B-cells was performed and cell density was evaluated in both myometrium and endometrium. RESULTS: Neutrophil density was increased in graft versus native uteri at D5 and D10 in myometrium and D10 in endometrium, and in endometrium was higher in the D5 than D2 graft (all P < 0.05). Infiltration of macrophages occurred from D2 in myometrium and from D5 in endometrium (P < 0.05, graft versus native). Density of CD8(+) cytotoxic T-cells increased in the graft versus native uteri at D5 in both uterine layers and for the graft versus D2 density (P < 0.01). In contrast, CD4(+) T-helper cells increased only transiently in graft endometrium at D5 (P < 0.05). Overall CD19(+) B-cell density was low, with no time-dependent changes in graft myometrium or endometrium. CONCLUSIONS: Acute rejection of an allogeneic uterus transplant in the mouse involves influx of predominately neutrophils, macrophages, CD8(+) T-cells and CD4(+) T-cells between D2 and D5 post-operatively.