Meiotic nondisjunction and sperm aneuploidy in humans.

Infertility is relatively common affecting approximately 1-in-6 couples. Although the genetic basis of infertility is increasingly being uncovered, the contribution of male infertility often remains unexplained. The leading cause of pregnancy loss and cognitive impairment in humans is chromosome aneuploidy. Sperm aneuploidy is routinely evaluated by fluorescence in situ hybridization. The majority of studies have reported similar findings, namely: (1) all men produce aneuploid sperm; (2) certain chromosomes are more prone to undergo chromosome nondisjunction; (3) infertile men typically have significantly higher levels of sperm aneuploidy compared to controls and (4) the level of aneuploidy is often correlated with the severity of the infertility. Despite this, sperm aneuploidy screening is rarely evaluated in the infertility clinic. Within recent years, there appears to be renewed interest in the clinical relevance of sperm aneuploidy. We shall examine the gender differences in meiosis between the sexes and explore why less emphasis is placed on the paternal contribution to aneuploidy. Increased sperm aneuploidy is often perceived to be modest and not clinically relevant, compared to the female contribution. However, even small increases in sperm aneuploidy may impact fertility and IVF cycle outcomes. Evidence demonstrating the clinical relevance of sperm aneuploidy will be discussed, as well as some of the challenges precluding widespread clinical implementation. Technological developments that may lead to widespread clinical implementation will be discussed. Recommendations will be suggested for specific patient groups that may benefit from sperm aneuploidy screening and whether preimplantation genetic testing for aneuploidy should be discussed with these patients.

Hierarchical radial and polar organisation of chromosomes in human sperm.

It is well established that chromosomes occupy distinct positions within the interphase nuclei, conferring a potential functional implication to the genome. In addition, alterations in the nuclear organisation patterns have been associated with disease phenotypes (e.g. cancer or laminopathies). The human sperm is the smallest cell in the body with specific DNA packaging and the mission of delivering the paternal genome to the oocyte during fertilisation. Studies of nuclear organisation in the sperm have postulated nonrandom chromosome position and have proposed a chromocentre model with the centromeres facing toward the interior and the telomeres toward the periphery of the nucleus. Most studies have assessed the nuclear address in the sperm longitudinally predominantly using centromeric or telomeric probes and to a lesser extent with whole chromosome paints. To date, studies investigating the radial organisation of human sperm have been limited. The purpose of this study was to utilise whole chromosome paints for six clinically important chromosomes (18, 19, 21, 22, X, and Y) to investigate nuclear address by assessing their radial and longitudinal nuclear organisation. A total of 10,800 sperm were analysed in nine normozoospermic individuals. The results have shown nonrandom chromosome position for all chromosomes using both methods of analysis. We present novel radial and polar analysis of chromosome territory localization within the human sperm nucleus. Specifically, a hierarchical organisation was observed radially with chromosomes organised from the interior to the periphery (chromosomes 22, 21, Y, X, 19, and 18 respectively) and polar organisation from the sperm head to tail (chromosomes X, 19, Y, 22, 21, and 18, respectively). We provide evidence of defined nuclear organisation in the human sperm and discuss the function of organisation and potential possible clinical ramifications of these results in regards to male infertility and early human development.

Sperm aneuploidy frequencies analysed before and after chemotherapy in testicular cancer and Hodgkin's lymphoma patients.

BACKGROUND Multicolour fluorescent in situ hybridization was utilized to detect sperm aneuploidy for chromosomes 13, 21, X and Y in testicular cancer and Hodgkin's lymphoma chemotherapy patients. METHODS Aneuploidy was assessed before, and 6, 12 and/or 18-24 months after, the initiation of chemotherapy, and compared with age matched controls. 635 396 sperm were scored blindly with 5000 sperm/patient/chromosome/ time point, where sperm was available. (First two phrases have been reversed). RESULTS Comparing testicular cancer and Hodgkin's lymphoma patients to each other and with controls, cancer-specific differences were identified. Hodgkin's lymphoma patients, particularly, exhibited significantly increased aneuploidy frequencies for all chromosomes throughout treatment. At 6 months, all cancer patients showed significantly increased frequencies of XY disomy and nullisomy for chromosomes 13 and 21. In general, aneuploidy frequencies declined to pretreatment levels 18 months after treatment initiation, but increased aneuploidy frequencies persisted in some chromosomes for up to 24 months. CONCLUSIONS Because of elevated aneuploidy frequencies prior to and up to 24 months from the start of chemotherapy, patients should receive genetic counselling about the potentially increased risk of an aneuploid conceptus from sperm cryopreserved prior to chemotherapy, and for conceptions up to 2 years after the initiation of treatment.

The role of sperm aneuploidy as a predictor of the success of intracytoplasmic sperm injection?

BACKGROUND: We present the first powered prospective study to assess whether sperm aneuploidy can predict the outcome of ICSI. METHODS: Our null hypothesis was that aneuploidy rates (AR) are identical in men who achieve successful (Group A) and unsuccessful (Group B) ICSI outcome. A power calculation yielded a sample number of 56 to achieve 80% power to reject our hypothesis at the 5% significance level. Samples for testing were obtained on the day of embryo transfer and tests were performed on raw pre-preparation samples. Sperm AR of chromosomes 13, 18, 21, X/Y were assessed using fluorescence in-situ hybridization (FISH) techniques (mean of 1223 sperm). RESULTS: There was no significant difference in any patient, seminal, cycle or laboratory characteristic between groups that may have affected outcome. Total AR (2.37 versus 1.18%, P = 0.01), as well as AR of chromosomes 18, X/Y and 18 + X/Y (1.48 versus 0.67%, P = 0.005) were significantly higher in Group B compared with Group A. Regression analysis confirmed these differences to be independent of other variables and showed a 2.6-fold change in odds of achieving a pregnancy for every 1% change in total AR. CONCLUSIONS: Our findings confirm a potential role for aneuploidy testing in the work-up of ICSI patients as a predictor of success, as well as in future genetic counselling. If confirmed, there may also be a place for a study of preimplantation genetic screening to improve ICSI success in men found to have high AR and ICSI failure.

Practicable approaches to facilitate rapid and accurate molecular cytogenetic mapping in birds and mammals.

Molecular cytogenetic mapping by FISH is a common feature of most genome projects as it provides a global, low-resolution overview of the genome and facilitates comparative genomics. An essential prerequisite for cytogenetic mapping is the ability to identify accurately the chromosome on which the clone (e.g. BAC) resides. This is not usually a barrier to human mapping as knowledge of the human karyotype is commonplace. For other species however accurate assignment can be problematic either because, as in birds, the karyotype is too complex to analyze by standard means or because of the paucity of individuals skilled to perform the karyotyping. Using chicken as a model we have developed a reproducible approach for accurate cytogenetic mapping that involves: a single colour FISH, measurement of the ratio of the size of the signal bearing chromosome to that of chromosome 8, and final assignment through a small series of dual colour experiments. Reference values for size ratios were established using base pair estimate information from the Ensembl browser. By this method cytogenetic mapping to highly complex karyotypes can be achieved in a small number of simple steps. We have also developed and tested a karyotyping tutorial programme adapted from one previously reported in this journal. That is, we have used pig as an example of a model species with a relatively tractable karyotype and demonstrated that scientists and students, even after only one hour using our tutorial, can readily identify pig chromosomes and thus make appropriate assignments using FISH. Simple, practicable means often provide preferable solutions than complex alternatives (e.g. m-FISH) to the solution of scientific problems. Such is the case for the approaches described here.

The evolution of the avian genome as revealed by comparative molecular cytogenetics.

Birds are characterised by feathers, flight, a small genome and a very distinctive karyotype. Despite the large numbers of chromosomes, the diploid count of 2n approximately 80 has remained remarkably constant with 63% of birds where 2n = 74-86, 24% with 2n = 66-74 and extremes of 2n = 40 and 2n = 142. Of these, the most studied is the chicken (2n = 78), and molecular cytogenetic probes generated from this species have been used to further understand the evolution of the avian genome. The ancestral karyotype is, it appears, very similar to that of the chicken, with chicken chromosomes 1, 2, 3, 4q, 5, 6, 7, 8, 9, 4p and Z representing the ancestral avian chromosomes 1-10 + Z; chromosome 4 being the most ancient. Avian evolution occurred primarily in three stages: the divergence of the group represented by extant ratites (emu, ostrich etc.) from the rest; divergence of the Galloanserae (chicken, turkey, duck, goose etc.)--the most studied group; and divergence of the 'land' and 'water' higher birds. Other than sex chromosome differentiation in the first divergence there are no specific changes associated with any of these evolutionary milestones although certain families and orders have undergone multiple fusions (and some fissions), which has reduced their chromosome number; the Falconiformes are the best described. Most changes, overall, seem to involve chromosomes 1, 2, 4, 10 and Z where the Z changes are intrachromosomal; there are also some recurring (convergent) events. Of these, the most puzzling involves chromosomes 4 and 10, which appear to have undergone multiple fissions and/or fusions throughout evolution - three possible hypotheses are presented to explain the findings. We conclude by speculating as to the reasons for the strange behaviour of these chromosomes as well as the role of telomeres and nuclear organisation in avian evolution.

Patients' preference for gender of urologist.

The NHS Plan is for a health service designed around the patient. Several studies have investigated patients' preference for the gender of their doctor. However, within these studies, the patients have predominantly been females. The preferences of urology patients, of whom the majority are males, have not previously been investigated. The preferences for gender of urologist, from 496 consecutive patients attending urology clinic in a district general hospital, were investigated by questionnaires. The majority of patients had no preference for gender of their urologist. Of the patients who did have a preference, females were more likely to prefer the same gender urologist than males.

The relationship between male infertility and increased levels of sperm disomy.

Sperm chromosome abnormalities cut across a number of areas relevant to ICC XV. The association between increased levels of sperm aneuploidy (usually disomy) and male infertility has implications for the sessions on reproduction, sex chromosomes, aneuploidy and meiosis and was, to the best of our knowledge, first reported in 1995. Since then most studies have reported similar increases of varying degrees but, despite this, a small number of laboratories have presented results that demonstrate no significant association. The purpose of this article is to review the state of the art in this area and to speculate as to reasons for the differences in reports from different laboratories. The findings are broken down by chromosome with studies of the sex chromosomes being broken down further to indicate meiotic stages of origin. We conclude that comparisons are difficult to make since many studies do not clearly define patient and control groups. Nevertheless, despite these and other differences (such as scoring criteria, technical differences, demographics, etc.), the consensus in the literature is that a strong correlation exists between sperm aneuploidy and male infertility. The nature of that relationship will be further defined when andrological criteria are more closely taken into account and protocols for preparation and scoring are standardised.

P2X and P2X receptor expression in human bladder urothelium and changes in interstitial cystitis.

OBJECTIVE: To investigate whether the expression of P2X(3) receptors (implicated in the pathophysiology of pain) is altered in human bladder urothelium from patients with interstitial cystitis (IC, a major symptom of which is pain), and as P2X(2) receptors can be co-expressed with P2X(3) receptors, to assess their expression also. PATIENTS AND METHODS: Bladder tissue samples were collected from patients undergoing cystectomy or radical prostatectomy. Patients with IC were diagnosed using the international criteria. RNA protein expression levels of both receptors were evaluated using reverse transcription-polymerase chain reaction (PCR), real-time quantitative PCR and Western blot analysis. RESULTS: P2X(2) was expressed in the human urothelium, in a glycosylated form. There was less gene expression of P2X(3) in IC urothelium, whereas P2X(2) gene expression was unchanged. This contrasted with the protein expression, which was increased for both P2X(2) and P2X(3). CONCLUSION: This is the first report of the expression of the P2X(2) receptor in human bladder urothelium. There was greater protein expression of both P2X(2) and P2X(3) in IC bladder urothelium which did not directly correlate with the gene expression. Changes in expression of P2X(2) and P2X(3) receptors may contribute to the pain that patients with IC have, and might provide novel drug targets.

Safety issues in assisted reproduction technology: Should men undergoing ICSI be screened for chromosome abnormalities in their sperm?

The incidence of aneuploidy in gametes of men undergoing ICSI has raised the prospect of there being risks associated with ICSI and the question of whether or not to screen men for sperm aneuploidy before treatment. We report results of a questionnaire undertaken to address how IVF staff perceive this problem, whether ICSI men are already being screened for sperm aneuploidy and the extent to which IVF specialists feel that there is merit in such a test. The results suggest that this is seen as a problem but most feel the risks outweigh the benefits. Most claimed their clinics do not screen sperm for aneuploidy but feel that there is merit in doing so. There are considerable benefits to screening i.e. couples would get additional information about the genetic repercussions of ICSI and could make informed decisions before treatment; screening would also facilitate the design of a large research study to give clearer answers on the safety of ICSI. However, we acknowledge counter arguments i.e. families would not necessarily benefit as most would have the ICSI procedure regardless of screen results; sex chromosome trisomies clinically are not severe enough to worry about in this context and there are other potential risks of ICSI that screening would not address.

Sequential organ scoring as a measure of effectiveness of care in the high-dependency unit.

High-dependency units are of benefit to patients and to the associated intensive care unit. However, the effectiveness of high-dependency care has not been quantified. We studied 100 consecutive patients whose high-dependency unit admission was longer than 48 h. The Logistic Organ Dysfunction System was used to assess organ dysfunction on a daily basis in these patients. The median Logistic Organ Dysfunction System score on admission was 3. The mean duration of stay was 4.3 days, and the median Logistic Organ Dysfunction System score on day 4 was 1. Sixty-six per cent of patients had a lower Logistic Organ Dysfunction System score at discharge. In the remaining patients, the commonest reason for no change or a higher Logistic Organ Dysfunction System score was renal dysfunction. High-dependency care is effective in the majority of patients in terms of improved physiological status and most patients were discharged with minor physiological dysfunction. This level of physiological dysfunction could be applied as a discharge threshold. Poorer patient outcome is associated with increased age and a longer stay on the unit. This poor outcome was commonly due to renal system dysfunction, and invariably followed surgery.

Embryo brain kinase: a novel gene of the eph/elk receptor tyrosine kinase family.

A new gene belonging to the Eph/Eck/Elk receptor tyrosine kinase family has been cloned from mouse brain. The gene maps to mouse chromosome 4. In the adult brain it is expressed exclusively and abundantly in the hippocampus. We propose to name it Ebk (embryo brain kinase), as in situ hybridisation shows expression in many parts of the developing mouse brain. The most abundant expression is in the subcommissural organ, and the earliest expression is in the forebrain neural folds, in rhombomeres 2-6, and in somites and heart. Other regions positive at various stages include the cochlear duct, trigeminal ganglion, lung, first branchial arch, and tooth primordia. Also positive are areas of mesenchyme underlying various epithelia during morphogenesis, especially in the mouth and nose, as well as in the eyelids and toes. We compare these patterns with the available data on the 12 other known members of this gene family. Most of them, like Ebk, are expressed in brain (especially adult hippocampus and embryonic rhombomeres) and in organs rich in epithelia (especially lung), although the spatial and temporal patterns differ. We suggest that combinatorial patterns of these receptors act as labels for the regional identity of neurons and epithelia, and could mediate fine control of neurite pathfinding and epithelial morphogenesis.