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Sinan Ozkavukcu, Nilay Kuscu, Dileyra Adiguzel, Guldane Cengiz-Seval, and Ciler Celik-Ozenci

Nilotinib is a second-generation tyrosine kinase inhibitor (TKI) that is widely used to treat patients with Philadelphia chromosome-positive chronic myeloid leukaemia (CML). TKIs provided a significant improvement in terms of survival rates and disease-free period in CML; however, there is insufficient knowledge about their side effects, including reproductive toxicity. Since nearly half of the CML patients are in their reproductive age, and newly announced indications cover the treatment of the paediatric age groups, concerns arise about the effects of these drugs on the reproductive system, as there are no controlled preclinical studies. We investigated acute and long-term gonadotoxic and teratogenic effects of nilotinib, utilising a mouse model that simulates various clinical scenarios. We observed significant testicular damage in mice receiving nilotinib according to Johnsen’s score analysis. Alterations were observed in female mice’s number of follicles, as the primordial follicle numbers significantly decreased. Proliferating cell number in both genders’ gonads decreased and apoptosis rate increased significantly. The nilotinib-received female and male mice’s pregnancy rates were low compared to controls. A significant decrease in the thickness of the spongiotrophoblast and decidual layers of the placenta was detected in pregnancies consisting of male and/or female mice treated with nilotinib. The results of this study establish a critical point of view for clinical translation and indicate the importance of consulting patients for directing them to fertility preservation and contraception options for both genders before nilotinib treatment.

Free access

T Leahy, J P Rickard, N C Bernecic, X Druart, and S P de Graaf

Ejaculation results in the confluence of epididymal spermatozoa with secretions of the accessory sex glands. This interaction is not a prerequisite for fertilisation success, but seminal factors do play a crucial role in prolonging the survival of spermatozoa both in vitro and in vivo by affording protection from handling induced stress and some selective mechanisms of the female reproductive tract. Reproductive biologists have long sought to identify specific factors in seminal plasma that influence sperm function and fertility in these contexts. Many seminal plasma proteins have been identified as diagnostic predictors of sperm function and have been isolated and applied in vitro to prevent sperm damage associated with the application of artificial reproductive technologies. Proteomic assessment of the spermatozoon, and its surroundings, has provided considerable advances towards these goals and allowed for greater understanding of their physiological function. In this review, the importance of seminal plasma will be examined through a proteomic lens to provide comprehensive analysis of the ram seminal proteome and detail the use of proteomic studies that correlate seminal plasma proteins with ram sperm function and preservation ability.

Free access

T Tharasanit, S Colleoni, G Lazzari, B Colenbrander, C Galli, and T A E Stout

Oocyte cryopreservation is a potentially valuable way of preserving the female germ line. However, the developmental competence of cryopreserved oocytes is presently poor. This study investigated whether the morphology of the cumulus complex surrounding an immature equine oocyte and/or the oocyte’s stage of maturation affect its cryopreservability. Compact (Cp) and expanded (Ex) cumulus oocyte complexes (COCs) were vitrified either shortly after recovery (germinal vesicle stage, GV) or after maturation in vitro (IVM); cryoprotectant-treated and -untreated non-frozen oocytes served as controls. In Experiment I, oocytes matured in vitro and then vitrified, or vice versa, were examined for maturation stage and meiotic spindle quality. Cp and Ex COCs vitrified at the GV stage matured at similar rates during subsequent IVM (41 vs 46% MII), but meiotic spindle quality was better for Cp than Ex (63 vs 33% normal spindles). Vitrifying oocytes after IVM resulted in disappointing post-warming spindle quality (32 vs 28% normal for Cp vs Ex). In Experiment II, oocytes from Cp and Ex COCs vitrified at the GV or MII stages were fertilized by intracytoplasmic sperm injection (ICSI) and monitored for cleavage and blastocyst formation. Oocytes vitrified prior to IVM yielded higher cleavage rates (34 and 27% for Cp and Ex COCs) than those vitrified after IVM (16 and 4%). However, only one blastocyst was produced from a sperm-injected vitrified–warmed oocyte (0.4 vs 9.3% and 13% blastocysts for cryoprotectant-exposed and -untreated controls). It is concluded that, when vitrification is the chosen method of cryopreservation, Cp equine COCs at the GV stage offer the best chance of an MII oocyte with a normal spindle and the potential for fertilization; however, developmental competence is still reduced dramatically.

Free access

Georgios Anifandis, Eleni Koutselini, Ioannis Stefanidis, Vassilios Liakopoulos, Constantinos Leivaditis, Themis Mantzavinos, and Nikolaos Vamvakopoulos

This prospective study was undertaken to reassess the prognostic value of leptin during critical stages of in vitro fertilization-embryo transfer (IVF-ET) and address its role in the functional staging of assisted reproductive technologies at the level of embryo quality. Serum and follicular fluid samples of 100 selected women undergoing the long IVF-ET protocol were collected for leptin and embryo quality determination. The highest serum leptin concentration (52.11 ± 4.27 ng/ml) was observed on ovum pick up day, while follicular fluid leptin was higher than all serum samples examined (62.59 ± 5.73 ng/ml). Serum leptin above 59.48 ± 7.6 ng/ml was associated with ‘poor’ embryo quality and above 56.87 ± 5.52 ng/ml with pregnancy failure. Elevated leptin concentrations were associated with reduced ovarian stimulation and response, follicle maturation, embryo quality and pregnancy success. Our findings suggest that leptin modulates embryo quality and may serve as a sensitive marker of IVF outcome.

Open access

Konstantina Nikolakopoulou and Margherita Y Turco

Infertility is a common problem in modern societies with significant socio-psychological implications for women. Therapeutic interventions are often needed which, depending on the cause, can either be medical treatment, surgical procedures or assisted reproductive technology (ART). However, the treatment of infertility is not always successful due to our limited understanding of the preparation of the lining of the uterus, the endometrium, for pregnancy. The endometrium is of central importance for successful reproduction as it is the site of placental implantation providing the interface between the mother and her baby. Due to the dynamic, structural and functional changes the endometrium undergoes throughout the menstrual cycle, it is challenging to study. A major advancement is the establishment of 3D organoid models of the human endometrium to study this dynamic tissue in health and disease. In this review, we describe the changes that the human endometrium undergoes through the different phases of the menstrual cycle in preparation for pregnancy. We discuss defects in the processes of endometrial repair, decidualization and acquisition of receptivity that are associated with infertility. Organoids could be utilized to investigate the underlying cellular and molecular mechanisms occurring in non-pregnant endometrium and early pregnancy. These studies may lead to therapeutic applications that could transform the treatment of reproductive failure.

Free access

E. A. Lenton, M. Hooper, H. King, A. Kumar, N. Monks, S. Verma, and J. Osborn

Sheffield Fertility Centre, 26 Glen Road, Sheffield, S7 1RA, UK and University Department of Obstetrics and Gynaecology, Jessop Hospital for Women, Leavygreave Road, Sheffield, S3 7RE, UK

Keywords: implantation; in-vitro fertilization; pregnancy; pregnancy loss; human chorionic gonadotrophin; human

Introduction

The endocrine characteristics of normal human pregnancy have been difficult to establish, chiefly because spontaneous pregnancies occur unpredictably. More reliable sources of early pregnancy data are conceptions following various assisted reproductive technologies although, unfortunately, many of these may not be useful for determining normal physiology, firstly, because there is multiple follicle development resulting from the use of exogenous gonadotrophins and, secondly, because human chorionic gonadotrophin (hCG) given to induce luteinization masks hCG from the implanting embryo. Furthermore, the practice, at least for in-vitro fertilization (IVF), of replacing up to 3 embryos renders assessment of the number of implantation sites uncertain.

In-vitro fertilization in the natural or spontaneous cycle may provide

Free access

J. Carroll, H. Depypere, and C. D. Matthews

Summary. Frozen–thawed oocytes have a reduced rate of fertilization (48·8%) when compared with unfrozen controls (97%). In this study we have used zona-drilling to bypass the zona pellucida and investigate whether the decreased rate of fertilization is due to freezing-induced changes in the zona pellucida which prevent sperm penetration. After zona drilling the fertilization rate of frozen–thawed oocytes (87·8%) was the same as for zona-intact unfrozen controls (88%), indicating that freeze–thaw-induced changes at the level of the zona pellucida were responsible for the decreased rate of fertilization.

To determine whether the changes were occurring during the manipulations before and after freezing or the complete freeze–thaw cycle, oocytes were exposed to the complete set of manipulations normally experienced during cryopreservation and appropriate control groups. A small but significant decrease in the rate of fertilization (82·8%) was apparent in oocytes exposed to the manipulations before and after freezing compared with controls (92·2%). The freeze–thaw-induced changes in the zona pellucida therefore occur primarily during the complete freeze–thaw cycle itself and not the manipulations before and after freezing and are responsible for the decreased rate of fertilization observed in frozen–thawed oocytes.

Keywords: cryopreservation; oocyte; zona pellucida; in-vitro fertilization; mouse

Free access

Lindsay A Hogan, Tina Janssen, and Stephen D Johnston

This review provides an update on what is currently known about wombat reproductive biology and reports on attempts made to manipulate and/or enhance wombat reproduction as part of the development of artificial reproductive technology (ART) in this taxon. Over the last decade, the logistical difficulties associated with monitoring a nocturnal and semi-fossorial species have largely been overcome, enabling new features of wombat physiology and behaviour to be elucidated. Despite this progress, captive propagation rates are still poor and there are areas of wombat reproductive biology that still require attention, e.g. further characterisation of the oestrous cycle and oestrus. Numerous advances in the use of ART have also been recently developed in the Vombatidae but despite this research, practical methods of manipulating wombat reproduction for the purposes of obtaining research material or for artificial breeding are not yet available. Improvement of the propagation, genetic diversity and management of wombat populations requires a thorough understanding of Vombatidae reproduction. While semen collection and cryopreservation in wombats is fairly straightforward there is currently an inability to detect, induce or synchronise oestrus/ovulation and this is an impeding progress in the development of artificial insemination in this taxon.

Free access

Mi Hou, Margareta Andersson, Chengyun Zheng, Anne Sundblad, Olle Söder, and Kirsi Jahnukainen

Testicular germ cell transplantation is a novel strategy for preservation of fertility in prepubertal cancer patients, but the risk of reseeding tumor cells into cured patients presently limits clinical application of this approach. To date, no systematic evaluation of the limitations of surface marker-based decontamination of testicular samples with acute lymphoblastic leukemia has been performed. Here, surface markers for leukemic (CD4 and major histocompatibility complex class I) and germ cells (epithelia cell adhesion molecule) in testicular samples infiltrated with Roser’s T-cell leukemia were identified. These markers were then used to delete leukemic cells and/or select for germ cells by flow cytometry (FACS). The resulting cell populations were analyzed by FACS, immunocytochemistry, or evaluation of leukemia transmission in syngeneic piebald variegated rats. Simple positive selection of germ cells or deletion of leukemic cells using specific surface markers was unable to effectively decontaminate testicular samples. The poor specificity of spermatogonial surface markers and aggregation of germ and leukemic cells limited the positive selection of germ cells, while immunophenotypic variation among lymphoblastic leukemia cells prevented adequate deletion of leukemic cells. Enzymatic treatment to disperse the testicular cells and feature of the intratesticular environment contributed to this immunophenotypic variation. Only germ cell selection in combination with leukemic cell deletion prevented leukemia transmission in association with intratesticular injection of the sorted cells. However, with such combined sorting, only 0.23% of the original testicular cells were recovered. With presently available techniques, flow cytometric purification of germ cells from a leukemic donor is not sufficiently effective or safe for clinical use.

Free access

Bruce F Kimler, Shawn M Briley, Brian W Johnson, Austin G Armstrong, Susmita Jasti, and Francesca E Duncan

Radiation damage due to total body irradiation (TBI) or targeted abdominal radiation can deplete ovarian follicles and accelerate reproductive aging. We characterized a mouse model of low-dose TBI to investigate how radiation affects the follicular and stromal compartments of the ovary. A single TBI dose of either 0.1 Gy or 1 Gy (Cesium-137 γ) was delivered to reproductively adult CD1 female mice, and sham-treated mice served as controls. Mice were euthanized either 2 weeks or 5 weeks post exposure, and ovarian tissue was harvested. To assess the ovarian reserve, we classified and counted the number of morphologically normal follicles in ovarian histologic sections for all experimental cohorts using an objective method based on immunohistochemistry for an oocyte-specific protein (MSY2). 0.1 Gy did not affect that total number of ovarian follicles, whereas 1 Gy resulted in a dramatic loss. At two weeks, there was a significant reduction in all preantral follicles, but early antral and antral follicles were still present. By five weeks, there was complete depletion of all follicle classes. We examined stromal quality using histologic stains to visualize ovarian architecture and fibrosis and by immunohistochemistry and quantitative microscopy to assess cell proliferation, cell death and vasculature. There were no differences in the ovarian stroma across cohorts with respect to these markers, indicating that this compartment is more radio-resistant relative to the germ cells. These findings have implications for reproductive health and the field of fertility preservation because the radiation doses we examined mimic scatter doses experienced in typical therapeutic regimens.