Human preovulatory oocytes have a higher sperm-binding ability than immature oocytes under hemizona assay conditions: evidence supporting the concept of "zona maturation"

OBJECTIVE: The purpose of this study was to investigate the sperm-binding potential of human oocytes at different stages of nuclear maturation under hemizona assay (HZA) conditions. DESIGN: This was a prospective study designed in a blinded fashion. SETTING: Academic research environment approved by the Institutional Review Board. PATIENTS: Surplus oocytes, donated by patients undergoing in vitro fertilization therapy after gonadotropin stimulation, were analyzed. Semen from a fertile donor was used in all assays. INTERVENTIONS: Five groups of oocytes were considered: (1) immature, prophase I; (2) metaphase I; (3) metaphase II; (4) inseminated, unfertilized metaphase II; and (5) immature, prophase I oocytes matured in vitro to metaphase II. Oocytes were stored in salt solution (pH 7.2) and microbisected before assay. MAIN OUTCOME MEASURE(s): Tight binding of sperm to the zona pellucida under HZA conditions was evaluated after 4 hours of gametes coincubation. RESULTS: Metaphase II oocytes (groups 3 and 4) had significantly higher binding than other groups (P = 0.0001). The mean value of the difference between the two halves (hemizona) was not significant, thus showing a small intra-assay variation for all maturational stages. CONCLUSIONS: Full meiotic competence of human oocytes is associated with an increased zona pellucida-binding potential.     

Influence of elevated pH levels on structural and functional characteristics of the human zona pellucida: Functional morphological aspects

Purpose A total of 86 fresh and salt-stored immature human oocytes derived from postmortem ovarian tissue were used for this study.Methods Oocytes were randomly incubated either in synthetic human tubal fluid medium (untreated zonae) or in a chemically defined medium (treated zonae).Results Sperm binding experiments using hemizona assay conditions exhibited a 10-fold increased binding of sperm to treated compared to untreated oocytes (272.7±43 versus 24.3±15 sperm bound, respectively; P<0.0001). pH recordings during incubation showed elevated pH levels of 8.1 compared to pH 7.2 among treated and untreated zonae, respectively. Ultrastructural examination showed a spongy appearance of the surface of treated zonae, whereas untreated zonae appeared compact with smooth surface.Conclusions The marked increase in sperm binding among treated zonae, together with the ultrastructural findings, suggest that the altered zona surface enhances sperm binding. The physiological maturational process of the zona pellucida might be manipulated in vitro, thus increasing sperm binding to the zona.Presented at the IXth World Congress on In Vitro Fertilization and Alternate Assisted Reproduction, April 3–7, 1995. Vienna, Austria.     

Impact of clinical history on fracture detection with radiography

The effect of knowledge of localizing symptoms and signs in the detection of fractures was studied. Forty radiographs of the extremities were examined twice by seven radiologists; the sessions were separated by 4 months. In 26 cases, a subtle fracture was present; 14 cases were normal. In half of the cases at each session, the precise location of pain, tenderness, or swelling was provided. The observer was asked to determine if the case was normal or abnormal (provide the exact location of the fracture) and to indicate the degree of confidence in the diagnosis. Responses were converted to a numeric scale for analysis. Analysis of receiver operator characteristic parameters indicates that clues regarding location of trauma facilitate detection of fractures. The improvement is based largely on an increased true-positive rate without an increased false-positive rate, regardless of the decision criteria of the radiologist (overall willingness to "overread" or "underread"). This has direct clinical applicability and reinforces the plea of radiologists for precise clinical information.     

Influence of clinical history upon detection of nodules and other lesions

The authors reexamined a suspected difference in the effects of clinical information upon detection of nodules vs. more diverse lesions by directly incorporating classification specificity into detection ROC analysis. Categorical prompts, correct for specific abnormalities, led to detection superior to unprompted reading when non-nodule trials (various lesion types) were analyzed. Trials that contained pulmonary nodules, or contained no lesions but had been preceded by "possible tuberculosis" or "rule out metastatic disease" prompts, failed to demonstrate the diagnostic prompt superiority. Perceptual responses may differ when nodules are compared with more complex lesions.     

Evaluation of three substitutes for Percoll in sperm isolation by density gradient centrifugation

Silane-coated silica particle solutions (ISolate(TM) and PureSperm)TM)) and iodixanol (OptiPrep(TM)) were compared to polyvinylpyrrolidone (PVP)-coated silica particles (Percoll(TM)) in their efficacy to recover spermatozoa by gradient centrifugation for use in assisted reproductive procedures. Efficacy was assessed in terms of percentages of sperm recovery, sperm vitality and motility, normal sperm morphology and normal sperm chromatin condensation. No significant difference was found in the recovery of spermatozoa for men with both normal sperm counts and oligozoospermia, between PVP-coated and silane-coated particle solutions. Iodixanol had significantly lower sperm recovery compared to the other products. Sperm vitality, progressive motility, normal morphology and normal chromatin condensation did not differ significantly between any of the sperm isolation products.      

A 10-Mb paracentric inversion of chromosome arm 2p inactivates MSH2 and is responsible for hereditary nonpolyposis colorectal cancer in a North-American kindred

Genomic deletions of the MSH2 gene are a frequent cause of hereditary nonpolyposis colorectal cancer (HNPCC), a common hereditary predisposition to the development of tumors in several organs including the gastrointestinal and urinary tracts and endometrium. The mutation spectrum at the MSH2 gene is extremely heterogeneous because it includes nonsense and missense point mutations, small insertions and deletions leading to frameshifts, and larger genomic deletions, the latter representing approximately 25% of the total mutation burden. Here, we report the identification and molecular characterization of the first paracentric inversion of the MSH2 locus known to cause HNPCC. Southern blot analysis and inverse PCR showed that the centromeric and telomeric breakpoints of the paracentric inversion map within intron 7 and to a contig 10 Mb 3' of MSH2, respectively. Pathogenicity of the paracentric inversion was demonstrated by conversion analysis. The patient's lymphocytes were employed to generate somatic cell hybrids to analyze the expression of the inverted MSH2 allele in an Msh2-deficient rodent cellular background. The inversion was shown to abolish MSH2 expression by both northern and western analysis. This study confirms that Southern blot analysis still represents a useful and informative tool to screen for and identify complex genomic rearrangements in HNPCC. Moreover, monoallelic expression analysis represents an attractive approach to demonstrate pathogenicity of unusual mutations in autosomal dominant hereditary conditions.     

The SCMC test: a reliable monitor for antispermatozoal antibodies

Seventeen couples (13%) were selected from a group of 129 infertilepatients according to the following criteria: (i) unexplainedinfertility for 3 years and (ii) <50% shaking spermatozoaduring SCMC testing. The couples were tested for sperm antibodiesafter a complete diagnostic work-up schedule. Post-coital testswere performed during the first menstrual cycle of the wife,followed by SCMC and sperm antibody titre testing. Ten malesand seven females were thus treated with 96 mg methylprednisolone.Nine (52%) of the 17 with sperm antibodies achieved a pregnancy.The results of the SCMC test were in all the cases indicativeof the actual sperm antibody titre. Reduction of the antibodytitre and a decrease in the percentage of shaking spermatozoaas detected by the SCMC test correlated well with the pregnancyrate amongst the patients.     

Multiplicity in polyp count and extracolonic manifestations in 40 Dutch patients with MYH associated polyposis coli (MAP)

Objective: To investigate the contribution of MYH associated polyposis coli (MAP) among polyposis families in the Netherlands, and the prevalence of colonic and extracolonic manifestations in MAP patients. Methods: 170 patients with polyposis coli, who previously tested negative for APC mutations, were screened by denaturing gradient gel electrophoresis and direct sequencing to identify MYH germline mutations. Results: Homozygous and compound heterozygous MYH mutations were identified in 40 patients (24%). No difference was found in the percentage of biallelic mutation carriers between patients with 10–99 polyps or 100–1000 polyps (29% in both groups). Colorectal cancer was found in 26 of the 40 patients with MAP (65%) within the age range 21 to 67 years (median 45). Complete endoscopic reports were available for 16 MAP patients and revealed five cases with gastro-duodenal polyps (31%), one of whom also presented with a duodenal carcinoma. Breast cancer occurred in 18% of female MAP patients, significantly more than expected from national statistics (standardised morbidity ratio = 3.75). Conclusions: Polyp numbers in MAP patients were equally associated with the attenuated and classical polyposis coli phenotypes. Two thirds of the MAP patients had colorectal cancer, 95% of whom were older than 35 years, and one third of a subset of patients had upper gastrointestinal lesions. Endoscopic screening of the whole intestine should be carried out every two years for all MAP patients, starting from age 25–30 years. The frequent occurrence of additional extraintestinal manifestations, such as breast cancer among female MAP patients, should be thoroughly investigated.