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Biomedical subjects

R J Aitken

Publications and source records attributed to R J Aitken.

At least 73 records · Page 4Linked to original sources
PubMed1997

The cell biology of fertilization.

Research into the cell biology of mammalian fertilization has been stimulated by the desire to provide a theoretical framework for the development of novel approaches to contraception and the need to understand the cellular basis of human infertility. The results of such studies have revealed a complex cascade of interactions initiated by the contact between capacitated spermatozoa on the oocyte-cumulus complex and culminating in sperm-oocyte fusion. In this review we shall examine our current understanding of the fertilization process, highlighting the strategic importance of recent findings and key areas where information is lacking.

Animals↗
PubMed1997

Field trial of a diluent for the transportation of human semen at ambient temperatures.

OBJECTIVE: To examine the ability of a citrate-yolk buffer extender to preserve human semen samples at ambient temperatures over a 25- to 30-hour period. DESIGN: Human semen samples were diluted 1:1 with citrate-yolk buffer or homologous seminal plasma and transported at ambient temperature between two distant locations (London to Edinburgh, United Kingdom). Various criteria of semen quality then were assessed before and after 25 to 30 hours storage and transportation in these diluents. SETTING: An institutional research laboratory and a private fertility clinic. PATIENT(S): Samples were provided by 21 donors of unknown fertility and 7 asthenozoospermic patients. INTERVENTION(S): The diluent used to preserve human semen comprised an egg yolk buffer supplemented with fructose and citrate. MAIN OUTCOME MEASURE(S): Aspects of semen quality assessed included movement, hyaluronate penetration, viability, acrosome reaction, and reactive oxygen species generation. RESULT(S): The deterioration of semen quality at ambient temperatures could be prevented by the presence of citrate-yolk buffer, permitting the accurate analysis of oxidative stress and human sperm function, 25 to 30 hours postejaculation. CONCLUSION(S): Citrate-yolk buffer offers considerable promise as a medium for the ambient temperature storage and transportation of human semen.

Acrosome↗
PubMed1997

Albumin gradients do not enrich Y-bearing human spermatozoa.

The aim of this study was to evaluate objectively whether or not discontinuous albumin gradients enrich the proportion of Y-bearing human sperm. A blinded, collaborative trial design was employed whereby a licensed centre prepared the sperm fractions using licensed procedures, coded the sperm slides and then sent them to an independent laboratory for determination of the X:Y ratio in each sperm fraction using X and Y chromosome-specific probes and double label fluorescence in-situ hybridization (FISH). The identification codes and FISH results were collated by an independent third observer. Two albumin gradient methods which are currently used by licensed centres for male sex pre-selection, protocol 3 and modified protocol 3, were tested. Essentially the same results were obtained for the two methods. Highly motile sperm fractions were recovered from the albumin gradients, and the recoveries of motile spermatozoa (1.3-8.5%) were within the optimal range reported to produce maximal enrichment of Y-bearing spermatozoa. FISH analysis, however, revealed no enrichment for Y-bearing spermatozoa with either method, and the overall X:Y ratios were not significantly different from 1.0. Some samples showed marginal enrichment of Y-bearing spermaotozoa, whereas others showed marginal enrichment of X-bearing spermaotozoa. In conclusion, this collaborative study has demonstrated that the protocol 3 and modified protocol 3 albumin gradient procedures do not enrich Y-bearing spermatozoa. The clinical use of albumin gradients for male sex preselection should be reconsidered in the light of this and other evidence.

Double-Blind Method↗
PubMed1996

The extragenomic action of progesterone on human spermatozoa is influenced by redox regulated changes in tyrosine phosphorylation during capacitation.

Capacitation had no effect on the ability of progesterone to elicit a rapid calcium transient in the acrosomal domain of human spermatozoa but had a marked influence of the ability of this steroid to induce a biological response. The development of this responsiveness to progesterone appeared to be redox regulated in that it was promoted by the stimulation of reactive oxygen species generation and inhibited by the presence of antioxidants, including catalase and membrane permeant thiols. The ability of redox conditions to influence the biological responsiveness of human spermatozoa did not involve changes in the dynamics of the calcium transients induced by progesterone but was causally linked with clear differences in tyrosine phosphorylation. We conclude that the ability of human spermatozoa to respond to the calcium transients induced by progesterone depends on a background of phosphotyrosine expression that can be profoundly influenced by the redox status of the spermatozoa during capacitation.

Humans↗
PubMed1996

Superoxide dismutase in human sperm suspensions: relationship with cellular composition, oxidative stress, and sperm function.

Sensitive techniques have been developed for monitoring superoxide dismutase (SOD) activities in human sperm preparations. In contradiction to the protective role normally assigned to SOD, populations of defective spermatozoa recovered from the low density region of Percoll gradients were found to have three times more SOD than functionally competent preparations pelleting in high density Percoll. SOD activity was negatively correlated with the movement characteristics of human spermatozoa and their capacity for oocyte fusion, and positively associated with the induction of peroxidative damage. SOD activity was also highly correlated with other markers of the cytoplasmic space, creatine kinase (CK), and glucose-6-phosphate dehydrogenase (G-6-PDH). We conclude that while SOD may play a physiological role in maintaining a balance between O2.- and H2O2, high levels of this enzyme are associated with impaired sperm function because (a) the human spermatozoon is highly susceptible to the cytotoxic effects of H2O2, (b) O2.- is an important mediator of normal sperm function, and (c) high SOD activities reflect errors in spermatogenesis associated with germ cell exfoliation and the retention of excess residual cytoplasm by the spermatozoa.

Acrosome↗
PubMed1996

Protective effect of antioxidants on the impairment of sperm motility by activated polymorphonuclear leukocytes.

OBJECTIVE: To test the ability of antioxidants to reduce the loss of sperm motility caused by reactive oxygen species generated by polymorphonuclear leukocytes (PML). DESIGN: Standardized preparations of leukocyte-contaminated semen were created by suspending known concentrations of purified spermatozoa and PML in seminal plasma. After the stimulation of reactive oxygen species generation with phorbol ester, the spermatozoa were washed and incubated in culture medium before an analysis of their movement. The ability of antioxidants to counteract the free radical-induced loss of sperm motility observed under these circumstances was assessed. SETTING: An institutional research laboratory. PARTICIPANTS: The semen was obtained from normal volunteers. INTERVENTIONS: The following were tested: vitamins C and E, dimethylsulfoxide, catalase, hypotaurine, N-acetylcysteine, and reduced glutathione. MAIN OUTCOME MEASURES: Reactive oxygen species generation was monitored by luminol-dependent chemiluminescence. Sperm motility was assessed manually and by computer-aided semen analysis. RESULTS: Consistent impairment of sperm motility and average path velocity was observed in the presence of activated PML. This effect was reduced by the concomitant presence of glutathione, N-acetylcysteine, hypotaurine, and catalase. CONCLUSION: Antioxidants can protect against the damaging effect of leukocyte-derived reactive oxygen species on sperm movement and may be of clinical value in assisted conception procedures.

Antioxidants↗
PubMed1996

Mesorectal excision for rectal cancer.

A series of 103 consecutive rectal cancers was prospectively documented. Laparotomy was performed in 78 patients of whom five did not undergo resection. Resection was considered curative unless there were liver metastases or biopsy-proven residual disease. Nine patients had a palliative resection. Thus 64 patients underwent a curative resection, 52 (81 per cent) of whom had an anterior resection of which 26 (46 per cent) were performed by trainees. There were three deaths after operation. Eight (14 per cent) patients developed a clinical anastomotic leak. Thirty patients were available for a minimum follow-up of 24 (mean 33, range 24-49) months. Four (13 per cent) patients developed distant recurrence. There were no isolated pelvic or anastomotic recurrences. One patient with distal recurrence may have had pelvic disease. The cumulative recurrence-free survival rate at 24 months was 84 per cent. These results support the suggestion that mesorectal excision may reduce local recurrence.

Adult↗
PubMed1996

Studies on the development of diluents for the transportation and storage of human semen at ambient temperature.

Storage of human semen samples at ambient temperature for 24 h resulted in a significant loss of sperm motility from a mean 45.1 +/- 1.8% to 13.8 +/- 1.1% (n = 148). This motility loss was associated with a significant increase in the osmolality of the seminal plasma and the induction of peroxidative damage to the spermatozoa. Both of these detrimental changes could be prevented by diluting the original semen sample 1:1 with a citrate-egg yolk, buffer (CYB). In the presence of this extender all aspects of semen quality were efficiently preserved for 24 h, including sperm movement, penetration of a cervical mucus substitute, the acrosome reaction and sperm-oocyte fusion. CYB extension also permitted the use of chemiluminescent tests of leukocyte contamination to be performed on semen samples stored for 24 h at ambient temperatures. As a preservation medium, CYB was found to be superior to alternative formulations lacking citrate and storage at ambient temperatures was preferable to 4 degrees C. Significant improvements in motility retention were also observed when CYB was supplemented with pentoxifylline, although this treatment significantly stimulated peroxidative damage in the spermatozoa. However, if the pentoxifylline was combined with antioxidants then this collateral peroxidative damage could be reduced and the performance of CYB significantly enhanced. These results have implications for the design of diluents permitting the long-term storage and transportation of human semen samples at ambient temperatures.

Animals↗
PubMed1996

Prediction of the in-vitro fertilization (IVF) potential of human spermatozoa using sperm function tests: the effect of the delay between testing and IVF.

To examine the diagnostic significance of several criteria of semen quality and to determine whether their prognostic value is eroded by the time interval between assessment and the attempt at in-vitro fertilization (IVF) with embryo transfer, 73 couples undergoing IVF and embryo transfer therapy were studied. The ability of human spermatozoa to achieve fertilization in vitro was examined in relation to the conventional semen profile, sperm morphology, the computer-aided assessment of sperm movement, ionophore-induced acrosome reaction, acridine orange staining, and chemiluminescent signals induced by phorbol ester and N-formyl-methionyl-leucyl-phenylalanine (FMLP). Spermatozoa were examined both in semen and after preparation on Percoll, some weeks prior to IVF. Fertilization rates were noted to be significantly correlated with elements of sperm movement characteristics, sperm morphology, and reactive oxygen species generation. Prediction of fertilization rates in a stepwise multiple regression analysis was obtained using four variables: sperm morphology, FMLP-induced chemi-luminescence and sperm movement characteristics (beat cross frequency and straightness) (r approximately 0.5). When multiple logistic regression analysis was used to predict which samples would achieve fertilization rates above and below a 50% threshold, three variables of predictive value including linearity, average path velocity and FMLP-induced chemiluminescence were selected. Combination of these variables classified the samples achieving good or poor fertilization with an overall accuracy of 83.6%. The time interval between semen assessment and IVF had little effect on the predictive value of these tests. In conclusion, the fertilizing ability of human spermatozoa is related to sperm morphology, attributes of sperm movement and reactive oxygen species production. The time delay between testing and IVF did not appear to affect predictive accuracy.

Acridine Orange↗
PubMed1996

The potential of the zona pellucida as a target for immunocontraception.

PROBLEM: To investigate the contraceptive potential of the zona pellucida. METHOD: Generation of antibodies against native and recombinant zona glycoproteins which have then been assessed for their capacity to disrupt sperm-zona interaction in vivo and in vitro. The animal model selected for these studies was the common marmoset and the end points examined were antibody titre, ovarian cyclicity and fertility. RESULTS: The fact that antibodies against the major zona glycoprotein, ZP3, block both the primary and secondary phases of sperm-zona interaction suggests that this molecule might have potential for contraceptive vaccine development. Active immunization of marmoset monkeys with native porcine ZP3 or recombinant human ZP3 produced long term infertility but also precipitated a premature decline in the primordial follicle population. CONCLUSIONS: Future studies will have to determine whether a safe, effective vaccine can be engineered by coupling unique B-cell epitopes from ZP3 to foreign T-cell antigens.

Animals↗
PubMed1996

On the use of paramagnetic beads and ferrofluids to assess and eliminate the leukocytic contribution to oxygen radical generation by human sperm suspensions.

PROBLEM: To develop a methodology to determine a) the leukocytic contribution to reactive oxygen species generation by human sperm suspensions and b) the therapeutic value of removing these cellular contaminants. METHODS: Leukocytes were removed with paramagnetic beads or colloidal ferrofluids coated with anti-CD45 antibody. The sperm suspensions were monitored for oxidant generation by chemiluminescence, leukocyte contamination by immunocytochemistry, and fertilizing potential using zona-free hamster oocytes. RESULTS: Percoll -prepared human sperm suspensions exhibited a competence for PMA-induced reactive oxygen species generation which was significantly correlated with leukocyte contamination. However, the purified spermatozoa remaining after paramagnetic bead treatment, also demonstrated an intrinsic capacity for PMA-responsive reactive oxygen species generation and, freed from the oxidative stress created by the leukocytes, exhibited a significantly enhanced capacity for sperm-oocyte fusion. CONCLUSIONS: Although human spermatozoa can generate reactive oxygen species, sperm function is inhibited by the additional oxidative stress created by contaminating leukocytes. Removal of these cells with paramagnetic beads enhances fertilizing potential.

Cell Separation↗
PubMed1996

The extragenomic action of progesterone on human spermatozoa: evidence for a ubiquitous response that is rapidly down-regulated.

Image analysis techniques have been used to demonstrate that progesterone induces a rapid calcium transient in the acrosomal domain of greater than 90% of human spermatozoa (n = 2354). These results are at variance with previous reports, suggesting that progesterone receptors are only expressed on a small subpopulation of these cells, by virtue of their ability to bind fluorescent probes incorporating progesterone 3- (O-carboxymethyl) oxime conjugated to BSA. In the present study, we could confirm that such probes only bound to a small proportion of human spermatozoa (3.01 +/- 0.29%; n = 7557) although 91.79 +/- 1.8% of the same sperm populations exhibited a calcium transient in response to progesterone. These results indicate that the binding of labeled progesterone conjugates to human spermatozoa does not reflect the size of the progesterone responsive population; the response elicited by this steroid is essentially ubiquitous. Progesterone action was shown to involve an influx of extracellular calcium via mechanisms that did not involve voltage sensitive- or second messenger operated-channels, phospholipase C, or G proteins. Despite previous evidence suggesting that progesterone action might involve a GABAA receptor/chloride channel, neither GABA nor the GABA agonist muscimol had any effect on intracellular calcium concentrations in human spermatozoa or influenced their functional competence. The only factor that disrupted the responses of human spermatozoa to progesterone was this steroid itself. Progesterone exposure induced a prolonged period of refractoriness to further stimulation that influenced the capacity of these cells to generate calcium transients, and their ability to exhibit a biological response to changes in intracellular calcium. There are implications in these results for our understanding of the extragenomic action of progesterone on human spermatozoa and the clinical manipulation of this system for the assessment and suppression of human sperm function.

Binding Sites↗
PubMed1996

An objective assessment of surgical training.

The aim of this study was to determine the feasibility of assessing surgical training from routine, prospectively collected data and to establish whether weighted workload assessed surgical training more objectively than caseload (case counting). The surgeons in this surgical unit prospectively documented details of all operations and endoscopic procedures (caseload) on a database. Over a six-month period the workload was calculated by weighting the caseload using Intermediate Equivalent (IE) values. Some 1827 procedures were documented. The three consultants performed 796 (44 per cent) procedures, the senior registrar (SR) 137 (7.5 per cent), the registrar 241 (13 per cent) and the three senior house officers (SHO) 644 (35 per cent). The consultant was first assistant in 185 (66 per cent) procedures performed by the SHOs, in 52 (61 per cent) by the registrar in 9 (13 per cent) by the SR. When assessed by caseload one SHO (as a representative example) performed 224 procedures compared to 137 by the SR. The IE workloads were 156 and 166 respectively. This better reflected the greater complexity of the operations performed by the SR. This study has shown that details of surgical training can be easily retrieved from existing administrative databases. This can be used to document the number and type of operations performed by a trainee and the degree of consultant supervision. The degree of surgical training is better assessed by weighted workload rather than caseload.

Education, Medical, Graduate↗
PubMed1996

Written discharge advice sheets reduce visits to the general practitioner.

A prospective audit of early post-operative morbidity in patients who would not normally receive routine outpatient review was undertaken. One-hundred-and forty-seven (92 per cent) of 162 patients invited returned for assessment. Thirty-five patients (24 per cent) had complications. These were of a minor nature with infected wounds being most numerous. Much of this morbidity appeared avoidable if the patients had received appropriate advice whilst in hospital. Also noted was the surprising frequency with which patients required to consult their general practitioner (GP) for guidance regarding an otherwise uncomplicated convalescence. Written advice sheets for the patients were drawn up and the study repeated. One-hundred-and-fifty (93 per cent) of 162 patients attended including 11 (7.3 per cent) who did not receive an advice sheet. Twenty-five (16.7 per cent) had complications. Although the overall complication rate was not significantly different there were significantly fewer wound infections in the second group (6 (4 per cent) versus 15 (10 per cent); p < 0.05). The number of GP visits was also reduced (24 (16.3 per cent) versus 13 (8.7 per cent); p < 0.05). Written post-operative advice sheets should be given to all patients following minor surgery.

Family Practice↗
PubMed1996

Training in surgical outpatients.

The consultants in this department, where the trainees are predominantly SHOs, organise their outpatient clinics such that one consultant sees mainly new patients (A), another mainly follow-up patients (C) and the third a mixture (B). A prospective audit was conducted to assess the impact of these different arrangements on training. Details of the training received were recorded at 32 consecutive clinics. A training episode (TE) occurred if the trainee and consultant jointly reviewed, or directly discussed, the patient in the clinic. A total of 550 patients attended and trainees were involved with 254 (46 per cent). A TE occurred in only 88 (16 per cent). The 235 (43 per cent) new patients produced 66 (28 per cent) TE and the 315 follow-up patients 22 (7 per cent) TE. Seventeen of 46 (37 per cent) procedures were a TE. The TE for the individual consultants were (new and follow-up): A 0, 8 (7 per cent); B 19 (22 per cent), 7 (8 per cent); C 47 (57 per cent), 7 (6 per cent). Outpatient training was greatly influenced by clinic organisation. Follow-up patients, who often have complex problems, rarely generate a TE. Training in surgical outpatients has not received the same attention as operative training and this deficiency needs to be addressed.

Consultants↗
PubMed1996

Comparative assessment of surgical training using objective criteria.

The aims of the study were to develop and validate an objective method of assessing the operative experience of surgical trainees. Data were retrieved from a prospectively recorded computer database of operating activity in a single surgical unit over a three-year period. Operations were weighted using intermediate equivalent (IE) values. The number of operations performed (caseload), IE workload, consultant involvement and a subjective assessment of operative ability by consultant was obtained for each of 3 SHO IIIs and 7 SHO Is. The total caseload correlated poorly with subjective grading. The total workload correlated well with the subjective assessment of the SHO IIIs and, excluding minor operations, with the SHO Is. SHO IIIs' workload consisted of a higher number of more complex procedures than SHO Is' and for both types of trainee the degree of complexity increased during the tenure of each one year post. A consultant was the assistant in 50 per cent of the intermediate and major operations performed by the SHO Is in the first quarter of each one year post dropping to 30 per cent in the remaining three-quarters. In conclusion an objective assessment was easily obtained and interpreted. Similar data from other surgical units are required to place this work in perspective and to allow formulation of guidelines.

Clinical Competence↗