The association of semen factors with the recovery of Ureaplasma urealyticum.

Ninety-six semen samples from males under examination for suspected subfertility were examined to determine if alterations in specific semen parameters could be associated with the presence or absence of Ureaplasma urealyticum. Forty-three (44.8%) of the specimens cultured positive for U. urealyticum. Both positive and negative specimens have a normal average motility rating. Sperm counts (millions per milliliter) were slightly higher in U. urealyticum positive samples. Average ejaculate volumes were identical between the two groups. Morphologic abnormalities were not found more frequently in U. urealyticum-positive samples. No statistically significant difference between seminal parameters of positive and negative samples were noted with the one-way analysis of variance test. Nonparametric analysis produced similar nonsignificant results.

Vital initiation of pregnancy (VIP) using human menopausal gonadotropin and human chorionic gonadotropin ovulation induction: Phase I--1981.

Laparoscopies for oocyte aspiration in 31 cycles were performed on 25 patients receiving human menopausal gonadotropin and human chorionic gonadotropin. Sixty oocytes were aspirated, of which 48 were considered preovulatory. Ninety-seven percent (58 of 60) of the oocytes were found in the original aspirate, and the remaining oocytes were found in either the first or second follicle wash. The fertilization rate per preovulatory oocyte was 33% (16 of 48), whereas on a per cycle basis it was 39% (12 of 31). A total of 15 conceptuses (2-cell = 5; 3-cell = 3; 4-cell = 7) were transferred to 12 patients, and two pregnancies were established. These pregnancies were established by transfers of 3-cell and 4-cell conceptuses at approximately 47 hours after insemination. Both pregnancies resulted in term deliveries of normal infants.

Correlation of postcoital evaluation with in vitro sperm cervical mucus determinations and ureaplasma cultures.

Fifteen trials were completed in 14 couples during an infertility evaluation. The postcoital test (PCT) was accomplished in a standardized manner. Also, standardized determinations of the sperm-cervical mucus penetration test (SCMPT) with the addition of cross-testing (X-test) utilizing bovine cervical mucus (BCM) and normal donor semen (NDS) were done. Male and female samples were also cultured for Ureaplasma urealyticum using U9-B indicator broth and A-7 agar. The PCT and SCMPT agreed in 87% (13/15) of the cases. Utilizing BCM and NDS, where possible, the causative factor was the cervical mucus in 54% (7/13); semen factor in 15% (2/13); both factors in 8% (1/13); and undetermined in 23% (3/13). U. urealyticum cultures were positive in 40% (6/15) of the cases. Analysis of the results revealed: (1) significant correlation between PCT and SCMPT (P less than 0.01); (2) no significant difference among the PCT, SCMPT, and X-test, indicating that the cervical mucus was the causative factor; and (3) no U. urealyticum correlation with the PCT or the SCMPT. Thus, laboratory SCMPT and X-test correlated with the PCT, providing additional information concerning the causative factor in infertility. The U. urealyticum status in cervical mucus and semen cannot be determined from the PCT nor the SCMPT.

Effects of human serum and plasma on development of mouse embryos in culture media.

Mouse embryos were cultured in Ham's F-10 medium (unsupplemented) and Ham's F-10 medium supplemented with either 15% fetal cord serum, 15% fetal cord plasma, 15% maternal serum, or 15% maternal plasma. None of the blood supplements significantly increased the numbers of embryos that developed to blastocysts, and the last three of the blood supplements inhibited embryo development. The cause of the inhibition was not identified. Estradiol concentrations of blood samples used as media supplements were found not to correlate with inhibition of embryo.

Evaluation of human fetal cord sera, Ham's F-10 medium, and in vitro culture materials with a mouse in vivo fertilization system.

A total of 54 human fetal cord sera, 42 lots of Ham's F-10 medium, and 36 lots of plastics were tested during a 10-month period with a mouse in vivo fertilization system. Two-cell embryos were collected from the oviducts of superovulated and mated (C57BL/6 X CBA)F1 mice. After collection, 2-cell embryos were distributed among test media and a modified Krebs-Ringer control medium. The test material passed quality control standards if a minimum of 80% of the original 2-cell embryos reached the blastocyst stage. Of the 54 cord sera examined, 8 (15%) were below our standards; 6 (14%) of the Ham's F-10 media and 4 (11%) of the plastics failed to pass minimum requirements. In all cases, the failed materials promoted slower-growing mouse embryos and increased the number of degenerates. It is our opinion that the mouse in vivo fertilization system has a valuable place in any in vitro fertilization program.

Vital initiation of pregnancy (VIP) using human menopausal gonadotropin and human chorionic gonadotropin ovulation induction: phase II--1981.

In a program for in vitro fertilization, laparoscopies for oocyte aspiration were performed on 24 patients receiving human menopausal gonadotropin and human chorionic gonadotropin. Of the 40 preovulatory oocytes that were recovered from these patients, 33 (83%) were fertilized and 30 (75%) cleaved and were transferred. Ten immature oocytes were collected, and attempts were made to mature these in vitro prior to insemination. All ten oocytes (100%) did fertilize, and seven (70%) cleaved and were transferred. Morphologic variation was noted between cleaving conceptuses, even in those conceptuses responsible for establishing pregnancies. Five pregnancies resulted from 19 embryo transfers (26%).

Maturation and fertilization of morphologically immature human oocytes in a program of in vitro fertilization.

Oocytes of varying stages of maturity were aspirated from follicles primed with either human menopausal gonadotropin (hMG) and human chorionic gonadotropin (hCG) or a combination of follicle-stimulating hormone (FSH), hMG and hCG. Of the aspirated oocytes from 44 cycles, 74 were considered to be immature by virtue of morphologic characteristics of the oocytes and the degree of intercellular expansion of the associated cumular and membrana granulosa cells. After incubation periods of 22 to 35 hours in a Ham's F-10-based culture medium, these immature oocytes were inseminated with sperm donated by the patient's husband. Ultimately, 44 conceptuses were transferred to the respective uteri of 30 patients. Eight pregnancies were established as a result of these 30 transfers, two of which resulted from the transfer of only developed immature oocytes.

Effects of season, environmental temperature, size of dams, and age of breeder males on numbers of embryos obtainable from superovulated mice.

Four factors were tested for their effects on numbers of embryos obtainable from mice injected with pregnant mare serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG) to induce superovulation. The factors tested were: (1) season of year; (2) environmental temperature (in the range of 17.8-23.7 degrees C); (3) age of breeder male mice (in the range of 10-76 weeks); and (4) size of female mice (in the range of 16-30 g body weight). Each of the first three factors was tested for its effect on mating and consequently on embryo yield; the fourth was tested for its effect on numbers of embryos yielded by those females that did mate. In all experiments hybrid female mice (C57BL/6J X CBA/J) were injected with 5 IU of PMSG and 5 IU of hCG and were crossed with males of the CD-1 strain. None of the factors tested was found to have a statistically significant effect on embryo yield. Other factors that should be examined for their effects on mating and/or ovulation are discussed.

Utilization of embryos from previously superovulated mice in an in vivo fertilization system.

When stringent quality control has been an integral part of an IVF-ET program, there has been an indication of more consistent pregnancy rates (8,9). We were therefore hesitant to introduce any variables, such as reutilizing previously stimulated mice, which might affect our quality-control system. However, from the results of our study there appeared to be no significant difference in the number of two-cell embryos retrieved between the control and the experimental groups. No difference was noted in the percentages of embryos reaching beta blastocyst. Although not significant, there did appear to be a decrease in mating when mice were stimulated more than once.

An indirect enzyme-linked immunosorbent assay (ELISA) for the detection and quantitation of antisperm antibodies.

Several recent comparative investigations using various assays to detect and quantitate levels of antibody to human spermatozoa have produced widely varying results. In an attempt to reduce test variability, an indirect enzyme-linked immunosorbent assay (ELISA) was devised to measure antisperm antibodies. A standardized protocol was adapted employing sperm adsorption to polystyrene microtiter plates, at a density of 10(5) sperm per well, serum and enzyme-conjugate incubation conditions at 37 degrees C for 60 min, and three ten-minute washes between incubations, using phosphate-buffered saline containing Tween-20. Using antihuman sperm antisera generated in rabbits, the ELISA was shown to yield significantly detectable antibody at dilutions of 1/16,384. The ELISA demonstrated approximately 89% reproducibility (ie, 100% minus the coefficient of variation) for an "intraassay" study wherein 300 determinations were performed on the same day on sperm from ten donors. However, when sperm from one donor were used in 30 determinations during ten assays over a six-month period, "interassay" reproducibility was approximately 51%. The ELISA was compared with macroagglutination, microagglutination, and immobilization tests, using rabbit antisperm serum and human sera from vasectomized males. Results of this study indicated that the ELISA was more sensitive, less subjective, and easier to perform than these other commonly used techniques.

Mating and embryo yield of mice injected with gonadotropins on specific days of the estrous cycle and in acyclic periods.

Pregnant mare serum gonadotropin and human chorionic gonadotropin, hormones that can be used to induce superovulation, were administered to hybrid female mice (C57BL/6J X CBA/J) on known days of the estrous cycle, or when the mice were known to be acyclic, in order to determine whether certain reproductive states of the animals either enhanced or inhibited effects of the exogenous gonadotropins. Days of the estrous cycle on which the hormones were given did not significantly affect the numbers of animals that mated and in most cases did not affect embryo yield. When hormone treatment was initiated on the day of estrus in the 5-day cycle, the embryo yield was greater than when treatment was initiated on the first day of diestrus but even the difference between these groups had only marginal significance. Mice that were not exhibiting regular estrous cycles, some of which had been rendered acyclic by exposure to constant light, did not differ from mice with regular cycles in numbers that mated or in embryo yield following gonadotropin administration. When gonadotropin injection of regularly cycling mice was initiated on metestrus, on the first day of diestrus, or on estrus, the vaginal cycle was retarded as indicated by failure of cornified cells subsequently to predominate in vaginal smears at proestrus, estrus, or metestrus but initiation of hormone treatments on the second day of diestrus advanced the occurrence of vaginal cornification. Following gonadotropin injection, vaginal cornification became less reliable as a predictor of the time at which the animals would mate and ovulate.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of embryo development by some maternal sera.

Blood sera obtained from patients in an in vitro fertilization and embryo transfer program at the time of oocyte retrieval were tested for their capacities to support embryo development by using them as supplements for media in which mouse embryos were cultured. The sera varied greatly in their capacities to support development of mouse embryos but there was no correlation between the effectiveness of the sera in supporting mouse embryo development and the achievement of pregnancy by the donors of the sera. The capacities of the serum samples to support embryo development also failed to correlate with any of the causes of infertility examined (endometriosis, ureaplasma infection, pelvic adhesive disease, and bilateral tubal occlusion). It was concluded that failure of maternal sera to support embryo development does not necessarily reduce the likelihood that the donors of the sera can become pregnant by in vitro fertilization and embryo transfer procedures.