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S Oehninger

Publications and source records attributed to S Oehninger.

At least 109 records · Page 6Linked to original sources

Validation of the hemizona assay (HZA) in a monkey model. II. Kinetics of binding and influence of cryopreserved-thawed spermatozoa.

We compared fresh and frozen-thawed cynomolgus monkey spermatozoa tight binding to the zona pellucida under hemizona assay (HZA) conditions. Monkey oocytes were recovered after superovulation and stored in salt solution. Matching hemizonae were obtained by micromanipulation. Semen, obtained by electroejaculation, was used fresh or was cyropreserved, thawed, and washed by swim-up separation. At the standard initial dilution of 500,000 motile sperm/ml (or 5 x 10(4) motile sperm/hemizona), binding was significantly higher for fresh sperm (P = 0.00004). For frozen-thawed samples, there was a linear increase in the number of tightly bound sperm with increasing sperm concentration (r = 0.95). At 1.5 x 10(6) motile sperm/hemizona, binding of frozen-thawed spermatozoa was similar to that of fresh at a standard concentration. Kinetic studies showed peak binding at 1 hr of gametes coincubation. We conclude that, in this monkey model, the HZA is a valuable bioassay for evaluation of sperm binding to the zona pellucida, the initial requisite for fertilization and embryo development.

Animals↗

Fucoidin binding activity and its localization on human spermatozoa.

We previously reported that fucoidin (a polymer of predominantly sulfated L-fucose) significantly inhibits: (1) tight binding of human sperm to human zona pellucida in vitro and (2) stimulation of the acrosome reaction by acid solubilized human zona pellucida. Here, we determined fucoidin binding activity on human spermatozoa and its localization on both live and permeabilized human sperm populations. A typical binding curve was demonstrated with biotinylated fucoidin. In competitive inhibition assays with unlabelled fucoidin or human sperm membrane extracts, IC50's were 4.0 micrograms/ml and 31.4 micrograms/ml, respectively. Fucoidin binding was localized over the acrosomal region of methanol-fixed human sperm and this pattern of binding significantly decreased from 92 +/- 3% to 74 +/- 6% with calcium ionophore A23187 treatment (p < 0.01). Binding of fucoidin-coated beads to live (non-permeabilized) human sperm was less than 1%. Addition of the detergent, Triton-X, to permeabilize sperm membranes resulted in a significant increase in binding (p = 0.001). These results provide evidence for the presence of a fucoidin binding compound in human spermatozoa that is localized to the membranes of the acrosomal region and can be extracted by a mild detergent extraction. Absence of binding by fucoidin to intact but not permeabilized spermatozoa suggests that the heteropolysaccharide binds to a receptor within the acrosomal matrix. However, further investigation is warranted to determine whether a fucoidin binding site is present both at the sperm's surface for the initial contact with the zona pellucida, and also for secondary binding after exposure of the acrosomal membranes.

Acrosome↗

The hemizona assay: its role in identifying male factor infertility in assisted reproduction.

OBJECTIVES: To identify male factor infertility among a group of patients in an assisted reproductive program (phase 1) and to evaluate the hemizona assay (HZA) in the diagnosis and prognosis of such a program (phase 2). DESIGN: The IVF performance of normal gametes in the Tygerberg program were critically evaluated. Female patients were classified as pure tubal factor infertility, having a normal FSH:LH ratio on day 3 of the menstrual cycle. All participating women produced three or more preovulatory oocytes at retrieval and were inseminated with sperm considered normal by all present diagnostic criteria. The total and normal fertilization rate thresholds were defined in that group. Using those thresholds, couples tested for sperm binding in the HZA (n = 48) were used and divided into two groups according to their fertilization rates, namely group 1, low fertilization (< 55%) and group 2, normal fertilization (> 55%). SETTING: University-based tertiary care center. PATIENTS: Ninety-nine couples (589 oocytes) with pure tubal factor infertility and normal male factor were used in phase 1. Forty-eight couples with normal and abnormal male factors that had both HZA performed and IVF treatment were included in phase 2. RESULTS: Investigation of the performance of normal gametes in 99 couples (589 oocytes) revealed the total fertilization rate (total number of oocytes fertilized/total number of oocytes inseminated) was (mean +/- SD) 88.6% +/- 16.8% and the normal fertilization rate (total number of oocytes with normal fertilization/total number of oocytes inseminated) was 81.3% +/- 22%. The minimum total fertilization rate that can be considered normal in the Tygerberg program using mean--2 SD is therefore 55% and for normal fertilization rate is 37%. The group with low fertilization rate (< 55%) showed a mean hemizona index (HZI) significantly lower; nevertheless, the distribution overlapping indicates a low discriminating power of the HZA. A sensitivity of 75% and a specificity of 75% were found; the positive and negative predictive values were 81% and 68%, respectively. CONCLUSIONS: The results indicated the HZA and HZI contribute important information and can serve in conjunction with other semen characteristics as useful tools during the diagnosis of the male factor in assisted reproduction.

Chorionic Gonadotropin↗

Effect of progesterone on human zona pellucida sperm binding and oocyte penetrating capacity.

OBJECTIVE: To determine if sperm exposure to P produces an enhancement in its fertilizing capacity. DESIGN: Sperm from fertile donors were exposed to P at 0.1 and 1.0 microgram/mL for 1 or 24 hours. The effects on hyperactivated (HA) motility at 1 and 4 hours, acrosome reaction (as determined by Pisum sativum agglutinin or T6/antibody techniques), on human zona pellucida binding (by using the hemizona assay), and on the penetrating ability (by using the zona-free hamster ova assay) were evaluated. RESULTS: Exposure to P at 1.0 microgram/mL enhanced HA motility after 1 and 4 hours of P exposure, the acrosome reaction after 24 hours' incubation, the number of sperm bound/hemizona after 1-hour incubation, and the penetration rates in the hamster ova assay at both incubation intervals. CONCLUSION: Sperm exposure to P enhances its fertilizing capacity in fertile men, and further investigation is warranted as a possible treatment for male factor patients.

Acrosome↗

The ability of the hemizona assay to predict human fertilization in different and consecutive in-vitro fertilization cycles.

The objective of this prospective study was to examine the ability of the hemizona assay (HZA) to predict fertilization outcome of mature, pre-ovulatory oocytes under in-vitro fertilization (IVF) conditions. Since a large number of patients were evaluated over a long period, the power of the HZA to prognosticate fertilization results in the same and subsequent (consecutive) IVF cycles of those same patients was assessed. For IVF, only metaphase II oocytes were used. For the HZA, both fresh oocytes donated by patients at the time of IVF and oocytes recovered from surgically removed ovarian tissue (and salt-stored) were used, and bisected by micromanipulation techniques. Matching hemizonae were co-incubated either with spermatozoa from the patient (test) or from a fertile man (control) for 4 h. The number of spermatozoa tightly bound to the zona was counted. Patients (n = 112) were divided into two groups based on HZA results (expressed as HZA index or HZI): HZI > or = 30% (n = 72) and < 30% (n = 40). The patients with HZI < 30% had significantly lower fertilization rates in both the HZA-IVF cycle and in subsequent cycles compared to patients with HZI > or = 30% (P < 0.03). Linear discriminant analysis indicated the HZA to have a sensitivity of 84%, and positive and negative predictive values of 85 and 70% respectively, for prediction of fertilization outcome in a total of 233 cycles. It was concluded that the HZA is a good predictor of fertilization rate in vitro, and can be used in the IVF setting to supply additional clinical information in male-factor patients.

Female↗

Functional and morphological features of spermatozoa microaspirated from the epididymal regions of cynomolgus monkeys (Macaca fascicularis).

In most species examined to date, spermatozoa mature during transit through the epididymis to become functionally competent to swim progressively, fertilize oocytes, and produce viable offspring. In the studies presented here, the status of spermatozoa obtained from the regions of the epididymis of the cynomolgus monkey (Macaca fascicularis) was examined. Epididymal fluid was collected by microaspiration from the caput, corpus, and cauda regions. The spermatozoa obtained were evaluated for morphology, motion parameters, and tight zona pellucida binding and were compared to spermatozoa collected by electroejaculation. Epididymal and ejaculated sperm differed morphologically only in the location of the cytoplasmic droplet. Motile sperm recovered by swim-up procedure exhibited a significantly higher proportion of sperm with more distal cytoplasmic droplets than the original aspirated samples (p < 0.02). Poor duration of movement was most noticeable for corpus spermatozoa, and to a lesser degree for cauda spermatozoa, when the motion parameters of velocity, linearity, and amplitude of the lateral head were examined. Zona pellucida binding was decreased only for sperm microaspirated from the caput region; sperm obtained from the corpus and cauda regions bound comparably to those collected via electroejaculation. These results suggest that by the time cynomolgus sperm reach the corpus region of the epididymis, they are functionally competent for tight zona binding. However, these nonhuman primate sperm may not complete the metabolic changes requisite for sustained sperm motility until reaching the most distal regions of the epididymis.

Animals↗

Prediction of fertilization in vitro with human gametes: is there a litmus test?

OBJECTIVES: The objectives of this study were as follows: (1) to evaluate the relationships between sperm concentration, morphologic pattern, motion parameters, and sperm-zona pellucida binding capacity and (2) to assess their ability to predict fertilization outcome under in vitro fertilization conditions. STUDY DESIGN: Semen samples from 44 infertile men were prospectively evaluated for density, morphologic pattern (strict criteria), computerized motion parameters (motility, velocity, and linearity), and hemizona assay (outcome expressed as hemizona assay index), and results were correlated with fertilization outcomes of preovulatory oocytes during in vitro fertilization. RESULTS: Of all sperm parameters, morphologic pattern was the best predictor of the ability of the sperm to bind to the zona pellucida. Hemizona assay index was the best predictor of fertilization rate. Stepwise regression analysis provided a model of hemizona assay index plus motility with highest predictability (R2 = 53.4%). CONCLUSIONS: (1) The influence of sperm morphologic pattern as a prognosticator of fertilization outcome is established in major part as a determinant of sperm binding ability to the zona pellucida; (2) the hemizona assay provides a robust index that is highly predictive of the potential of human gametes to achieve fertilization and highlights its use in infertility and contraception testing.

Cluster Analysis↗

Performance of cryopreserved pre-embryos obtained in in vitro fertilization cycles with or without a gonadotropin-releasing hormone agonist.

OBJECTIVE: To evaluate the viability and potential for pregnancy of cryopreserved/thawed pre-embryos obtained after ovarian stimulation using gonadotropin-releasing hormone agonist (GnRH-a) adjunct therapy. DESIGN: Retrospective clinical evaluation of all patients receiving a gonadotropin ovarian stimulation protocol (follicle-stimulating hormone/human menopausal gonadotropin [FSH/hMG]) with/without GnRH-a. SETTING: Academic tertiary clinical care unit. PATIENTS: Patients receiving leuprolide acetate (LA)/FSH/hMG (n = 136: LA in the luteal phase; long protocol) were compared with patients receiving FSH/hMG alone (n = 130) within the same time-frame in our program (April 1987 through October 1989). INTERVENTIONS: All patients had both a cycle in which pre-embryos were transferred fresh and a cycle of thaw of cryopreserved pre-embryos (frozen at the pronuclear stage in a slow freeze-thaw protocol using 1,2 propanediol) transferred in monitored natural cycles. MAIN OUTCOME MEASURES: Groups were similar in age, etiology of infertility, and cycle day 3 serum FSH levels; a significantly higher (P less than 0.001) number of preovulatory oocytes was recovered in the GnRH-a group. Both groups of patients were transferred an equal number of pre-embryos at the time of IVF. Cycles with frozen/thawed pre-embryos were evaluated based on the analysis of the three main variables that demonstrate cryopreservation efficiency: survival rate, implantation rate, and term pregnancy rate (PR). RESULTS: Non-GnRH-a group (113 transfers): pre-embryo survival, 71.5%; PR/transfer, 24.7%; implantation rate, 16.0%; GnRH-a group (125 transfers): pre-embryo survival 71.6%; PR/transfer, 32.8%; implantation rate, 12.0% (no significant differences). CONCLUSIONS: The use of GnRH-a produced pre-embryos of equal aptitude for development after cryopreservation at the pronuclear stage when compared with a similar gonadotropin stimulation treatment without GnRH-a.

Adult↗

Peritoneal fluid from patients with endometriosis decreases sperm binding to the zona pellucida in the hemizona assay: a preliminary report.

OBJECTIVE: To evaluate the effect of peritoneal fluid (PF) from patients with endometriosis on gamete interaction under hemizona assay (HZA) conditions. DESIGN: The HZA was used to study the effect of PF from patients with endometriosis on sperm binding to the zona pellucida using media and normal PF as controls. SETTING: The patients were collected from a university hospital infertility clinic. PATIENTS: Peritoneal fluid from 16 women being evaluated for infertility or sterilization who were found to have endometriosis at surgery was used. Three normal patients, who were being sterilized, had PF that was used as a control. RESULTS: Results suggest that there is a significant reduction in the number of tightly bound sperm to the zona surface in endometriosis specimens as reflected in the hemizona index and that this effect is directly related to the stage of the disease. CONCLUSIONS: Although the sample is small, this methodology may help to elucidate one of the mechanisms responsible for endometriosis-associated infertility.

Ascitic Fluid↗

Embryo transfer to the uterus or the fallopian tube after in vitro fertilization yields similar results.

The outcome of ZIFT and IVF-ET was compared in consecutive nontubal factor patients in a prospective fashion. Groups did not differ in characteristics and were matched by the number of prezygotes/pre-embryos transferred. Overall, implantation, pregnancy, miscarriage, and ongoing PRs were not statistically different. These results suggest that ZIFT offers no significant advantage over IVF-ET for the treatment of nontubal infertility.

Adult↗

Successful fertilization and pregnancy outcome in in-vitro fertilization using cryopreserved/thawed spermatozoa from patients with malignant diseases.

Cryopreservation of spermatozoa before treatment is the only proven effective method available to circumvent the sterilizing effect of therapy in some patients with malignant diseases. Because of impaired sperm quality after freezing and thawing in-vitro fertilization/embryo transfer (IVF/ET) was indicated in 10 patients (12 cycles) during 1986-1990. The patient's mean age was 33.4 +/- 1.6 years. The following diagnoses were made: seminoma (1), testicular carcinoma (3), leiomyosarcoma of the prostate (1), Wegener's granulomatosis (1), non-Hodgkin's (1) and Hodgkin's lymphoma (3). When motile spermatozoa could be recovered after thawing, the total fraction of motile spermatozoa after swim-up separation ranged from 0.2 to 4.2 x 10(6) spermatozoa/ml (eight patients, nine cycles). In all these cases, insemination was performed with multiple oocytes per dish. Fertilization was achieved when swim-up recovered a mean of 1.8 +/- 0.5 x 10(6) spermatozoa/ml and when insemination was performed with at least a calculated concentration of motile spermatozoa of 1 x 10(5) spermatozoa/oocyte. The fertilization rate of preovulatory oocytes was 60%. Four patients achieved a pregnancy: two of them delivered a single healthy baby, one delivered triplet healthy babies and one had a preclinical abortion. In two patients (three cycles), no motile spermatozoa were recovered after thawing, and micromanipulation of oocytes for assisted fertilization was performed. Although fertilized oocytes were transferred, those couples did not achieve a pregnancy. Patients with lymphopathies had the best results, whilst those with testicular neoplasms had the poorest outcome, thus suggesting a poor gametogenic function in the non-affected testis. These results give hope to some patients with malignant diseases to maintain their reproductive capacity through sperm banking and IVF/ET.

Adult↗

Sonographic appearance of the endometrium: the predictive value for the outcome of in-vitro fertilization in stimulated cycles.

We evaluated the predictive value for pregnancy of the endometrial thickness and pattern assessed by vaginal sonography on the day of human chorionic gonadotrophin (HCG) injection and the day of embryo transfer in 74 stimulated cycles for in-vitro fertilization (IVF) which were analysed prospectively. Thickness was measured from the echogenic interface of the endometrial-myometrium junction in transverse fundal sections. The distribution of endometrial pattern on the day of HCG was 19 A (poor quality) cases (25.7%) and 55 B (good quality) cases (74.3%). On the day of embryo transfer, 16 out of the 19 A cases (84%) remained as A pattern, while the remaining three cases (16%) had changed to B pattern; of the 55 B cases, 29 (53%) remained the same, while 26 cases (47%) changed to A pattern. There was no significant correlation between the endometrial pattern on the day of HCG and/or on the day of embryo transfer and the peak serum oestradiol levels, the number of preovulatory oocytes aspirated, the serum progesterone levels and oestradiol: progesterone ratio on the day of transfer. In contrast, on the day of HCG, endometrial thickness correlated with serum oestradiol levels on that day (P = 0.02). The presence of pattern A on the day of HCG was associated with a lower pregnancy rate [three out of 19 cases, (15.8%)], compared to pattern B [16 out of 55 cases, (29.1%)], although this was not statistically significant (P greater than 0.05). On the day of embryo transfer, comparable pregnancy rates between A pattern [11 out of 42 cases, (26.1%)] and B pattern [eight out of 32 cases, (25%)] were found.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗

Pure human follicle stimulating hormone has a role in the treatment of severe male infertility by assisted reproduction: Norfolk's total experience.

Fifty patients [79 in-vitro fertilization (IVF) cycles] with severe male factor infertility were included in an experimental clinical trial running from October 1987 to March 1991 to assess the potential of systemic follicle stimulating hormone (FSH) treatment to improve sperm fertilizing ability in IVF. Two groups were defined: a secondary group (24 patients, 33 IVF cycles) with a history of failed fertilization in previous IVF attempts and a primary group (26 patients, 46 IVF cycles) with poor sperm parameters which suggested that fertilization would not occur according to previously established criteria. Basic semen analysis and a battery of endocrine radioimmunoassays [serum FSH, luteinizing hormone (LH), oestradiol, prolactin and testosterone] were performed in these patients. Bioactive-FSH and LH were also determined in some patients. For this study, pure FSH was administered (150 IU i.m. three times per week) for at least 3 months, after which the semen analysis and endocrine tests were repeated. Although no significant changes were observed after FSH therapy, either in the endocrine profile or in the basic semen parameters, except for FSH radioimmunoassay levels, the fertilization rate of pre-ovulatory oocytes was significantly improved from 2 to 54.4% in the secondary group; the primary group showed a 52.3% fertilization rate. Eighteen clinical pregnancies were achieved, 11 in the primary group and seven in the secondary group, giving 30 and 26% term pregnancy rates per transfer respectively. These results, which are in complete agreement with our preliminary study, re-emphasized the benefits of systemic FSH administration as an adjunct to assisted reproduction in selected cases of severe male factor infertility.(ABSTRACT TRUNCATED AT 250 WORDS)

Embryo Transfer↗

Male infertility: the impact of assisted reproductive technologies.

Spermatozoal abnormalities are present in up to 40% of infertile couples. Multiple etiologies may be responsible for these disorders, including pretesticular, testicular, and ductal causes. An adequate andrologic consultation, including history, physical examination, and repeated semen analysis, should be performed routinely in these cases. Evaluation of the basic sperm parameters, including sperm concentration, motion parameters (as evaluated by computer-assisted semen analysis), and sperm morphology (as judged by strict criteria), constitute the first obligatory step for a critical evaluation of male factor patients. Patients in whom fertilization disorders are suspected should be evaluated through bioassays of sperm-oocyte interaction, including the heterologous sperm penetration assay and hemizona assay, a bioassay of sperm-zona binding capacity highly predictive of in vitro fertilization outcome. Male disorders can be subjected to specific therapies (surgical or medical), or to empiric therapeutic modalities. Assisted reproductive technologies have enhanced our understanding of the physiopathology of spermatozoal disorders and also have ostensibly improved pregnancy rates in male factor patients.

Female↗

Comparison of sperm binding potential of uninseminated, inseminated-unfertilized, and fertilized-noncleaved human oocytes under hemizona assay conditions.

In this study, human oocytes obtained after ovarian hyperstimulation for in vitro fertilization (IVF) and gamete intrafallopian transfer (GIFT) were utilized to evaluate sperm/zona pellucida binding potential. Three groups of oocytes were evaluated: 1) uninseminated; 2) inseminated-unfertilized; and 3) fertilized-uncleaved. All oocytes had undergone germinal vesicle breakdown at the time of retrieval and were salt-stored (pH 7.2) for not more than 30 days. Sperm binding was recorded under hemizona assay (HZA) conditions using spermatozoa from eight fertile men (HZA control) and from 1) four teratozoospermic (HZA test) and 2) four normozoospermic (HZA test) infertile men. First, the mean numbers (+/- SD) of sperm tightly bound for fertile controls and teratospermic men to hemizonae from uninseminated oocytes were 69.7 +/- 16 and 14.5 +/- 7, respectively (P = 0.02). Likewise, hemizonae from uninseminated oocytes bound 102.0 +/- 19 and 114.0 +/- 28, respectively, for fertile controls and normospermic men (P = 0.5). Second, hemizonae obtained from inseminated-unfertilized IVF oocytes bound 44.2 +/- 12 and 19.7 +/- 6 for fertile controls and teratospermic men, respectively (P = 0.02). This category of oocytes bound 100.5 +/- 7 and 108.5 +/- 11 sperm, respectively, for fertile controls and normospermic semen (P = 0.3). Third, HZA results of fertilized but uncleaved oocytes showed a mean number of tightly bound sperm of 6.0 +/- 4 compared with 65.0 +/- 1 in control, uninseminated oocytes using fertile sperm. These results demonstrate that uninseminated and inseminated-unfertilized human oocytes, salt-stored under controlled pH conditions, give reliable information regarding sperm binding potential under HZA conditions.

Female↗

Sperm selection capacity of the human zona pellucida.

Previous hemizona assay (HZA) results have illustrated a positive and significant correlation between the percentage of morphologically normal spermatozoa in the semen and the number of spermatozoa tightly bound to the zona pellucida. The present study was designed to evaluate the morphologic features using strict criteria of spermatozoa tightly bound to the zona pellucida. Semen samples of 4 normozoospermic and 11 teratozoospermic men were used to compare the percentage of normal spermatozoa in the semen with that found 1) after swim-up separation and 2) bound to the zona under HZA conditions. The mean (+/- SEM) % normal forms for normozoospermic men in semen, after swim-up and zona-bound spermatozoa were 21.5 +/- 1.6, 27.5 +/- 2.9, and 44.8 +/- 3.4, respectively. A significantly higher % of normal forms were found among zona-bound sperm compared to swim-up forms (p = 0.02) and seminal sperm (p = 0.02). The mean % of normal sperm forms present in semen, after swim-up and zona pellucida-binding for teratozoospermic men, were 3.7 +/- 0.9, 5.8 +/- 1.6 and 15.6 +/- 3.1, respectively. Significant differences existed between the % of normal sperm forms found in the swim-up and zona-bound spermatozoa (P = less than 0.01 and P = less than 0.0003, respectively) compared to the original ejaculates. Results indicate that a selective process against abnormal spermatozoa occurs at the site of the zona pellucida.(ABSTRACT TRUNCATED AT 250 WORDS)

Cell Separation↗

Hemizona assay: use of fresh versus salt-stored human oocytes to evaluate sperm binding potential to the zona pellucida.

The hemizona assay (HZA) has a high predictive value for in vitro fertilization (IVF) results. Oocyte quality plays a significant role in the validation of this test. The question was asked whether human salt-stored oocytes (up to 30 days) are damaged and subsequently lose their sperm binding capacity when compared to fresh human oocytes. Equivalent binding in both the salt-stored and the fresh group of oocytes was observed in the hemizonae incubated with normal semen as well as in their matching halves incubated with semen from an infertile man. Based on the results, we conclude that salt-stored oocytes (pH 7.0) give reliable information regarding sperm binding potential under HZA conditions.

Female↗

Recurrent failure of in vitro fertilization: role of the hemizona assay in the sequential diagnosis of specific sperm-oocyte defects.

The results of predictive fertilization bioassays (hemizona assay, hamster ova-human sperm penetration assay), in vitro fertilization treatment, fertile donor cross-match tests with either sperm or oocytes, and oocyte micromanipulation for assisted fertilization were used to establish a pathophysiologic diagnosis in cases of recurrent failed fertilization in vitro. Disorders of sperm function manifested at the level of zona binding, zona penetration, oolemma fusion, and pronuclear decondensation as well as oocyte anomalies were considered to represent the specific gamete defects that led to abnormal sperm-oocyte interactions (i.e., failed fertilization). Our findings show that the sequential application of bioassays can elucidate specific sperm or oocyte defects that characterize functional abnormalities among sperm populations.

Acrosome↗