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Endometrial biopsy was performed in 27 infertile women participating in the IVF program. Their mean age was 31.8 years, 33% of the women being over 35 years old. The average duration of infertility was 6.9 years. The superovulation protocol consisted of hMG/hCG in 5 cases, of combined GnRH analog/gonadotropin therapy in 20 women, and 2 patients received combined contraceptive pill/gonadotropin treatment. Judging by hormonal profiles, follicular growth rate and number of oocytes retrieved, the response to stimulation was normal. The mean estradiol (E2) levels increased from 132.7 pg/ml on day -5 (SEM = 9.67) to 1272 pg/ml (SEM = 103.7) on the day of hCG administration and to 1813 pg/ml (SEM = 209.6) 1 day later. One day before the hCG application, the mean progesterone and LH levels were 1.34 ng/ml and 8.38 IU/ml, respectively. Only one patient had clinical hyperstimulation syndrome. Ova were harvested in all women, the mean number of oocytes being 7.7 (SEM = 0.83) per patient. In all 27 cases lack of fertilization or faulty ovum cleavage were observed. Thus, an endometrial biopsy (EB) was performed 72 h after oocytes retrieval. The mean estrogen and progesterone levels on the EB day were 610.9 pg/ml (SEM = 78.44) and 45.4 ng/ml (SEM = 7.53), respectively. Histologic examination of the endometrium showed normal secretory endometrium consistent with day 16-17 of spontaneous ovulatory cycle. Two women who received combined contraceptive pills/gonadotropin therapy showed inactive endometrium with subnuclear vacuoles and decidual reaction in the stroma similar to that observed in women on estrogen-progestin birth control medication.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Endometrial morphology and hormonal profiles in in vitro fertilization patients. 158 76

In an attempt to identify the embryos and cycles that have the best chances of resulting in establishment of pregnancies, after in vitro fertilization-embryo transfer (IVF-ET) treatment, the concentrations of sex hormone-binding globulin (SHBG) and cortisol-binding protein (CBP) were measured, using two new enzyme-linked immunosorbent assays in serum and follicle fluid (FF) from 30 women (125 FF) undergoing IVF-ET. The concentrations were compared to those of total estradiol and total progesterone, and correlated to oocyte cleavage and the establishment of pregnancies. Serum concentrations of CBP were significantly higher in women who became pregnant (1469 +/- 108) nM (+/- SEM] than in those who did not (CBP, 1200 +/- 58 nM; P less than or equal to 0.05). The concentrations of SHBG were not significant different in these two groups of women (72.4 +/- 9.3 and 60.8 +/- 4.2 nM, respectively; P greater than or equal to 0.10). By contrast, in FF significantly higher concentrations of both SHBG and CBP were found in women achieving pregnancy (SHBG, 56.1 +/- 2.8 nM; CBP, 1198 +/- 37 nM) than in those who did not (SHBG, 45.5 +/- 1.4 nM; P less than or equal to 0.001; CBP, 1079 +/- 29 nM; P less than or equal to 0.01). A positive correlation was found between serum and FF levels of both SHBG (r = 0.85; P less than or equal to 0.001) and CBP (r = 0.70; P less than or equal to 0.001). FF levels of estradiol and progesterone did not differ regardless of whether the oocyte cleaved. However, a significant reduction of estradiol was found in fluid from follicles in which the oocyte cleaved and resulted in pregnancy (3046 +/- 180 nM) than in fluid from follicles in which the oocyte cleaved but without establishment of pregnancy (4162 +/- 282 nM; P less than or equal to 0.001). There was no correlation between estradiol and SHBG and between progesterone and CBP. However, levels of FF progesterone above 15,000 nM combined with CBP concentrations above the mean concentration found in FF (1,127 nM) were related with oocyte cleavage in 87% of the cases. The overall cleavage rate is 56%. The higher levels of SHBG and CBP in serum compared to those in FF, and the positive relationship between serum and FF levels suggest that both proteins arise from the circulation. The similar levels in serum and FF indicate that neither SHBG nor CBP is responsible for maintaining the concentration gradient of estradiol and progesterone from follicle to plasma.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Levels of steroid-binding proteins and steroids in human preovulatory follicle fluid and serum as predictors of success in in vitro fertilization-embryo transfer treatment. 222 95

A quantitative bioassay was used to measure the concentration of platelet activating factor (PAF) in medium in which human embryos produced by IVF had been cultured and in various other biological fluids. Following extraction and partial purification, 121 of 228 (53%) media samples in which single human embryos were cultured for 24 h had PAF levels greater than found in corresponding control media. This was assigned as embryo-derived PAF and the corresponding embryos termed 'PAF-positive'. Medium from those PAF-positive embryos transferred to patients who achieved an ongoing pregnancy had a mean PAF concentration of 295 +/- 107 nM (mean +/- SEM, n = 55), which was significantly greater (P less than 0.03) than media of PAF-positive embryos transferred to patients who failed to become pregnant (75 +/- 27 nM, n = 66, t-test). The embryos with the faster cleavage rates tended to secrete more PAF (P less than 0.01). Although a greater proportion of culture media derived from embryos transferred to patients who achieved a pregnancy were PAF-positive (66 out of 121, 54.5%) compared with those transferred to patients who failed to achieve a pregnancy (55 out of 121, 45.4%), this was not significant (P greater than 0.05). It was observed that 13% of women who achieved a pregnancy had embryos transferred which did not produce significant amounts of PAF in vitro. This occurred in 26% of women not achieving pregnancy.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Measurement of human embryo-derived platelet-activating factor (PAF) using a quantitative bioassay of platelet aggregation. 235 16

In order to establish criteria for selection of the best ova in in-vitro fertilization-embryo transfer (IVF-ET) programes we have examined the follicular fluid (FF) levels of plasminogen activator (PA), collagenolytic activity, progesterone (P) and alpha 2 macroglobulin (alpha 2M) and related them to the success of pregnancy. PA activity was similar in FF of pregnant and nonpregnant cycles, 13.8 +/- 3.9 mU/ml versus 14.6 +/- 2.9 (mean +/- SEM) respectively. By contrast, FF from pregnant cycles exhibited lower collagenolytic activity (49.6 +/- 3.9% versus 67.9 +/- 3.0; P less than 0.001). Likewise, in a semi-quantitative assay of alpha 2M, only 18.4% of the aspirates from pregnant cycles showed a precipitation line, whereas 76.8% of those from non-pregnant cycles were positive. Levels of P in aspirates from pregnant cycles were in the intermediate range, as compared with those from non-pregnant cycles (0.06-5.5 micrograms/ml versus 0.02-12.0 micrograms/ml). All these assays can be completed before ET and performed in IVF-ET programmes. In conclusion, it seems that a combination of follicular alpha 2M levels and collagenolytic activity, and to a lesser extent addition of P assay, may serve as good criteria for selecting the best embryos for establishment of pregnancy.
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PMID:Follicular fluid contents as predictors of success of in-vitro fertilization-embryo transfer. 244 20

An important factor influencing the pregnancy rate after in vitro fertilization-embryo transfer (IVF-ET) appears to be the number of embryos transferred to the uterus. In this study, the influence of oocyte maturity and embryo quality on pregnancy rate was assessed in patients undergoing IVF-ET. Ovarian hyperstimulation was performed by human menopausal gonadotropin (hMG [n = 29]), clomiphene citrate (CC)/hMG (n = 81), and hMG/follicle-stimulating hormone (FSH [n = 13]) protocols. Oocyte maturity was graded on a scale from 1 to 5 based on the morphology of the ooplasm, cumulus mass, corona radiata, and membrana granulosa cells. Embryos were graded according to the symmetry of the blastomeres and the presence or absence of fragmentation. Mature preovulatory oocytes yielded the highest fertilization rates. No differences were found among the protocols in terms of fertilization rate, embryo quality, or pregnancy rate. When all protocols were combined, patients who conceived had a significantly higher number of embryos transferred than those who did not conceive (3.6 +/- 0.1 [mean = SEM] versus 2.7 +/- 0.1). When embryo quality was compared, there was no difference in the number of "B" embryos transferred between patients who conceived and those who did not (1.2 +/- 0.2 versus 1.2 +/- 0.1), but the patients who conceived had significantly more "A" embryos transferred (1.6 +/- 0.3 versus 0.8 +/- 0.1). These data suggest that the treatment protocol did not determine embryo quality. Furthermore, the increase in pregnancy rates seen with an increase in embryos transferred is the result of the transfer of more "A" embryos.
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PMID:The influence of oocyte maturity and embryo quality on pregnancy rate in a program for in vitro fertilization-embryo transfer. 250 52

Multidisciplinary andrology deals with clinical application and modern technology for the evaluation and differential diagnosis of male infertility with emphasis on morphological, anatomical, biochemical, immunological, hereditary, and microbiological parameters. Little is known about the effects of diet, disease, stress, environmental, and drugs on male-related unexplained infertility of couples. Regional, national, and international centers of multidisciplinary andrology should provide (1) extensive and unique clinical services; (2) a computerized "patient referral center," (3) self-learning packages (slides/tape programs) for patients; and (4) a computer link to the National Library of Medicine and the Drug Information Center. Specialized laboratories and clinics can be served by expert consultants, visiting professors, bilingual and well-trained clinicians, nurses, laboratory technologists, computer operators, and related allied health personnel. Patient education pamphlets, updated every few years, can be distributed during training workshops when an extensive network of remote teleprinters can be utilized. Qualified client location may install a printer to allow on-site printing of reports in the shortest possible time. Special mailing containers are provided to clients who wish to mail their laboratory specimens. Other clinical services may include the following: 1. Central source of communication and information in andrology; 2. International roster of multidisciplinary andrology centers; 3. Patient referral to centers and consultations for developing countries; 4. Screening of husbands and wives for in vitro fertilization/embryo transfer (IVF/ET); 5. Screening of couples with unexplained infertility for sexually transmitted diseases (STDs) including AIDS; 6. Exchange of research material and methodology; 7. Coordination of multicenter research; 8. Organizing training workshops for clinicians, nurses, and laboratory technicians; 9. Establishing a repository of films, video tapes, slides, catalogs, instrumentation, books, SEM photos, and atlases; 10. Publication and editorial assistance; 11. Consultation for the appropriate selection, purchase, and quality control of instrumentation (all on one computer system); 12. Evaluation of new diagnostic tools for idiopathic infertility and fertility regulation; 13.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Interdisciplinary andrology. STDs. AIDS research. 330 63

In order to evaluate the prognostic value of the heterologous ovum penetration test (HOP-test) the results of this test were compared with the fertilization rate of human ova in a programme for in vitro fertilization and embryo transfer (IVF/ET). Sperm from 29 husbands (23 normozoospermic and 6 with an abnormal semen profile) were exposed on one occasion to approximately 30 hamster ova in the HOP-test and on 1 to 3 occasions to 1 to 4 preovulatory oocytes obtained from the respective wives. The mean penetration rate (+/- SEM) of the hamster ova was 43 +/- 4% (range: 0-62%) for the normozoospermic men, and 23 +/- 6% (range: 0-47%) for the men with abnormal semen profiles. In 20 out of 23 couples in which the husbands were normozoospermic, sperm penetrated the hamster ova as well as they fertilized human ova; however, in one couple, a false-positive result was obtained (penetration of the hamster ova and no fertilization of the human oocytes) and in one couple a false-negative result occurred. One negative IVF result was correctly predicted by the HOP-test. In the 6 patients with disturbed sperm motility no correct positive or negative results were obtained, whilst 4 false-positive and 2 false-negative results occurred using the HOP-test. Although the number of patients with disturbed sperm motility was small, the data suggest that the HOP-test is of limited value in predicting fertility in an IVF program for couples with reduced fertility.
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PMID:Prognostic value of the heterologous ovum penetration test for human in vitro fertilization. 384 Oct 90

Cloned blastocysts developed in vitro for 7 d had a mean number of cells (82.86 +/- 5.35) as evaluated by nuclei counting in serial optical sections using confocal microscopy, after staining with propidium iodide. This number was not significantly different from that of control IVF embryos cultured under the same conditions during the same period (mean = 88.89 +/- 7.53). Semi-thin sections revealed that most of the blastocysts had an inner cell mass (10/12) and a blastocoele. Under transmission electron microscopy, the trophectoderm appeared well differentiated as a polarized epithelium with apical microvilli and lateral junctions including desmosomes with bound intermediate filaments. The cytoplasm sometimes contained immature mitochondria or a large number of residual bodies. About half of the blastocysts examined had a large amount of cellular debris in the perivitelline space or inside the blastocoele cavity. The cloned blastocysts were also able to hatch in vitro by day 8 and SEM indicated a normal morphology of the trophectoderm cells with numerous apical microvilli. The high number of excluded or degenerating cells found in some embryos may partially explain early embryonic mortality that follows transfer. However, these observations do not give a clear explanation for the high incidence of fetal losses.
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PMID:Cellular evaluation of bovine nuclear transfer embryos developed in vitro. 853 65

Four experiments determined the kinetics of in vitro maturation and fertilization of cat oocytes and the effects of prolonged cold storage of ovaries before oocyte recovery on in vitro maturation/in vitro fertilization (IVM/IVF) success. Domestic cat ovaries were collected at ovariohysterectomy and stored at 4 degrees C in PBS until oocyte recovery and culture in Eagle's minimal essential medium (EMEM) containing FSH, LH, oestradiol and BSA for maturation. In Expt 1, meiotic maturation was assessed at 0, 12, 24, 38 and 48 h of culture. After 24 h, > 61% of oocytes were in telophase I or metaphase II. In Expt 2, oocytes were recovered from ovaries stored for 24, 48 or 72 h and cultured in EMEM for 24 h. There was no difference among groups (P > 0.05) in the ability to achieve nuclear maturation (mean +/- SEM, 57.1 +/- 5.3%, 60.4 +/- 5.4%, 55.4 +/- 15.1% for 24, 48 and 72 h, respectively). Fertilization and embryo development after insemination at 16, 24, 32, 40 and 48 h of culture were examined in Expt 3. Of 98 oocytes inseminated at 32 h, 69% cleaved, 59% developed into morulae and 13% into blastocysts, more (P < 0.05) than those oocytes inseminated at earlier and later times. Development to blastocysts occurred after insemination at 16 (1.2%), 24 (9.1%) and 32 (13.3%) h of culture, but not after insemination at 40 or 48 h. Expt 4 involved cold storage of ovaries for 24, 48 or 72 h before oocyte recovery and insemination at 32 h of culture (the optimal time measured in Expt 3). Compared with storage for 24 h, fertilization success was lower (P < 0.05) in the 48 and 72 h groups, and, although 9.1% of inseminated oocytes from the 24 h storage group developed to blastocysts, none (P < 0.05) achieved this stage after 48 or 72 h of storage. These results indicate that domestic cat oocytes reach nuclear maturity by 24 h in culture and can be fertilized and develop to blastocysts optimally after insemination at 32 h. Oocytes recovered from ovaries stored at 4 degrees C for up to 72 h are capable of reaching telophase I or metaphase II in vitro. However, only oocytes stored within the ovary for 24 h produce blastocysts, indicating that the ability to achieve nuclear maturation is an inadequate indicator of fertilization and developmental competence.
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PMID:Development to blastocysts of domestic cat oocytes matured and fertilized in vitro after prolonged cold storage. 866 38

In bovine in vitro embryo production, the IVM step is rather successful with 80% of the oocytes reaching the MII stage. However, the extent to which the process limits the yield of viable embryos is still largely unknown. Therefore, we compared embryonic developmental capacity during IVC of IVF oocytes which had been matured in vitro with those matured in vivo. In vitro maturation was carried out for 22 h using oocytes (n = 417) obtained from 2- to 8-mm follicles of ovaries collected from a slaughterhouse in M199 with 10% fetal calf serum (FCS), 0.01 IU/mL LH, and 0.01 IU/mL FSH. In vivo matured oocytes (n = 219) were aspirated from preovulatory follicles in eCG/PG/anti-eCG-superovulated heifers 22 h after a fixed time GnRH-induced LH surge; endogenous release of the LH surge was suppressed by a Norgestomet ear implant. This system allowed for the synchronization of the in vitro and in vivo maturation processes and thus for simultaneous IVF of both groups of oocytes. The in vitro developmental potential of in vivo matured oocytes was twice as high (P < 0.01) as that of in vitro matured oocytes, with blastocyst formation and hatching rates 11 d after IVC of 49.3 +/- 6.1 (SEM; n = 10 heifers) vs 26.4 +/- 1.0% (n = 2 replicates), and 39.1 +/- 5.1% vs 20.6 +/- 1.4%, respectively. It is concluded that IVM is a major factor limiting in the in vitro production of viable embryos, although factors such as the lack of normal preovulatory development of IVM oocytes contributed to the observed differences.
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PMID:Improved in vitro embryo development using in vivo matured oocytes from heifers superovulated with a controlled preovulatory LH surge. 1073 99


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