The effect of follicular fluid reactive oxygen species on the outcome of in vitro fertilization.

OBJECTIVE: To examine the presence of reactive oxygen species (ROS) in the follicular fluid of women undergoing in vitro fertilization (IVF) and identify its role in pregnancy outcome. STUDY DESIGN: In this prospective study, ROS and total antioxidant capacity (TAC) levels were measured by the chemiluminescence method in the follicular fluid of 53 women. Age, number of oocytes recovered, percentage of oocytes fertilized, ROS and TAC levels were compared in women who did and did not become pregnant. RESULTS: Patients who become pregnant had significantly higher log-transformed ROS levels (1.01 +/- 0.14, P = 0.031 than those who did not (0.69 +/- 0.08). Women with endometriosis or male factor infertility who became pregnant had significantly higher ROS levels (1.44 +/- 0.23 and 1.31 +/- 0.19) than those who did not (0.60 +/- 0.17 and 0.67 +/- 0.16; P < .006 and P < .01). CONCLUSION: Follicular fluid ROS, at low concentrations, may be a potential marker for predicting success in IVF patients.

When days are numbered: calendar structure and the development of calendar processing in English and Chinese.

Unlike English, Chinese uses a numerical system for naming months and days. This study explored whether this difference in naming affects the development of simple calendar calculation. Eight- and 10-year-old children as well as undergraduates in China and the United States were asked to name the day or month that comes a specified time before or after a given day or month. In each age group Chinese speakers primarily used calculation based on calendar names to solve these tasks, while English speakers primarily resorted to reciting the names. The magnitude of these differences was substantial; on difficult tasks Chinese fourth graders performed at speeds comparable to those of English-speaking adults. Implications for models of how linguistic structure affects cognition are discussed.

Previous fertilization failure with conventional in vitro fertilization is associated with poor outcome of intracytoplasmic sperm injection.

OBJECTIVE: To evaluate the outcome of intracytoplasmic sperm injection (ICSI) in patients with a history of fertilization failure with conventional IVF. DESIGN: Retrospective analysis of 2 years of clinical experience with ICSI. SETTING: Clinical IVF-ET program in a tertiary care referral center. PATIENT(S): The results of ICSI performed between January 1995 and December 1996 were compared between patients with normal semen parameters and a history of fertilization failure (< 20% of oocytes) with conventional IVF and patients with male factor infertility. INTERVENTION(S): In vitro fertilization with ICSI. MAIN OUTCOME MEASURE(S): Parameters examined included oocyte survival, fertilization, embryo cleavage, implantation rates, and clinical pregnancy rates. RESULT(S): Fertilization was achieved with ICSI for all patients during the study period. Although oocyte survival and fertilization did not differ between groups, the pregnancy and implantation rates for patients with a history of idiopathic fertilization failure (20% and 6%, respectively) were significantly lower than those for other patients undergoing ICSI (47% and 22%, respectively). CONCLUSION(S): The outcome of ICSI varied depending on the indication for treatment. Patients who had a history of failed or poor fertilization in vitro with apparently normal semen parameters had significantly lower pregnancy and implantation rates than did patients with either obstructive azoospermia or impaired semen quality.

Donor insemination. Prevalence of antibodies to cytomegalovirus.

OBJECTIVE: To determine the prevalence of seropositivity to cytomegalovirus in women presenting for donor insemination and in their partners. STUDY DESIGN: Retrospective examination of test results for patients presenting for donor insemination. Two hundred eighteen couples presenting for donor insemination were studied. RESULTS: Overall, 37% of women were seropositive for cytomegalovirus. Their partners had a slightly, although not significantly, lower rate of seropositivity. No clear relationship between patient age and rate of seropositivity was found. CONCLUSION: The rate of seropositivity to cytomegalovirus in couples presenting for donor insemination appears to be lower than that reported for the general population.

Preferences for intracytoplasmic sperm injection versus donor insemination in severe male factor infertility: a preliminary report.

With the advent of intracytoplasmic sperm injection (ICSI), our programme noted a drop in the number of couples using donor insemination (DI) for severe male factor infertility. Over the first 8 months in which our infertility programme offered both treatments, 27 consecutive couples scheduled for ICSI and 15 consecutive couples scheduled for DI were evaluated Since all patients in our infertility programme beginning in-vitro fertilization (IVF) with planned ICSI or starting DI undergo a semi-structured psychological interview, the psychologist's clinical notes as well as the medical chart were reviewed and coded retrospectively to determine factors related to a couple's treatment choice. Couples who chose IVF-ICSI over DI had a higher occupational status and included husbands with higher educational levels. Their most common motivation was to have the husband's biological child (93% of couples in the ICSI group). The most common motivation for choosing DI (60% of DI couples) was that IVF was not financially affordable. Choice of treatment was not related to psychological adjustment, the husband having prior biological children, or his risk of passing on a genetic defect to offspring. These preliminary data raise the concern that, with the success of ICSI, DI may change in the USA from being an option dictated by semen quality to a second choice treatment utilized for economic reasons.

Follicle size and implantation of embryos from in vitro fertilization.

OBJECTIVE: To determine the effect of increased follicle size at the time of hCG administration on the outcome of in vitro fertilization (IVF). METHODS: Fifty women immediately before 50 immediately after a protocol change were reviewed retrospectively. The criterion for hCG administration was changed from a leading follicle size of at least 17 mm to one of at least 20 mm (average of two dimensions). Changes in follicle size and number, oocytes recovered, fertilization rate, embryo development, implantation rate, and pregnancy rate were examined. Women over 40 years of age, those undergoing intracytoplasmic sperm injection, and those whose stimulation did not include GnRH agonists were excluded. RESULTS: The number of large follicles increased, but peak estradiol concentrations and number of oocytes recovered did not differ when hCG was administered at a follicular size of at least 20 mm. The numbers of oocytes fertilized, embryos replaced, and embryos cryopreserved were not statistically different. The percentage of cleaved fertilized oocytes was greater and the degree of embryo fragmentation smaller when hCG was administered at an increased follicular size. The implantation rate was higher and clinical and ongoing pregnancy rates tended to be higher when hCG was administered at a larger follicle size. CONCLUSION: The administration of hCG at a larger follicle size appears to have a beneficial effect on the outcome of IVF. Embryo quality-as reflected by decreased fragmentation, increased cleavage, and increased implantation rate-appears to be improved when hCG is delayed until two or more follicles reach at least 20 mm in diameter.

The significance of elevated progesterone at the time of administration of human chorionic gonadotropin may be related to luteal support.

PURPOSE: Our purpose was to determine if the progesterone concentration on the day of HCG administration is associated with the establishment of pregnancy in IVF patients. METHODS: Concentrations of progesterone on the day of hCG were examined retrospectively in 293 patients with ovarian hyperstimulation induced by GnRH agonist with hMG and/or FSH. Patients were grouped based on progesterone concentration: < 0.9, 0.9 to < 1.2, 1.2 to < 1.5, and > or = 1.5 ng/ml. Oocytes recovered, fertilization rate, and pregnancy rate were compared among groups. RESULTS: Patients with a higher concentration of progesterone had increased concentrations of estradiol and greater numbers of oocytes retrieved. No differences were found for fertilization rate, polyploid fertilization, or pregnancy rate. It was noted that there was an association between the type of luteal support and the effect of high preovulatory progesterone on pregnancy rate. CONCLUSIONS: These data suggest that oocytes and embryos derived from patients with high preovulatory progesterone are not of a reduced quality. The association of high progesterone concentrations with a reduced rate of pregnancy varied with the type of luteal support.

Variation in recovery of motile sperm after preparation by a simple Percoll gradient technique.

PURPOSE: Numerous techniques have been used to prepare sperm for assisted reproduction technology. Density-gradient centrifugation with Percoll is becoming a method of choice. This study reviewed the results of a simple two-layer discontinuous Percoll gradient. METHODS: We reviewed retrospectively results obtained from 208 semen specimens prepared for in vitro fertilization by a discontinuous Percoll gradient. RESULTS: Overall results (mean +/- SD) were a recovery of 21 +/- 16% of total sperm, recovery of 38 +/- 30% of motile sperm, and a motility in the final sperm preparation of 88 +/- 10%. Specimens with higher initial concentrations had higher motility in the final preparation. Higher total recovery and higher motility in the final preparation were found for specimens with a higher initial motility. CONCLUSIONS: This simple two-layer Percoll technique is rapid and inexpensive and reliably produces a final sperm preparation with desirable characteristics. Even though specimens with poor initial parameters yielded final preparations with excellent characteristics, fertilization rates were still related to the initial semen parameters.

Endometriosis: medical and surgical intervention.

Pregnancy rates with endometriosis-associated infertility may be improved by laparoscopic surgery or laparotomy for moderate to severe disease. Surgery for minimal to mild disease does not increase pregnancy rates. Medical treatment has not been shown to increase fecundity for any stage of the disease. Pregnancy rates with assisted reproductive technology for endometriosis appear to be comparable with those for tubal disease that are also treated with assisted reproductive technology. Medical and surgical treatments for pelvic pain with endometriosis are both effective, but surgery avoids the side-effects associated with drugs and may result in a lower recurrence rate.

In vitro development and implantation rates of fresh and cryopreserved sibling zygotes.

OBJECTIVE: To compare the developmental potential of cryopreserved human zygotes after thawing to sibling zygotes that were transferred without cryopreservation. METHODS: Retrospective analysis of embryo data and pregnancy outcome for all in vitro fertilization (IVF) patients who had sufficient zygotes to allow fresh embryo transfer and cryopreservation of additional sibling zygotes for later use. RESULTS: Zygotes survived cryopreservation at a high rate (87%). After thawing, cryopreserved zygotes developed at rates similar to those of fresh zygotes. Pregnancy occurred at similar rates after replacement of fresh embryos (27.9%) or replacement of cryopreserved-thawed zygote-derived embryos (24.3%). CONCLUSIONS: Human embryos can be cryopreserved at the pronuclear zygote stage with little loss of developmental potential. Cryopreservation allowed a reduction in the number of embryos transferred during an IVF retrieval cycle, thereby reducing the occurrence of multiple pregnancy. The total cumulative pregnancy rate per retrieval cycle doubled when pregnancies from cryopreserved-thawed zygotes were added to those originating from fresh zygotes.

Covering embryo cultures with mineral oil alters embryo growth by acting as a sink for an embryotoxic substance.

PURPOSE: Assisted reproductive technologies require the use of culture media for gametes and embryos. Culture media and conditions intended for use in human in vitro fertilization are tested prior to use by evaluating their ability to support normal development of mouse embryos. RESULTS: Development to blastocysts of two-cell mouse embryos was equivalent for embryos cultured in center well dishes alone (93 +/- 10%, mean +/- SD) and embryos cultured in center well dishes with an overlay of 1 ml of mineral oil (94 +/- 5%). In contrast, no development of two-cell-stage mouse embryos occurred in an embryotoxic medium when cultured in center well dishes. However, when this medium was covered with an overlay of mineral oil, development to blastocyst was no different than that of controls (96 +/- 5%). Similar results were found with cultures of one-cell-stage mouse embryos. CONCLUSION: These results indicate that in some circumstances the presence of mineral oil can significantly alter the outcome of embryo culture. Further, these results reinforce the necessity of directly testing the exact culture conditions to be used for assisted reproductive technologies.

The effect of partial zona dissection on the fertilizing capacity of chronically obstructed mouse caput epididymal sperm.

PURPOSE: Having observed that sperm from the chronically obstructed caput epididymis fertilize poorly in vitro, we investigated the effect of partial zona dissection (PZD) on fertilization in a murine model of unilateral proximal epididymal obstruction. Cleavage rates were compared for zona-intact oocytes and PZD oocytes incubated with sperm from the following epididymal segments: the obstructed caput, the contralateral nonobstructed caput, the contralateral cauda, and a sperm-free preparation to control for parthenogenesis. RESULTS: Unilateral epididymal obstruction resulted in significantly higher sperm counts in the obstructed caput compared to the nonobstructed caput, although there was no difference in motility. Cleavage rates for ova incubated with sperm from the obstructed caput and the nonobstructed caput were uniformly poor and did not differ significantly from those for the sperm-free controls. Cauda sperm fertilized significantly better than all other sperm groups (P < 0.05). Partial zona dissection did not improve cleavage rates in any group. CONCLUSION: We conclude that sperm from the chronically obstructed caput epididymis, like sperm from the normal caput, are unable to fertilize ova, and PZD does not enhance this poor fertilizing capacity. Furthermore, the finding that PZD does not improve the fertilizing capacity of the presumably mature cauda sperm in our mouse model suggests that any beneficial effect of PZD may be strain-specific.

One hundred unstable bladders.

OBJECTIVE: To investigate the correlation between presenting symptoms and cystometric data in patients with urodynamically proven detrusor instability. METHODS: A retrospective review was conducted of standardized urodynamic data bases and cystometrograms from the gynecologic urodynamics laboratories at Duke University Medical Center, Durham, North Carolina, and Grady Memorial Hospital, Atlanta, Georgia. RESULTS: Of the 100 patients reviewed, 86% complained of urge incontinence and 78% complained of urinary urgency; however, 76% also complained of stress incontinence. Statistical evaluation of symptoms by analysis of variance showed no significant correlations between the symptoms of urinary frequency and urgency and any of the cystometric indices. Patients with motor urge incontinence had significantly smaller mean cystometric bladder capacities than patients without this complaint (299.9 +/- 132.5 versus 553.6 +/- 173.6 mL; P < .01). One-third (34) of the patients had mixed incontinence due to genuine stress incontinence as well as detrusor instability. Compared with patients suffering from detrusor instability alone, patients with mixed incontinence had significantly larger cystometric bladder capacities (396.0 +/- 172.9 versus 308.1 +/- 154.8 mL; P < .02) and a lower amplitude of the maximum detrusor contraction (38.8 +/- 21.7 versus 49.9 +/- 25.1 cm H2O; P < .04). CONCLUSIONS: Women with detrusor instability represent a diverse population with a wide variety of symptoms and urodynamic findings. Patients with mixed incontinence may represent a subpopulation distinct from those with pure detrusor instability. Further research that attempts to describe other discrete subpopulations of patients with detrusor instability may improve our understanding of this troublesome clinical problem.

Transferrin- and albumin-directed expression of growth-related peptides in transgenic sheep.

Chimeric genes containing either the mouse transferrin (Trf) enhancer/promoter fused to the structural sequences encoding bovine growth hormone (GH) or the mouse albumin (Alb) enhancer/promoter fused to the gene for human growth hormone-releasing factor (GRF) were microinjected into sheep zygotes. A low percentage of resulting transgenic sheep chronically expressed the respective genes, resulting in elevated plasma concentrations of circulating GH or GRF, respectively. Growth hormone-releasing factor expression induced elevated plasma levels of endogenous GH production. In addition, elevated levels of circulating insulin-like growth factor-I were observed in the bovine GH-expressing Trf transgenic sheep. Growth of these founder transgenic sheep relative to controls were not enhanced. In part, this may be due to the development of the diabetic condition exhibited by both transgenic groups. These results demonstrate that the mouse Trf and Alb enhancer/promoters are active in sheep and suggest that alternate strategies for expressing growth-related genes may be required to modulate growth in sheep.

Effect of bovine growth hormone gene expression, sex and age on plasma gonadotropins, estrone and testosterone in prepuberal pigs.

Chronic supraphysiological blood levels of growth hormone (GH) may retard sexual maturation in swine. Pigs used in this study included four founder transgenic pigs (two gilts and two boars) expressing a mouse transferrin (TF) promoter fused to a bovine (b) GH structural gene, 13 second- or third- generation transgenic pigs (seven gilts and six boars) expressing a mouse metallothionein (MT) promoter fused to a bGH structural gene and 16 control littermates (eight gilts and eight boars). Blood plasma levels of LH, FSH, estrone and testosterone were measured to determine whether expression of bGH genes altered secretion of hormones between 80 and 180 days of age. Presence of a bGH gene was detected by hybridization of DNA in dot blots of tail biopsies. Expression of a bGH gene was detected by radioimmunoassay of plasma bGH. In four TFbGH founder transgenic pigs bGH ranged from 164 to 1948 ng/ml; in one MTbGH transgenic boar of line 3104 bGH was 1211 ng/ml; and in 12 pigs of line 3706 bGH ranged from 25 to 190 ng/ml. Expression of bGH in transgenic pigs lowered (P = .0192) plasma LH with no significant differences between sexes, had no significant effect on plasma FSH and lowered plasma estrone (P = .0001) and testosterone (P = .0269) in boars (but not gilts). Plasma estrone and testosterone were higher (P = .0001) in boars than in gilts. Plasma FSH was higher (P = .0001) in gilts than boars and decreased (P = .0001) with advancing age in gilts but not in boars.(ABSTRACT TRUNCATED AT 250 WORDS)

Immunoreactive inhibin in follicular fluid is related to meiotic stage of the oocyte during final maturation of the porcine follicle.

The concentration and content of inhibin was determined in individual porcine follicles from gilts ovariectomized at various times after the onset of estrus. In one experiment, gilts (n = 5) were ovariectomized at 0, 10, or 20 hr after the onset of estrus and the follicular fluids from all large follicles individually aspirated. In a second experiment, gilts (n = 6) were ovariectomized at 21, 24, 27, 30, or 34 hr after the onset of estrus; follicular fluids were aspirated; and each oocyte was stained and evaluated for cytogenetic stage of meiotic maturation. Inhibin was determined in diluted follicular fluids with a radioimmunoassay based on a synthetic peptide replica of part of the alpha subunit of porcine inhibin. Inhibin values were expressed in terms of thousands of units (kU) of a World Health Organization inhibin standard (86/690). Concentration of inhibin did not vary among hours (overall mean 248 kU/ml). Total follicular content of inhibin also was not different among hours (overall mean 57 kU/follicle). When follicles were classified on the basis of the maturation of the oocyte, significant differences were found. Concentration of inhibin in follicles with a germinal vesicle-stage oocyte was 138 kU/ml, whereas follicles with more mature oocytes had concentrations of between 204 and 254 kU/ml. Follicular content of inhibin showed a similar pattern with 34.9 kU/follicle at germinal vesicle stage, increasing to 42.5-56.1 kU/follicle at later stages. Quantities of inhibin were also negatively skewed and were positively correlated to follicular content of estradiol and dermatan sulfate.

Starting to add worse: effects of learning to multiply on children's addition.

A major stumbling block in acquiring a new skill can be integrating it with old but related knowledge. Learning multiplication is a case in point, because it involves integrating new relations with previously acquired arithmetic knowledge (in particular, addition). Two studies explored developmental changes in the relations between single-digit addition and multiplication. In the first study, third-graders, fifth-graders, and adults performed simple addition or multiplication in mixed- and blocked-operations formats. Substantial interfering effects from related knowledge were found at all age levels, but were more pronounced for younger subjects. Thus in the early stages of learning multiplication, one consequence of learning a new operation is interference in performance of an earlier, related, but less recently studied skill. Consideration of error patterns supported the view that the problem of integrating operations is a prominent one even in the early stages of mastering multiplication. Patterns of errors were generally consistent across all age groups, and all groups were much more likely to give a correct multiplication response to an addition problem than the reverse. A second, longitudinal study confirmed this finding, showing evidence for impaired performance of addition over time within individual children (second-, third-, and fourth-graders) tested on simple addition and multiplication over a 5-month period. Analysis of reaction times for addition indicated that second-graders in advanced math classes and third-graders in regular math classes tended to slow down over the year in responses to addition problems. Fourth-graders, on the other hand, tended to increase their speed of addition over the course of the year. Multiplication showed a different pattern during this period, with no evidence for slowing among children who were able to perform this task. Disruption of previously learned knowledge in the course of acquiring new skills provides evidence that new knowledge and old knowledge are being integrated. This kind of non-monotonic development may provide an empirical method for determining the functional limits of a domain of knowledge.

Inhibin suppresses human chorionic gonadotropin secretion in term, but not first trimester, placenta.

Inhibin is produced by the placenta, with serum concentrations rising throughout pregnancy. In contrast, hCG serum concentrations peak in the first trimester and are 80-90% lower at term. This study was designed to determine the effect of inhibin on hCG secretion both early and late in gestation. Villus tissue from 3 term and 3 first trimester (8-10 week) placentas was maintained in an in vitro explant culture model for 5 days. Tissue from each placenta was incubated with control medium in 24 replicate wells for the first 72 h. During the final 48 h, 12 wells received control medium, and 12 wells received medium containing 1% rabbit antiserum raised against the alpha-subunit (residues 1-32) of the human inhibin peptide. The antiserum demonstrated biological activity by increasing serum FSH concentrations in an immature female rat bioassay. The relative increase in hCG secretion at the conclusion of days 4 and 5 in control and antiserum-treated groups for each first trimester and term placenta were compared to pretreatment hCG concentrations on day 3. The relative increases in hCG secretion of first trimester control groups on day 4 (mean +/- SD, 34 +/- 11%) and day 5 (63 +/- 23%) were compared to those in antiserum-treated groups on day 4 (39 +/- 13%) and day 5 (54 +/- 5%) and showed no significant difference between groups on either day. The same comparison in term cultures showed the relative increases in hCG secretion of control groups on day 4 (31 +/- 10%) and day 5 (64 +/- 50%) to be significantly lower than those in antiserum-treated groups on day 4 (100 +/- 41%) and day 5 (150 +/- 108%; P less than 0.001). These findings suggest that inhibin suppresses hCG secretion in term, but not first trimester, placentas.