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Results for “In Vitro Fertilization (IVF)”

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PubMed2006

In vitro fertilization of in vitro matured canine oocytes using frozen-thawed dog semen.

Experiments were conducted to evaluate in vitro fertilization (IVF) of in vitro matured (IVM) bitch oocytes using dog spermatozoa frozen in three different extenders. Sperm-rich fraction from eight ejaculates of five dogs was frozen in each one of three egg yolk Tris extenders with additional: (A) 1.4 g citric acid and 0.8 g glucose; (B) 0.7 g citric acid and 3.5 g glucose; or (C) 1.4 g citric acid and 0.8 g fructose (all with 5% glycerol in 100 mL milliQ water). Thawed sperm were co-incubated with IVM bitch oocytes for 6 h. Oocytes were fixed and evaluated under an epifluorescence microscope; penetrated oocytes were defined as those having sperm heads in the perivitelline space or in the oocyte cytoplasm. Higher penetration rates (P < 0.05) were obtained in oocytes cultured with spermatozoa frozen in extenders B and C than those frozen in extender A (33.1, 34.2 and 26.4%, respectively).

Animals↗

PubMed2004

Regulation of ribosomal RNA gene expression in porcine oocytes.

In vitro production (IVP) of porcine embryos including in vitro maturation (IVM) of oocytes followed by in vitro fertilization (IVF) and in vitro culture (IVC) of the resultant embryos may result in live offspring, but it is still associated with great inefficiencies probably due to incomplete cytoplasmic maturation of the oocytes in vitro. Therefore, fundamental knowledge on the regulation of transcription during the oocyte growth phase when the messengers and protein synthetic machinery necessary for oocyte developmental competence are formed, is of great importance. In mammals, synthesis of RNA, up to 60-70% of which is ribosomal (rRNA), increases during oocyte growth and reaches a peak at the beginning of follicular antrum formation. In oocytes at the end of the growth phase, acquisition of full meiotic competence coincides with a markedly decreased rRNA transcriptional activity in the gametes. Our recent studies on the porcine oocyte growth phase have revealed a deeper molecular and biological insight into the complex regulation of rRNA transcription at different stages of follicular development. The data indicate that the so-called pocket protein, p130, is involved in the down-regulation of rRNA transcription at the end of the oocyte growth phase through an inhibition of the action of upstream binding factor (UBF). The latter protein is necessary for the function of RNA polymerase I (RNA Pol I), which is the actual enzyme driving rRNA gene transcription. Moreover, rRNA transcription also appears to be down-regulated by a decrease in the expression of mRNA encoding PAF53, an RNA Pol I-associated factor also required for the polymerase to exert its action. At the ultrastructural level, these molecular changes are paralleled by marginalization of the fibrillar centres of the oocyte nucleolus followed by compaction of the nucleolus into an inactive sphere of fibrils.

Animals↗

PubMed1999

Morphologic evaluation and actin filament distribution in porcine embryos produced in vitro and in vivo.

Porcine embryos produced in vitro have a small number of cells and low viability. The present study was conducted to examine the morphological characteristics and the relationship between actin filament organization and morphology of porcine embryos produced in vitro and in vivo. In vitro-derived embryos were produced by in vitro maturation, in vitro fertilization (IVF), and in vitro development. In vivo-derived embryos were collected from inseminated gilts on Days 2-6 after estrus. In experiment 1, in vitro-derived embryos (</= 8-cell stage) collected 12-48 h after IVF were separately fixed, stained by orcein, and examined under phase contrast microscopy. It was found that 27% of 2-cell, 74% of 3-cell, 51% of 4-cell, and 74% of 5- to 8-cell-stage embryos were abnormal in morphology. Morphological anomalies included fragmentation (no nucleus in one or more than one blastomere) and/or binucleation (two nuclei in one or more than one blastomere). In experiment 2, actin filament distribution of the embryos at 2-cell to blastocyst stages that were produced in vivo and in vitro were stained by rhodamine-phalloidin and examined by confocal microscopy. Actin filaments were distributed in all in vivo-derived embryos at the cell cortex, and at the joints of cells and perinucleus in 2- to 8-cell-stage embryos and in some cells of morulae and blastocysts. Actin filaments were also distributed in the cortex and at the joints of cells of all in vitro-produced embryos. However, only 20% of in vitro-produced embryos at 2- to 8-cell stages had perinuclear actin filaments in all blastomeres. Most in vitro-produced embryos had fewer perinuclear actin filaments or did not have perinuclear actin filaments in some blastomeres. Fragmentation and binucleate blastomeres were not observed in in vivo-derived embryos. In vivo-derived Day 5 (136.5 +/- 60.4 nuclei per blastocyst) and Day 6 (164.5 +/- 51.9 nuclei per blastocyst) blastocysts had significantly (p < 0.001) more cells than in vitro-produced Day 6 blastocysts (37. 3 +/- 11.7 nuclei per blastocyst). In experiment 3, when cytochalasin D, an inhibitor of microfilament polymerization, was included in the culture medium, it prevented 2- to 4-cell-stage embryos from developing to the blastocyst stage. These results indicate that abnormal actin filament distribution is one possible reason for abnormal embryo cleavage and small cell numbers in pig embryos produced in vitro. Culture conditions that mediate normal actin filament distribution may result in an improvement in embryo quality.

Actins↗

PubMed1988

The influence of stimulation regimes and luteal phase support on the outcome of IVF.

Successful in-vitro fertilization (IVF) requires the generation of a number of healthy embryos and adequate priming and maintenance of a receptive endometrium. Both of these factors are influenced by ovarian stimulation regimes and are responsive to endocrine manipulation. Detailed analysis of follicular phase patterns of hormone secretion have established an inverse relationship between luteinizing hormone (LH) and egg/embryo quality which is not discernible on microscopic examination. The advent and extensive use of LHRH agonists in ovarian stimulation has gone some way towards optimizing embryo fitness by suppressing LH secretion and this has exposed the endometrial contribution to a failure of implantation to closer examination. The supraphysiological production of oestradiol and progesterone from the large numbers of follicles developed during ovarian stimulation cannot be without effect on the endometrium. Data presented in this brief review show how our understanding of ovarian stimulation and its influence on the outcomes of IVF has advanced and highlights the opportunities for manipulation of the endometrium to encourage implantation.

Female↗

PubMed2003

Capacity of adult and prepubertal mouse oocytes to undergo embryo development in the presence of cysteamine.

The present study was carried out to study de novo glutathione (GSH) synthesis and to evaluate the effect of stimulating GSH synthesis during in vitro maturation (IVM) of adult and prepubertal mouse oocytes on the embryo developmental rate. Adult (8 weeks old) and prepubertal mice (24-26 days old) were primed with 5 IU of PMSG and oocytes were retrieved from the ovary 48 hr later for IVM. After IVM (18 hr) Cumulus oocyte complexes (COC) were in vitro fertilized (IVF) and in vitro culture (IVC) in order to observe embryo development. The IVM medium was supplemented with: 0, 25, 50, 100, or 200 microM of cysteamine. To study the novo GSH synthesis, 5 mM BSO was added during IVM of adult or prepubertal oocyte. Developmental rates up to blastocyst were recorded for each group. Experiments also included a group of ovulated oocytes (in vivo matured) after priming with PMSG and HCG. After IVM of adult mice oocytes, an improvement was observed on embryo development in all the supplemented groups when compared with the untreated group (P < 0.05). No differences were observed in blastocyst rate among IVM oocytes with cysteamine and ovulated oocytes. Prepubertal IVM mouse oocytes had a lower cleavage rate compared with ovulated oocytes (P < 0.05). Cysteamine failed to improve prepubertal oocytes developmental rates (P > 0,05). 2-cell embryos, coming from IVM prepubertal oocytes and ovulated oocytes had the same preimplantation developmental rate up to the blastocyst stage. In prepubertal, and adult oocytes an inhibition of embryo development was observed when buthionine sulfoximide (BSO), a specific inhibitor of the gamma-glutamylcysteine synthetase, was added during oocyte maturation (P < 0.01). In conclusion, an improvement in mouse embryo development was observed when cysteamine was added to the IVM medium of adult mice oocytes. In prepubertal oocytes cysteamine addition during oocyte maturation failed to improve embryo developmental rates. The presence of BSO lowered or completely blocked blastocyst development. This proves that, de novo GSH synthesis during oocyte maturation of adult and prepubertal oocytes undoubtedly plays an important role in embryo development. The improvement on oocyte competence observed in adult mice oocytes is probably related to intracellular GSH synthesis stimulated by cysteamine. Nevertheless the reason why cysteamine failed to improve prepubertal oocytes competence remains as an open question.

Animals↗

PubMed1995

A rapid method for mRNA detection in single-cell biopsies from preimplantation-stage bovine embryos.

Major questions concerning the control of development and gene expression at the cellular level are still unanswered. Nowhere is this more evident than during the earliest stages of development and embryogenesis. This study describes the detection of specific gene transcripts in single cells derived from bovine embryos. Following in vitro fertilization (IVF) and in vitro culture (IVC) of bovine embryos, small groups of cells and even single blastomeres from 32 to 64-cell embryos were micromanipulated into individual tubes for analysis of cytoplasmic RNAs. Reverse transcriptase-PCR was applied to cell lysates for the amplification of beta-actin mRNA transcripts. Primers were designed to flank an intron expected to be present within genomic DNA sequences, thus allowing for simple differentiation between DNA- and RNA-derived amplification products. Using a 50-cycle amplification profile, a 260 bp band could be seen as a PCR product derived from a single blastomere following electrophoresis in an ethidium bromide-stained agarose gel. The identity of the band was verified by DNA sequence determination and diagnostic restriction digestion. Lysates derived from single blastomeres in this way have been used for simultaneously phenotyping multiple RNA products. This capability allows the spatial analysis of gene expression and development within embryos from the earliest stages of cellular differentiation.

Journal Article↗

PubMed1994

Effect of recovery method on yield of bovine oocytes per ovary and their developmental competence after maturation, fertilization and culture in vitro.

An important aim of an oocyte recovery method is to maximize the number of oocytes per ovary which can be employed for in vitro maturation (IVM), in vitro fertilization (IVF) and in vitro culture (IVC). In this study, primary bovine oocytes were collected by 2 methods: aspiration of visible follicles (2 to 6mm in diameter) or surface dissection in which the ovary surface is finely dissected. The oocytes were classified on the basis of cumulus cover and cytoplasmic appearance. The total number of oocytes and the yield of good-quality oocytes recovered per ovary by surface dissection and aspiration were 44.2 and 13.9 and 13.5 and 4.6 (P<0.05), respectively. When a sample group of selected oocytes recovered by each method was measured, no significant difference was found in the mean diameter (144.11m vs 142.54m). A representative sample of good-quality oocytes recovered by each method was put through the IVM/IVF/IVC procedure: no significant difference in cleavage rate, cleavage index or blastocyst yield was found. However, when the blastocyst yield was compared on a per ovary basis, a significant difference was observed in favor of surface dissection (3.30+/-0.46 vs 0.96+/-0.16;P<0.05). When unselected oocytes recovered by surface dissection of the ovaries were put through the standard embryo production system, an average of 15.4 blastocysts per dam was obtained.

Journal Article↗

PubMed1995

[MESA (microsurgical epididymal sperm aspiration) and IVF (in vitro fertilization). A therapy concept in treatment of male infertility].

Microsurgical epididymal sperm aspiration (MESA), or sperm microaspiration retrieval technique (SMART), in conjunction with in vitro fertilization is a successful therapy in male infertility. From November 1991 to March 1994 a total of 29 attempts at MESA with subsequent IVF were made. Of 48 aspirations, 37 were successful and 13 attempts at IVF were possible, 6 of which were successful with 10 subsequent embryo transfers. In all, 3 pregnancies were achieved and 1 boy was born. In conclusion, microsurgical spermaspiration in conjunction with in vitro fertilization is a way of treating male infertility with a chance of achieving paternity with the partner's own sperm; the chances are probably better with intracytoplasmatic sperm injection.

Embryo Transfer↗

PubMed1996

Cumulative pregnancy rate following in-vitro fertilization: the significance of age and infertility aetiology.

During the years 1984-1992, 951 couples completed 2252 in-vitro fertilization (IVF) treatment cycles at the In-Vitro Fertilization Unit of The Chaim Sheba Medical Centre. This study was conducted to evaluate the success of IVF using the cumulative pregnancy rate (CPR), with special emphasis on the optimal number of treatment cycles, the age of the patients and female infertility factors. It was found that the CPR showed a constant rise during the six initial IVF treatments (56% CPR), and plateaued in the subsequent three cycles (63% CPR). Various female infertility factors did not influence this rate. Women > or = 40 years of age have a significantly lower CPR. Thus, it was concluded that the CPR in IVF declined after the sixth initial treatment cycle, and in women > or = 40 years of age. The infertility factor did not significantly influence CPR.

Adult↗

PubMed2000

TransgenICSI reviewed: foreign DNA transmission by intracytoplasmic sperm injection in rhesus monkey.

This brief review considers the status of transgenesis by intracytoplasmic sperm injection (ICSI) with nonhuman primates. GFP expressing rhesus macaques embryos (mean = 34.6%; N = 81) were produced by ICSI using rhodamine-tagged DNA encoding the green fluorescence protein (GFP) gene bound on sperm. Rhodamine signal was lost at the egg surface during in vitro fertilization (IVF) but could be traced by dynamic imaging during ICSI within the egg cytoplasm. GFP gene was expressed as early as the 4-cell stage in ICSI embryos but not in embryos produced by in vitro fertilization (IVF). The percentage of GFP expressing blastomeres increased during embryogenesis to the blastocyst stage. Three offspring resulted from seven embryo transfers-a set of anatomically normal twins (a male and a female) stillborn 35 days premature, and a healthy male born at term. Although transgene was not detected in the offspring, the successful production of live primates using DNA bound sperm by ICSI suggests an alternative route to creating transgenic animals. It also raises concern regarding transmission of infectious material during ICSI.

Animals↗

PubMed2006

Comparative study of the efficacy and tolerability of two vaginal progesterone formulations, Crinone 8% gel and Utrogestan capsules, used for luteal support.

OBJECTIVE: To compare the efficacy and tolerability of two different types of vaginal progesterone (P), Crinone 8% gel (Fleet Laboratories Ltd., Watford, United Kingdom) and Utrogestan capsules (Laboratories Besins International, Paris, France), used for luteal support after in vitro fertilization (IVF) cycles. DESIGN: Cohort study. SETTING: In Vitro Fertilization Polyclinic, Zagreb, Croatia. PATIENTS: A total of 285 women aged < or =37 years undergoing IVF-embryo transfer treatment. INTERVENTIONS: Patients were treated with either Crinone 8% vaginal P gel (90 mg) administered daily, or Utrogestan vaginal capsules (2 x 100 mg) administered three times daily. Progesterone was administered from the day of oocyte retrieval (day 0) to menses or, in a case of pregnancy, until week 12. MAIN OUTCOME MEASURE: Clinical pregnancy rate. The tolerability and acceptability of both preparations were determined by a questionnaire given to patients. RESULTS: The similar rates of clinical pregnancies (33.1% vs. 30.9%) [corrected] were obtained by using either Crinone 8% vaginal P gel or Utrogestan vaginal capsules. Overall tolerability and acceptability were significantly better in the Crinone group than in the Utrogestan group. CONCLUSIONS: The efficacy of the two vaginal P formulations was nearly the same, but the tolerability and acceptability of Crinone 8% gel were superior, in the opinion of patients.

Administration, Intravaginal↗

PubMed2006

Guidelines for the number of embryos to transfer following in vitro fertilization.

OBJECTIVE: To review the effect of the number of embryos transferred on the outcome of in vitro fertilization (IVF), to provide guidelines on the number of embryos to transfer in IVF-embryo transfer (ET) in order to optimize healthy live births and minimize multiple pregnancies. OPTIONS: Rates of live birth, clinical pregnancy, and multiple pregnancy or birth by number of embryos transferred are compared. OUTCOMES: Clinical pregnancy, multiple pregnancy, and live birth rates. EVIDENCE: The Cochrane Library and MEDLINE were searched for English language articles from 1990 to April 2006. Search terms included embryo transfer (ET), assisted reproduction, in vitro fertilization (IVF), ntracytoplasmic sperm injection (ICSI), multiple pregnancy, and multiple gestation. Additional references were identified through hand searches of bibliographies of identified articles. VALUES: Available evidence was reviewed by the Reproductive Endocrinology and Infertility Committee and the Maternal-Fetal Medicine Committee of the Society of Obstetricians and Gynaecologists of Canada and the Board of the Canadian Fertility and Andrology Society, and was qualified using the Evaluation of Evidence Guidelines developed by the Canadian Task Force on the Periodic Health Exam. BENEFITS, HARMS, AND COSTS: This guideline is intended to minimize the occurrence of multifetal gestation, particularly high-order multiples (HOM), while maintaining acceptable overall pregnancy and live birth rates following IVF-ET.

Adult↗

PubMed1999

Maternal serum alpha-fetoprotein and human chorionic gonadotrophin in pregnancies conceived after intracytoplasmic sperm injection and conventional in-vitro fertilization.

Data in the Caucasian population suggest that maternal serum alpha-fetoprotein (AFP) and unconjugated oestriol concentrations are reduced and human chorionic gonadotrophin (HCG) concentrations are elevated in pregnancies conceived after in-vitro fertilization (IVF), leading to a higher than expected Down's syndrome screen-positive rate. There are no previous reports on the serum marker values in pregnancies conceived after intracytoplasmic injection (ICSI). Between 1996 and 1998, we measured maternal serum total HCG and AFP concentrations between 15 and 20 weeks gestation in 42 in-vitro fertilization (IVF) pregnancies and 23 ICSI pregnancies with known normal outcome. The results were compared with that of 2799 naturally occurring singleton pregnancies who were known to have a normal outcome. Median AFP multiple of the median (MOM) in ICSI pregnancies was significantly reduced to 0.76 compared with both that of the controls and that of the IVF pregnancies. For the IVF pregnancies, median HCG MOM was elevated to 1.15, and median AFP MOM was reduced to 0.88 compared with the controls, but these differences were not statistically significant. In both the IVF and ICSI pregnancies the changes might result in a falsely high Down's syndrome risk. In particular, the reduced AFP concentration in ICSI pregnancies was substantial. If this preliminary finding is substantiated by other series, the appropriate adjustment needs to be made to allow for valid interpretation of the screen result and to avoid an unnecessarily high false positive rate.

Adult↗

PubMed1998

Studies on the origin of redox enzymes in seminal plasma and their relationship with results of in-vitro fertilization.

Glutathione (GSH), GSH peroxidase (GPX), GSH reductase (GRD), superoxide dismutase (SOD) and catalase-like enzyme activity were quantified in seminal plasma from normozoospermic patients, men with known distal ductal occlusion, proven fathers and male partners of couples receiving in-vitro fertilization (IVF) treatment for both male and female causes. Glutathione was non-detectable (< 2.5 microM) in seminal plasma. None of the enzyme activities per unit volume were lower in semen from vasectomized men, suggesting that they did not originate substantially from the testis or epididymis. The strongest relationships between enzyme activities and accessory gland markers were between zinc and GRD (r = 0.678), SOD (r = 0.602) and GPX (r = 0.548), suggesting a largely prostatic origin of these enzymes. Only weak relationships between accessory gland markers and catalase-like activity suggested a multi-glandular source of this enzyme. There was no relationship between the activity of any of the enzymes in the IVF patients with their fertilization rates in vitro or the establishment of pregnancy after IVF. Nor was there any correlation of enzyme activity with the morphology and percentage of motile spermatozoa in semen or with the percentage motility of spermatozoa immediately after swim-up or after overnight incubation. These findings suggest that the protective enzymes in the seminal plasma are contributed largely by the prostate and little by the epididymis, and that in most cases of IVF, they have no major influence on the outcome.

Adult↗

PubMed1993

In vitro fertilization: mandatory IVF coverage in health insurance.

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Canada↗

PubMed1990

Analysis of immunosuppressive molecules associated with murine in vitro fertilized embryos.

Supernatants from mouse in vitro fertilized (IVF) oocyte cultures may suppress in vitro lymphocyte proliferation stimulated by concanavalin A (Sigma, St. Louis, MO). Supernatants conditioned by incubation with mouse epididymal sperm alone were even more inhibitory. Thin-layer chromatography suggested the polyamines spermine in sperm and spermine plus spermidine in IVF embryo supernatants were responsible. Putrescine was not suppressive. In vitro fertilized oocytes from old CBA/J-strain mice (greater than 20 weeks) that suffer age onset infertility lacked suppression and manifest cleavage arrest that could be partially reversed by adding spermine to the cultures. The failure of IVF oocytes to produce adequate quantities of polyamines could lead to failure of implantation due to division arrest. A possible in vivo role of polyamines as immunosuppressor factors is discussed.

Animals↗

PubMedDate not supplied

Culture and selection of viable blastocysts: a feasible proposition for human IVF?

In human in-vitro fertilization (IVF) embryos are routinely transferred to the uterus on day 2 or day 3 of development. Resultant implantation and pregnancy rates are disappointingly low, with only approximately 10% of embryos transferred leading to a live birth. The ability to culture embryos to the blastocyst stage should help to resolve this problem by synchronizing the embryo with the female reproductive tract, and by identifying those embryos with little developmental potential. Co-culture has offered a possible means of producing blastocysts capable of high implantation rates. However, recent developments in the field of embryo physiology and metabolism have led to the formulation of new sequential serum-free culture media capable of supporting the development of viable blastocysts in several mammalian species, including the human. It is therefore proposed that blastocyst transfer should be considered for routine use in human IVF. The high viability of blastocysts cultured in the appropriate sequential media means that fewer embryos are required for transfer to achieve a pregnancy, culminating in fewer multiple births. Furthermore, the development of suitable non-invasive tests of embryo viability should further increase the overall success of human IVF by the ability to select before transfer those blastocysts most able to establish a pregnancy.

Amino Acids↗

PubMed2000

Increased pregnancy rates after IVF/ET with intravenous immunoglobulin treatment in women with elevated circulating C56+ cells.

Intravenous (IV) immunoglobulin (Ig) has been previously shown to increase pregnancy rates after previously failed in vitro fertilization (IVF) embryo (ET) attempts in women who are efficient embryo producers (fertilize at least 50% of oocytes retrieved and generate at least 3 embryos/cycle). Women experiencing implantation failure have a higher frequency of elevated percentage of circulating CD56+ (natural killer) cells (>12%) than fertile women (3-12%). To evaluate the effects of IVIg on pregnancy rates in women with elevated percentage of circulating CD56+ cells, 32 women who had previously failed IVF/ET (>12 embryos transferred without pregnancy) were studied. Pregnancy and live birth rates with and without IVIg were compared in the same woman. All 32 women had previously failed to conceive after at least 12 ET, were efficient embryo producers and had persistently elevated plasma concentrations of CD56+ cells. Each woman received IVIg 500mg/kg prior to ET. If serum hCG concentrations were positive for pregnancy, IVIg was continued at 500mg/kg/mo until 28 weeks gestation. Pregnancy rates with and without IVIg were 56% and 9% (P<0.0001). The rate of live birth was 38% with IVIg and 0% without IVIg (P<0.0001). IVIg enhances pregnancy and live birth rates in women with elevated circulating CD56+ cells who have a history of implantation failure. Despite technologic advances leading to enhancement of fertilization rates after in vitro fertilization (IVF) (1, 2) implantation rates after embryo transfer (ET) have not increased significantly (3) over the last 20 years (4). Implantation rates after IVF/ET are influenced by the quality of the embryos and receptivity of the endometrium (3-9). Endometrial receptivity involves both hormonal (10-13) and immunologic (14-29) factors. Among the immunologic factors that play a crucial role in successful implantation are natural killer (NK) cells (14-18). NK cells present within the decidua that express CD56(but lack CD 16) have been associated with successful implantation (14-18). A deficiency of decidual CD56+ CD16- cells (18) and an increase in circulating CD56+ cells (25, 26) have been observed in women experiencing implantation failure. Women experiencing implantation failure after IVF and multiple ET have been successfully treated with intravenous (IV) immunoglobulin (Ig) (27). IVIg reduces activation of NK cells and NK killing activity both in vitro (29) and in vivo (30-31). This reduction in activation of NK cells is essential for normal implantation to occur (14). To further define the role of IVIg for treatment of implantation failure, pregnancy and live birth rates were compared before and after IVIg treatment in women undergoing IVF/ET who had elevated levels of circulating CD56+ cells.

Adult↗