A.A. Toledo

Human endometrial prostaglandin E2 binding sites and their profiles during the menstrual cycle and in pathologic states

Endometrial tissue from uteri of 35 nonpregnant, premenopausal women was assayed for prostaglandin E2 and F2 alpha binding site content as a function of the phase of the menstrual cycle and the pathologic state. For all specimens, tritium-labeled prostaglandin F2 alpha, binding was very low (less than 8 fmol/mg of protein) or undetectable regardless of the phase of the menstrual cycle or pathologic state or in the presence or absence of 10 mumol/L of indomethacin, a prostaglandin synthetase inhibitor. However, tritium-labeled prostaglandin E2 binding was detected in every specimen and was independent of the presence or absence of indomethacin. Binding of tritium-labeled prostaglandin E2, as determined by Scatchard analyses, was biphasic (dissociation constant approximately 1 nmol/L; dissociation constant for low-affinity sites approximately 10 nmol/L) for both proliferative and secretory endometrial tissue. However, the total number of prostaglandin E2 binding sites, determined from Scatchard or single-point analyses, was significantly higher (p less than 0.01) in proliferative endometrium compared to secretory endometrium. In addition, for endometrium from the proliferative phase of the menstrual cycle, the diagnosis of abnormal uterine bleeding was associated with higher (p less than 0.01) tritium-labeled prostaglandin E2 binding than diagnosis of dysmenorrhea, stress urinary incontinence and uterine prolapse, or pelvic inflammatory disease. Endometrial specimens with the last four diagnoses did not differ significantly (p greater than 0.1) from each other.

Electroejaculation in combination with in vitro fertilization and gamete micromanipulation for treatment of anejaculatory male infertility

Objective: Failure to ejaculate may be overcome by use of electroejaculation. However, such semen samples are often unsuitable for therapies like intrauterine insemination. The combination of electroejaculation with in vitro fertilization, including gamete micromanipulation, should improve chances of fertilization and pregnancy in such cases. Study Design: Within a private infertility clinic electroejaculation in combination with intrauterine insemination was carried out in 18 cycles (10 couples). Four couples went on to receive therapy by electroejaculation plus in vitro fertilization, along with six other couples (15 cycles total) with semen too poor for intrauterine insemination. Results: One term pregnancy arose in the electroejaculation-intrauterine insemination group, and one term pregnancy plus one continuing pregnancy arose from two couples (three cycles) who underwent in vitro fertilization with conventional insemination after electroejaculation. Six couples (nine cycles) had embryos arising only from gamete micromanipulation transferred, and this yielded two term pregnancies, one spontaneous abortion, and a biochemical pregnancy. Two couples (three cycles) failed to achieve fertilization even with micromanipulation; however, donor-inseminated eggs gave rise to two term pregnancies and one continuing pregnancy in these patients. Conclusions: This report confirms the feasibility of in vitro fertilization in conjunction with electroejaculation and extends the therapy to incorporate gamete micromanipulation.

Effect of coculture on subsequent survival and implantation of cryopreserved human embryos

PurposeThis retrospective analysis was designed to assess the performance of human embryos following cryopreservation based on whether they were originally developed in standard culture medium (65 cycles, 223 embryos) or cocultured on partial monolayers of bovine oviductal epithelial cells (63 cycles, 198 embryos). Embryo cryosurvival and implantation were compared between the study group and the contemporaneously matched controls.ResultsDuring a 2-year period when no factors of the cryopreservation program were altered, 63 transfers of 159 surviving thawed control cleavage-stage embryos (71.3% survival) that were 54% intact gave rise to 11 viable pregnancies (17.5%/ET), to yield an implantation rate of 6.9% per embryo. Sixty-three transfers of 147 thawed cocultured embryos (74.2% survival) that were 61% intact gave rise to 17 viable pregnancies (27%/ET), which gave an implantation rate of 13.6% per embryo that was significantly higher than the control group (P< 0.05).ConclusionCoculture of embryos prior to cryopreservation does not appear to improve cryosurvival; however, it does improve implantation postthaw compared with embryos following standard culture prior to cryopreservation.

Poor implantation of cryopreserved reinsemination-fertilized human embryos

OBJECTIVE To investigate whether a poor rate of implantation after in vitro fertilization (IVF) was due to poor embryonic/endometrial synchrony during the original IVF cycle, we have cryopreserved reinseminated-fertilized embryos for later more synchronous replacement after thawing. The chance of implantation of fresh reinseminated fertilized human oocytes is approximately one tenth that of timely fertilized embryos. STUDY DESIGN AND DATA: Retrospective study of 35 original oocyte collections in which initial normal fertilization was 47.3% (129/273 oocytes), with 49.6% fertilization (67/135) upon reinsemination. Of these, 70 initially fertilized and 67 reinsemination-fertilized embryos were cryopreserved, and 50 initially fertilized and all 67 reinsemination-fertilized embryos were subsequently thawed with 72% and 63% cryosurvival, respectively, (not significant). SETTING Private infertility clinic. RESULTS In 11 cycles, 23 thawed initially fertilized embryos (group A) were replaced with a 21.7% implantation rate per embryo; in 10 cycles, 13 initially fertilized and 12 reinsemination-fertilized embryos (group B) were replaced together with an 8% implantation rate; finally, in 16 cycles, 30 reinsemination-fertilized embryos (group C) were replaced with a 3.3% implantation rate (group A versus group C: P = 0.076). Comparison of clinical pregnancies between these groups was significantly different (6/11 versus 1/16; P = 0.0427). CONCLUSION Reinsemination-fertilized embryos survive freezing as well as initially fertilized embryos, but better embryonic/endometrial synchrony does not improve chances of their implantation.

Simultaneous Detection of Chromosomes X, Y, 13, 18, and 21 by Fluorescence In Situ Hybridization in Blastomeres Obtained from Preimplantation Embryos

Purpose.Simultaneous fluorescence in situ hybridization (FISH) was used in a preimplantation genetic diagnosis program to determine which embryos were normal for chromosomes X, Y, 13, 18, and 21.Methods:Single blastomeres were disrupted and attached to glass slides using acetic acid and ethanol. Using a ratio mixture of chromosome enumeration DNA probes in combination with locus-specific identifier DNA probes, FISH was performed for the identification of chromosomes X, Y, 13, 18, and 21.Results:Fourteen couples enrolled in IVF produced 134 embryos for biopsy. Blastomeres subjected to five-color FISH revealed that 22% of embryos were normal for chromosomes X, Y, 13, 18, and 21. In addition, 52% were abnormal and no results could be detected for 25%. Twelve couples underwent embryo transfer, two couples did not receive embryos due to lack of any normal embryos, and three couples became pregnant.Conclusions:The simultaneous detection of five-color FISH is a feasible method to detect aneuploidy in preimplantation embryos from women of advanced maternal age.

Exposure of preimplantation embryos to platelet-activating factor increases birth rate.

AbstractProblem: Platelet-activating factor (PAF) plays a significant role in fertility. Preimplantation stage embryos produce PAF (ePAF) which is required for development. PAFs mechanism of action is receptor-mediated and its presence has been reported in the developing mouse and human embryo. Exposure of preimplantation stage mouse embryos results in higher implantation rates. However, the effect of such treatment on live-birth rates and birth weights has not been reported. Therefore, the objective the study was to determine the effect of exposing preimplantation mouse embryos to PAF on subsequent birth rate and weight. Design: Two-cell stage preimplantation stage mouse embryos exposed to PAF (10−7 M) for 15 min prior to intraoviductal transfer. Methods: Preimplantation stage embryos were recovered from eCG/hCG primed BDF1 female mice. Embryos were exposed to synthetic PAF (10−7 M) for 15 min. PAF-treated embryos were transferred to the oviducts of pseudopregnant female CD-1 female mice. Superovulated and cultured BDF1 embryos not treated with PAF served as in vitro controls and naturally ovulated embryos with no collection/culture served as in vivo controls. Embryos were permitted to develop to term (18–21 days). The number of pups born per litter and litter weights subsequently were recorded. Results: A total of 160 BDF1 mouse embryos were collected, treated, and transferred (20 per CD-1 recipient) as described. There was a significant (P<0.05) increase in the number of pups born to the PAF treatment group (56/80; 70%) as compared to the control group (44/80; 55%). There was also a significant difference (P<0.05) in litter birth weights between the PAF (1.31 g/litter) and controls groups (1.25 g/litter). There was a significant difference (P<0.05) in birth weights between the PAF treatment group and the in vivo group (1.51 g/litter). There was a significant difference in birth weights between the in vitro control and in vivo groups (1.51 g/litter). There were no observational malformaties to pups born in any group. Conclusions: Brief exposure of preimplantation stage embryos to PAF will result in a significant increase of delivery rates (pups/litter) as well as birth weights. However, the increase of birth weight was significantly below that found naturally. Additional studies are warranted to elucidate the mechanism of PAFs action in the preimplantation stage embryo and subsequent uterine development.

Treatment of male infertility and idiopathic failure to fertilize in vitro with under zona insemination and direct egg injection

Objective: Failure to fertilize eggs in vitro may be countered by micromanipulation of gametes to place selected spermatozoa underneath the zona pellucida of the egg or directly into the egg, thereby improving chances of fertilization and production of viable embryos. Analysis of our clinical data for assisted fertilization was undertaken to assess those factors of relevance in this therapy, and a description of our procedures are given. Study Design: Retrospective analysis of 85 cycles (73 couples) of in vitro fertilization and embryo transfer performed at a private infertility clinic, in which micromanipulation for assisted fertilization was used to overcome either severe male factor infertility or idiopathic failure to fertilize, was performed. Results: In 60 cycles where only embryos from under zona insemination were available for uterine transfer, 15 singleton and two twin pregnancies occurred (28.3% viable pregnancy rate per transfer, 14.1% embryonic implantation). In 14 of these cycles embryos arose only after repeated under zona insemination adding more spermatozoa; this accounted for four of the singleton and one of the twin pregnancies (38.5% pregnancy rate, 22.2% embryonic implantation). No embryos arose from partial zona dissection performed in five cycles on sibling eggs. However, in 16 cycles conventional insemination yielded fertilization in six cycles, and mixed transfer of these embryos and sibling embryos from under zona insemination gave rise to one pregnancy from four transfers (pregnancy rate 25%, embryonic implantation 7.1%). Likewise, in nine cycles donor spermatozoa yielded fertilization in eight cycles, and mixed transfer with sibling embryos fertilized by under zona insemination with partners spermatozoa gave rise to two pregnancies from five transfers (pregnancy rate 40%, embryonic implantation 15.8%). Fertilization and pregnancy rates did not differ whether couples suffered either from male factor infertility or from previous idiopathic fertilization failure. Direct egg injection of a single spermatozoon into 105 eggs gave an 88.6% egg survival and 32.3% fertilization. Mixed transfers with sibling embryos from conventional and under zona insemination yielded one triplet, one twin, and three singleton pregnancies. Conclusions: Overall, a 24.7% (21/85) viable pregnancy rate per cycle initiated occurred when only embryos from assisted fertilization were available. This strongly indicates that assisted fertilization made a real contribution in cases where either insufficient spermatozoa were available for conventional insemination or in cases where previous fertilization failure had arisen. The wide range of seminal parameters were found to be unhelpful in defining chances of success with assisted fertilization.

The prevention of adhesion formation by nonsteroidal antiinflammatory drugs: an animal study comparing ibuprofen and indomethacin

The efficacy of two nonsteroidal antiinflammatory drugs, ibuprofen and indomethacin, in the prevention of postoperative adhesions was examined. Thirty-three guinea pigs were randomly divided into three groups: a control group (n = 11), an ibuprofen group (n = 11), and an indomethacin group (n = 11). All of the animals received standardized injuries, and adhesions were graded 4 weeks later. Both treatment groups were found to have significantly fewer (P less than 0.01) adhesions when compared with the control group with no difference among the two treatment groups. It is concluded that ibuprofen and indomethacin are equally effective in reducing postoperative adhesions.

Fertilization Potential of Human Sperm Is Correlated with Endogenous Platelet-Activating Factor Content

AbstractPurpose: Platelet-activating factor (PAF) is a potent signaling phospholipid that is found in mammalian sperm and has a positive correlation with fertility. Whereas PAF is present in human sperm, there are no relational reports on its content and the cells fertilization potential. Therefore, the study objective was to determine if PAF content in capacitated-induced sperm is related to fertilization potential as determined by the sperm penetration assay (SPA). Methods: Endogenous sperm lipids were measured for PAF content by a specific radioimmunoassay following insemination of zona pellucida-free hamster ova. Data were analyzed by regression analysis and Students t test. Results: Regression analysis revealed a positive and significant relation (R2 = 0.806; P < 0.05) between PAF content in human sperm and SPA outcome (pass: ≥5.0; fail: <5.0, penetrations/ova). Patients that passed (22.61 ± 5.21 picomoles/106) the SPA had significantly (P < 0.01) higher PAF levels in their sperm than patients that failed (12.91 ± 1.76 picomoles/106 cells) the test. Conclusions: PAF content in capacitated-induced sperm has a significant and positive relationship with fertilization potential. Fertilization potential may be predicted by measuring PAF levels in capacitation-induced human sperm. Determining PAF content in capacitated human sperm may be a beneficial diagnostic tool for the infertility specialist.

Evaluation of the first one hundred and thirty one live births following oocyte cryopreservation from a single ivf program utilizing a standardized vitrification technique

OBJECTIVE: Oocyte cryopreservation by vitrification is an efficient procedure, however, limited data is available addressing the safety of this technology. For this reason we analysed live birth outcomes obtained after egg freezing in a comparable patient population. DESIGN: Retrospective, matched control study. MATERIALS AND METHODS: From December 2006 to December 2009, live birth outcomes from 90 deliveries following vitrified donation cycles & from 112 fresh egg donation cycles were tabulated and analysed. Cryopreservation of oocytes was performed by minimum volume vitrification. Oocytes were fertilized by 40 hrs after HCG adminisration (fresh egg donation) or 2-3 hrs after oocyte warming (cryo egg donation) Results were analysed by the One-way Anova or the Fisher’s exact tests, as appropriate (P<0.05). RESULTS: Parameters of live birth outcomes are reported in Table 1.