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Levels of apoptosis in human granulosa cells seem to be comparable after therapy with a gonadotropin-releasing hormone agonist or antagonist.

OBJECTIVE: To compare levels of apoptosis in granulosa cells from women treated with the gonadotropin-releasing hormone (GnRH) agonist triptorelin or the GnRH antagonist cetrorelix. DESIGN: Randomized, prospective study. SETTING: University hospital. PATIENT(S): Thirty-two women undergoing assisted reproduction techniques after ovulation induction with recombinant follicle-stimulating hormone (FSH) plus GnRH agonist or antagonist. INTERVENTION(S): Granulosa cells were isolated from follicular aspirates after oocyte removal. MAIN OUTCOME MEASURE(S): Apoptosis was assessed with Annexin V binding assay, terminal deoxynucleotidyl transferase (TdT)-mediated nick-end labeling (TUNEL) assay, flow cytometric analysis of DNA, and ultrastructural analysis of cell morphology in transmission electron microscopy. Serum and follicular hormonal levels were also determined. RESULT(S): Annexin V binding and TUNEL assays revealed comparable percentages of apoptosis in the two groups under investigation. Analysis of DNA histograms revealed a similar cell cycle distribution in the two groups. Ultrastructural analysis only occasionally displayed patterns of chromatin margination in apoptotic cells. The mean concentrations of all the follicular fluid steroid hormones evaluated (E2, T, and P) were significantly lower in the GnRH antagonist-treated group. CONCLUSION(S): Therapy with a GnRH agonist or antagonist is associated with comparable levels of apoptosis in granulosa cells.

Adult↗

Development and evaluation of a qualitative reverse-transcriptase nested polymerase chain reaction protocol for same-day viral validation of human immunodeficiency virus type 1 ribonucleic acid in processed semen.

OBJECTIVE: To develop a method for same-day validation of processed semen in the setting of assisted reproductive techniques (ART) with patients who are seropositive for human immunodeficiency virus, type 1 (HIV-1). DESIGN: Laboratory experiments. SETTING: University hospital. PATIENT(S): Volunteers who are HIV-1 seronegative and seropositive. INTERVENTION(S): Evaluation of the sensitivity of a reverse-transcriptase (RT)-nested polymerase chain reaction (PCR) in HIV-1 RNA-positive blood plasma, in artificially infected blood plasma and semen, and in 85 semen samples of 29 HIV-1-seropositive volunteers. Semen was submitted to gradient separation, followed by swim-up. MAIN OUTCOME MEASURE(S): Qualitative detection of HIV-1 RNA in blood plasma and in different parts of semen preparation by using RT-nested PCR, PCR inhibition control by dilution of samples, and an internal control. RESULT(S): The detection limit of our PCR was 20 HIV-1 RNA copies per milliliter. Among seropositive patients, RNA was detected in 25% of fresh semen, 36.5% of seminal plasma, 27.5% of gradient supernatants, and 7.1% of final preparations before the migration-sedimentation stage. Positive final preparations were observed in patients who had blood viral loads of >/=20,000 HIV-1 RNA copies per milliliter. Inhibition was present in 17.6% of seminal plasma and in 20% gradient supernatants and in 2 final preparations among 69 tested. Among 25 preparations tested after the migration-sedimentation stage, 2 were positive (1 patient; 70,000 HIV-1 RNA copies per milliliter). CONCLUSION(S): The RT-nested PCR detects low viral load and allows the validation of semen preparations of HIV-1-seropositive patients for ART on the day of sampling. For this purpose, the validation is performed on spermatozoa that are obtained after gradient separation before swim-up. Inhibition of the PCR must be controlled by using an internal control that is well-designed to explore the detection limit of the method.

Cells, Cultured↗

[Male genital tract infection: the point of view of the virologist].

Attention to viral infection of the male genital tract has been renewed over the last 15 years as a result of the prolific ongoing research on AIDS. Epidemiological studies of the virus in sperm and male genital tract contributes to the understanding of STD physiopathology and helps assessing their impact on male fertility. Recent advances in this field have allowed to offer Assisted reproductive techniques to couples with chronic viral infection, under strict and specific protocols. This paper presents an overview of these recent developments.

Cross Infection↗

Myomas and assisted reproductive technologies: when and how to act?

The effect of myomas on reproductive outcome has been the subject of many studies; however, a definitive answer is still missing. Therefore, the authors have tried to outline some guidelines for the management of women who have uterine myomas and desire to conceive. The location and size of the myomas are the two parameters that influence the success of a future pregnancy. Subserosal myomas seem to have little, if any,effect on reproductive outcome, especially if they are up to 5 to 7 cm in diameter. Intramural myomas that do not encroach upon the endometrium also can be considered to be relatively harmless to reproduction, if they are smaller than 4 to 5 cm in diameter. This is the ambiguous gray zone of the subject, and where research should be focused before a consensus can be established. Myomas that compress the uterine cavity with an intramural portion (submucous myoma type II) and submucous myomas significantly reduce pregnancy rates, and should be removed before assisted reproductive techniques are used. Hysteroscopic myomectomy is the gold standard for the treatment of submucous myomas. For other myomas, abdominal myomectomy, or laparoscopic myomectomy--when the experience of the surgeon and the facilities are sufficient--are the best alternatives. In most of the literature, the pregnancy rates were increased and the miscarriage rates were decreased after surgery with these two techniques. Other alternative treatment modalities, such as CUV, laparoscopic myolysis, or MRI-guided focused ultrasound, are to be monitored and evaluated thoroughly before they are applied as routine procedures.

Clinical Trials as Topic↗

Ovarian control for assisted reproduction in the domestic cat and wild felids.

Of the 37 felid species, all but the domestic cat are classified as threatened with extinction in all or part of their native range. Additionally, the domestic cat is a valuable model for human biomedical research. Propagating some wild felids as well as domestic cat populations serving as human models is a major challenge primarily due to difficulties in transporting animals between facilities to ensure the pairing of genetically matched individuals, behavioral incompatibility between pairs and low fertility. Artificial insemination (AI) and in vitro fertilization/embryo transfer (IVF/ET) are powerful tools for helping manage rare populations. Developing successful assisted reproductive techniques requires knowledge of the female reproductive cycle and precise control of ovarian activity. Successful ovarian stimulation for AI and IVF/ET has been achieved in at least one-third of all cat species. However, sensitivity to a given gonadotropin treatment appears highly species-specific, and poor responses are common, particularly in felid species that exhibit spontaneous ovulations. Furthermore, current gonadotropin regimens have been demonstrated to perturb female reproductive function often leading to reduced fertility. Overall, ovarian response to exogenous hormonal stimulation has been highly variable, and pregnancy success after AI or IVF/ET remains low (<20%) in most species. Therefore, there is an immediate need to develop improved regimens that would allow more predictable ovarian responses in felids. We contend that recent research involving the use of progestins to control the ovary prior to gonadotropin stimulation shows promise for providing consistent ovarian stimulation in felids.

Animals↗

Updated meta-analysis of recombinant follicle-stimulating hormone (FSH) versus urinary FSH for ovarian stimulation in assisted reproduction.

OBJECTIVE: To determine the relative efficacy of recombinant FSH (rFSH) and urinary FSH (uFSH) for ovarian stimulation in assisted reproductive techniques (ART). DESIGN: Systematic review and meta-analysis of randomized, controlled trials comparing rFSH and uFSH. SETTING: Infertility centers providing treatment with ART. PATIENT(S): Patients undergoing IVF with or without ICSI. INTERVENTION(S): Controlled ovarian stimulation using uFSH or rFSH (follitropin alpha or follitropin beta). MAIN OUTCOME MEASURE(S): Primary: rate of clinical pregnancy per cycle. Secondary: rates of spontaneous abortion, multiple pregnancy, and severe ovarian hyperstimulation syndrome (OHSS); total gonadotropin dose; serum E(2) level and number of follicles at hCG administration; number of oocytes retrieved. RESULT(S): Eighteen trials were included in the meta-analysis. Subgroup analyses demonstrated higher pregnancy rates with both follitropins compared with uFSH. However, statistical significance was reached only in the follitropin alpha versus uFSH comparison in IVF cycles, with an additional pregnancy for every 19 patients treated. Fewer units of rFSH than uFSH achieved the same E(2) level and oocyte yield. No differences were found between treatments in rates of spontaneous abortion, OHSS, and multiple gestation. CONCLUSION(S): rFSH produced higher pregnancy rates per cycle than uFSH when follitropin alpha was used in IVF, and the total gonadotropin dose required was lower.

Abortion, Spontaneous↗

Embryo technologies in the horse.

Recent studies demonstrated that zwitterionic buffers could be used for satisfactory storage of equine embryos at 5 degrees C. The success of freezing embryos is dependent upon size and stage of development. Morulae and blastocysts <300 microm can be slowly cooled or vitrified with acceptable pregnancy rates after transfer. The majority of equine embryos are collected from single ovulating mares, as there is no commercially available product for superovulation in equine. However, pituitary extract, rich in FSH, can be used to increase embryo recovery three- to four-fold. Similar to human medicine, assisted reproductive techniques have been developed for the older, subfertile mare. Transfer of in vivo-matured oocytes from young, healthy mares into a recipient's oviduct results in a 70-80% pregnancy rate compared with a 30-40% pregnancy rate when the oocytes are from older, subfertile mares. This procedure can also be used to evaluate in vitro maturation systems. In vitro production of embryos is still quite difficult in the horse. However, intracytoplasmic sperm injection (ICSI) has been used to produce several foals. Cleavage rates of 60% and blastocyst rates of 30% have been reported after ICSI of in vitro-matured oocytes. Gamete intrafallopian tube transfer (GIFT) is a possible treatment for subfertile stallions. Transfer of in vivo-matured oocytes with 200,000 sperm into the oviduct of normal mares resulted in a pregnancy rate of 55-82%. Oocyte freezing is a technique that has proven difficult in most species. However, equine oocytes vitrified in a solution of ethylene glycol, DMSO, and Ficoll and loaded onto a cryoloop resulted in three pregnancies of 26 transfers and two live foals produced. Production of a cloned horse appears to be likely, as several cloned pregnancies have recently been produced.

Animals↗

Clinical value of preimplantation genetic diagnosis.

The clinical application of preimplantation genetic diagnosis (PGD) has provided an alternative approach for the prevention of affected pregnancies in couples at high reproductive risk. The frequent contribution of genetic factors to infertility problems makes PGD of particular value for assisted reproductive practices. In addition, the selection of euploid embryos for transfer has a strong impact on IVF efficiency as aneuploidies are the main cause of spontaneous abortions and implantation failures. In this study, the clinical outcome in PGD cycles is presented. The list of monogenic disorders for which PGD is performed is rapidly extending and the safety of the procedure has lead to an increasing interest among couples at high reproductive risk. Following PGD for aneuploidy, a higher implantation rate and a lower incidence of spontaneous abortions are obtained in patient categories where aneuploidy is a prominent cause of reproductive failure. In view of these findings, PGD has become an integral part of assisted reproductive techniques for the prevention of affected pregnancies and improvement of IVF efficiency.

Adult↗

Vasovasostomy: multiple children and long-term pregnancy rates.

PURPOSE: To determine the long-term patency and pregnancy rates after vasovasostomy and to determine the likelihood of having more than 1 child after this procedure. MATERIALS AND METHODS: We performed a retrospective examination of 73 patients that were at least 4 years out from vasovasostomy at our institution. Patients and their partners completed questionnaires that focused on their pre- and post-procedure fertility, vasal obstructive interval, time to pregnancy, and the number of children conceived. All patients were asked to obtain a current semen analysis. RESULTS: From a population of 73 patients with proven prior fertility, 43 could be contacted for data collection. Of the 43 men, 39 (91%) reported that they had actively tried to father children. These 39 men are a mean of 84.8 months (range, 4 to 10 years) out from their surgery, with a mean vasal obstructive interval of 87 months (7.4 years). All men denied using assisted reproduction techniques except the father of the only twins, who reported the use of ICSI for female-factor infertility. He was excluded from further pregnancy calculations. Forty-five percent of couples achieved at least 1 pregnancy, whereas 6 of 17 (35%) conceived 2 children. Mean time to conception was 14.7 months for the first child and 51 months for the second (range, 13 to 108 months). In those that did not conceive (n = 21), the mean obstructive interval was 106 months versus 81 months for those that did conceive. This time interval was not statistically different (P > 0.05). Mean maternal age was significantly less in those that did versus those that did not conceive (32.2 vs 36.6 years) (P < 0.01). We also noted that 10 of 21 were with a different spouse that had never been pregnant. Three of these 10 had known female-factor fertility problems. Semen analyses were obtained in 6 of the 21 patients that did not father children. Only 1 of the 6 was azoospermic. Thirteen (33%) desired more than 1 child at the time of vasectomy reversal. CONCLUSIONS: After vasovasostomy in patients with a vasal obstructive interval of 7 years, up to 45% of couples may achieve pregnancy and up to 35% could go on to conceive a second child. Long-term anastomosis patency after the procedure is estimated to be approximately 60%. Maternal age is a significant factor in predicting a couple's success in conceiving a child. Vasovasostomy remains a highly effective option for restoration of fertility in vasectomy patients, while offering the opportunity for multiple pregnancies with only a single intervention.

Anastomosis, Surgical↗

[Sophrology: a different tool for infertile couples].

Because of the high degree of complexity of assisted reproduction techniques (ART), the human and conscious dimensions of infertility problems are often neglected. Different strategies may help infertile couples coping with infertility and related treatments; among these, Caycedian sophrology relies on the cognitive, emotional, and somatic aspects of consciousness. In the present article, the authors report on their experience with sophrologic support for infertile patients by a midwife qualified in caycedian sophrology. Overall, since 1988, 310 couples have benefied from this kind of support, with an average of 10 sophrologic trainings per patient. Whereas some couples consider sophrology as a short time training to better cope with any particular aspect of their infertility treatment, others want to undertake more profound work on their body scheme. The authors wish to call the attention of ART professionals to this kind of medical support for infertile couples, and also to the particular role of midwives with sophrologic competence in an ART center.

Adult↗

Current concepts and novel applications of LH activity in ovarian stimulation.

Luteinizing hormone (LH) is a crucial physiological regulator of the human menstrual cycle. LH activity is also contained in many medications used to treat anovulation and to stimulate multiple folliculogenesis for assisted reproduction techniques. However, LH activity had previously been regarded as just a contaminant of follicle-stimulating hormone (FSH)-containing products and deemed potentially detrimental for reproductive function. Novel experimental and clinical evidence now suggests that the administration of pharmacological amounts of LH activity, instead of being harmful, is therapeutically advantageous, particularly in the support and modulation of ovarian folliculogenesis. The aim of this article is to provide an overview of the effects of LH activity administration in ovarian stimulation and to outline novel unconventional gonadotropin regimens that might improve the efficacy, safety and convenience of ovulation induction.

Anovulation↗

The potential risks of abnormal transmission of mtDNA through assisted reproductive technologies.

The recent introduction of more invasive assisted reproductive techniques offers the possibility to provide a wider treatment profile to patients. However, some of these technologies are of considerable concern as they are fraught with the possible transmission of genetic abnormalities to the offspring they create. To date, much analysis of these technologies has been conducted at the chromosomal DNA level. While some analysis has been conducted on the extranuclear, mitochondrial genome (mtDNA), this has been mainly descriptive. In the vast majority of cases, it appears that mtDNA is maternally inherited. The impact that leakage of sperm mtDNA transmission might have for the offspring is discussed in the light of the recent identification of sperm mtDNA presence in a patient with mtDNA disease. The implications of introducing donor mtDNA into a recipient oocyte through both cytoplasmic and nuclear transfer are also discussed. Again, the implications for offspring survival are discussed and suggestions made as to why the techniques might provide valuable insights into mtDNA transmission, replication and transcription. In order to be confident that patients and their offspring are being offered safe treatment, it is argued that potentially some of these treatments may be of considerable benefit in the future but significant scientific research is required before these treatments can be effectively employed in the clinic.

Animals↗

To add or not to add LH: comments on a recent commentary.

From a period of time when LH activity supplementation in stimulation protocols for assisted reproductive technology was questioned, recent scientific evidence seems to prove a role of LH activity in optimizing the clinical results of certain subgroups. Until now, apart from the hypogonadotrophic hypogonadal patient, two subgroups of patients have attracted attention: patients over 35 years of age, and normogonadotrophic patients with an initial low response to recombinant FSH (rFSH). However, in a recent paper, another subgroup of patients has been identified: normogonadotrophic patients with high endogenous LH levels (>1.99 IU/l) on day 8 of stimulation after a long GnRH agonist down-regulation. By adding recombinant LH (rLH) from day 8 to the stimulation protocol, this subgroup of patients achieved good implantation rates, similar to those of patients with lower LH levels. Interestingly, the dynamics of the endogenous LH levels during down-regulation in this group of patients differed from those of patients with lower LH levels. The immunoreactivity of the LH molecule does not necessarily reflect the bioactivity, which might explain the beneficial effect of LH activity supplementation in patients with high endogenous LH levels after down-regulation. Future studies should be focused on subgroups to further explore the effect of LH activity supplementation in assisted reproduction techniques.

Adult↗

Embryo implantation: the molecular mechanism remains elusive.

Low rates of implantation are an impediment to more efficient assisted reproduction techniques. Improved endometrial receptivity and embryo preparation should lead to higher pregnancy rates, lower rates of early pregnancy failure and fewer multiple pregnancies. As the first site of contact between embryo and endometrium, the luminal epithelium (LE) is responsible for the non-receptive status of proliferative and early secretory tissue, and transformation to receptivity in the mid-secretory phase presumably requires alterations in expression, organization or activation of adhesion systems. Luminal cells are less abundant than their glandular counterparts, and are under-represented in global tissue datasets. Furthermore, alterations in cell surface composition can be readily accomplished by mechanisms that do not rely on altered transcription or translation. Current data from in-vitro models are consistent with initial attachment to mucin in the apical glycocalyx, perhaps via a carbohydrate-mediated interaction, after which the epithelial phenotype is modified by a medium- or short-range embryonic signal. A cascade of interactions follows, mediating embryo migration across the epithelium. Strikingly, numerous potential mediators of adhesion at implantation are located in the lateral rather than the apical surface of LE cells. Attached embryos appear to gain rapid access to this highly adhesive lateral membrane domain.

Adhesiveness↗

Prevention of multiple pregnancy following IVF in Spain.

Since the development of assisted reproduction techniques most countries have witnessed increased rates of multiple pregnancy. Despite the guidelines proposed by various scientific societies these rates continue to be abnormally high. In Spain, as in other Mediterranean countries, a greater number of embryos are transferred than in northern and central European countries and the incidence of multiple pregnancies is greater in comparison. Effective strategies must be established to prevent multiple pregnancy without reducing overall pregnancy rates. In the authors' institute, taking into account the authors' experience, the relevant literature, and despite the limitation of retrospective studies, it is recommended that a maximum of two embryos are transferred in young women with good quality embryos at the time of transfer. The transfer of three embryos is only recommended in women >or=38 years who have one or no good quality embryos available at the time of transfer. The responsibility for preventing multiple pregnancy lies with health professionals, who must be aware of the risks involved in twin and triplet pregnancy. Couples must be provided with objective information before starting an IVF cycle. Professional societies should highlight the problem and make suitable recommendations.

Adult↗

Diagnostics in assisted human reproduction.

The World Health Organization (WHO) estimates that 50-80 x 10(6) couples in the world are infertile, i.e. 7-15% of all couples of reproductive age (15-45 years old). The term infertility refers to couples who have been unable to conceive children for at least 1 year of regular unprotected intercourse. Although the frequency and origin of infertility varies, approximately 40-60% of the aetiology of infertility in the population studied is due to female causes. The introduction of assisted reproduction techniques, which offer couples the best opportunities for pregnancy, has opened a vast field of knowledge in reproductive biology. The medical history must be taken meticulously, with enough time to obtain the largest possible amount of relevant information. It is recommended that both partners be included in the initial work-up. In recent decades, technological progress has been favoured with the development of a great variety of diagnostic tests. Moreover, a very valuable tool helping to solve this problem has gained strength and is available to everybody - so-called 'evidence-based medicine'. There are three categories in infertility diagnostic tests. The first category includes tests showing a well-established correlation with pregnancy, e.g. semen analysis, tubal patency with hysterosalpingography or laparoscopy, and ovulation detection. The second category includes patients whose results are not consistently related to pregnancy. Here, these tests include sperm penetration assay in the zona-free hamster oocyte, post-coital tests, sperm penetration into cervical mucus, and tests to detect anti-sperm antibodies. The third group includes patients whose tests are not correlated with pregnancy. The relevant tests include endometrial biopsy, the presence of varicocele, and Chlamydia detection tests. Tests for infertility are often expensive. Since these patients are usually anxious and eager and will do almost anything to have a child, care must be taken to avoid exploitation of their hopes with unnecessary procedures. This text now reviews and updates the procedures used to investigate infertility.

Acrosome Reaction↗

Why and how should multiple pregnancies be prevented in assisted reproduction treatment programmes?

Although most professional societies have issued guidelines to diminish the number of embryos to be transferred during assisted reproductive techniques, the incidence of multiple pregnancies remains unacceptably high. The burden of morbidity and mortality seems to increase substantially with each fetus in a multiple gestation. As a result, there has been growing debate on the need to prevent multiple pregnancies. The infertility specialists who can solve the infertility problem are usually shielded from the complications of multiple pregnancies. If they were involved in the delivery and, more particularly in the care of multiple pregnancies (both financially and socially), their attitude would probably change. IVF centres should gradually reduce the mean number of embryos per transfer in terms of the cost:benefit ratio. A further reduction to one single embryo per transfer in good cases would be similarly acceptable. Laboratory expertise is of vital importance, especially in terms of embryo culture, embryo selection, and freezing and thawing techniques in embryo transfer programmes for reducing the number of transferred embryos.

Cost-Benefit Analysis↗

Implications of reproductive technologies for birth and developmental outcomes: imprinting defects and beyond.

Assisted reproductive techniques (ARTs) have been widely used over the past two decades to help infertile couples conceive. Recent studies on the ART-conceived population have raised concern about the possible risks of these techniques, in particular with regard to increased incidence of growth and developmental disorders. Some of these effects might be linked to genomic imprinting defects, although current evidence does not allow definite conclusions to be drawn. This review summarises studies that have examined effects of gamete and embryo manipulations on imprinted genes, and discusses the evidence for and against effects of ARTs on offspring health, and in particular imprinting-related conditions.

Animals↗