Showing papers by "Medical Research Council published in 1991"

Methods for producing members of specific binding pairs

Abstract: A member of a specific binding pair (sbp) is identified by expressing DNA encoding a genetically diverse population of such sbp members in recombinant host cells in which the sbp members are displayed in functional form at the surface of a secreted recombinant genetic display package (rgdp) containing DNA encoding the sbp member or a polypeptide component thereof, by virtue of the sbp member or a polypeptide component thereof being expressed as a fusion with a capsid component of the rgdp. The displayed sbps may be selected by affinity with a complementary sbp member, and the DNA recovered from selected rgdps for expression of the selected sbp members. Antibody sbp members may be thus obtained, with the different chains thereof expressed, one fused to the capsid component and the other in free form for association with the fusion partner polypeptide. A phagemid may be used as an expression vector, with said capsid fusion helping to package the phagemid DNA. Using this method libraries of DNA encoding respective chains of such multimeric sbp members may be combined, thereby obtaining a much greater genetic diversity in the sbp members than could easily be obtained by conventional methods.

Recombinant altered antibodies and methods of making altered antibodies

Abstract: An altered antibody is produced by replacing the complementarity determining regions (CDRs) of a variable region of an immunoglobulin (Ig) with the CDRs from an Ig of different specificity, using recombinant DNA techniques. The gene coding sequences for producing the altered antibody may be produced by site-directed mutagenesis using long oligonucleotides.

Psychological Stress and Susceptibility to the Common Cold

Abstract: Background. It is not known whether psychological stress suppresses host resistance to infection. To investigate this issue, we prospectively studied the relation between psychological stress and the frequency of documented clinical colds among subjects intentionally exposed to respiratory viruses. Methods. After completing questionnaires assessing degrees of psychological stress, 394 healthy subjects were given nasal drops containing one of five respiratory viruses (rhinovirus type 2, 9, or 14, respiratory syncytial virus, or coronavirus type 229E), and an additional 26 were given saline nasal drops. The subjects were then quarantined and monitored for the development of evidence of infection and symptoms. Clinical colds were defined as clinical symptoms in the presence of an infection verified by the isolation of virus or by an increase in the virus-specific antibody titer. Results. The rates of both respiratory infection (P<0.005) and clinical colds (P<0.02) increased in a dose-response manner...

Human IgE, IgG4 and resistance to reinfection with Schistosoma haematobium

Abstract: A well recognized feature of the immune response to parasitic helminth infections, including schistosomiasis, is the production of large amounts of specific and nonspecific IgE1,2. Immunological pathways involving IgE can lead to damage to the developing schistosomulum and it has been suggested that responses involving IgE could have evolved as protection against helminth infections. There has been no epidemiological evidence to support this idea and the only significant IgE responses known in man are those involved in the pathogenesis of allergic disease. Here we measure serological response during reinfection with S. haematobium and demonstrate that IgE antibodies in man can be beneficial. Our results support the hypothesis that the slow build-up of IgE to high levels and the early production of IgG4 antibodies, which may block IgE pathways are responsible for delaying the development of protective immunity to S. haematobium.

Transcription factor AP-2 is expressed in neural crest cell lineages during mouse embryogenesis.

Abstract: We have analyzed the expression pattern of transcription factor AP-2 in mouse embryos to evaluate the potential of AP-2 as a regulator during vertebrate development. A partial cDNA encoding AP-2 was isolated from a mouse embryo cDNA library and used to prepare probes to measure AP-2 mRNA levels by RNase protection and RNA in situ hybridization. Between 10.5 and 15.5 days of embryogenesis, the relative abundance of AP-2 mRNA is greatest at 11.5 days and declines steadily thereafter. RNA in situ hybridization analysis of embryos between 8.5 and 12.5 days of gestation identified a novel expression pattern for AP-2. The principle part of this expression occurs in neural crest cells and their major derivatives, including cranial and spinal sensory ganglia and facial mesenchyme. AP-2 is also expressed in surface ectoderm and in a longitudinal column of the spinal cord and hindbrain that is contacted by neural crest-derived sensory ganglia. Additional expression of AP-2 occurs in limb bud mesenchyme and in meso-metanephric regions. This embryonic expression pattern is spatially and temporally consistent with a role for AP-2 in regulating transcription of genes involved in the morphogenesis of the peripheral nervous system, face, limbs, skin, and nephric tissues.

Humanization of a mouse monoclonal antibody by CDR-grafting: the importance of framework residues on loop conformation.

Abstract: A mouse monoclonal antibody (mAb 425) with therapeutic potential was 'humanized' in two ways. Firstly the mouse variable regions from mAb 425 were spliced onto human constant regions to create a chimeric 425 antibody. Secondly, the mouse complementarity-determining regions (CDRs) from mAb 425 were grafted into human variable regions, which were then joined to human constant regions, to create a reshaped human 425 antibody. Using a molecular model of the mouse mAb 425 variable regions, framework residues (FRs) that might be critical for antigen-binding were identified. To test the importance of these residues, nine versions of the reshaped human 425 heavy chain variable (VH) regions and two versions of the reshaped human 425 light chain variable (VL) regions were designed and constructed. The recombinant DNAs coding for the chimeric and reshaped human light and heavy chains were co-expressed transiently in COS cells. In antigen-binding assays and competition-binding assays, the reshaped human antibodies were compared with mouse 425 antibody and to chimeric 425 antibody. The different versions of 425-reshaped human antibody showed a wide range of avidities for antigen, indicating that substitutions at certain positions in the human FRs significantly influenced binding to antigen. Why certain individual FR residues influence antigen-binding is discussed. One version of reshaped human 425 antibody bound to antigen with an avidity approaching that of the mouse 425 antibody.

Fatal myocardial infarction in the Scottish adjuvant tamoxifen trial. The Scottish Breast Cancer Committee.

Abstract: OBJECTIVE--To investigate the incidence of fatal myocardial infarction in women in the two randomised arms of the Scottish adjuvant tamoxifen trial. DESIGN--Retrospective review of hospital notes to determine with the greatest possible certainty women who had died of an acute myocardial infarction. SETTING--Scottish Cancer Trials Office, the University of Edinburgh. PATIENTS--1070 postmenopausal women with operable breast cancer who were randomised to receive either adjuvant tamoxifen for five years or until relapse (539 patients) or tamoxifen for at least six weeks on the confirmation of first recurrence (531 patients). MAIN OUTCOME MEASURES--Incidence of fatal myocardial infarction in women with no known or suspected systemic cancer. RESULTS--Of the 200 women who died in the adjuvant tamoxifen arm of the trial, 44 were free of cancer at death and 10 of these died of myocardial infarction. In the observation arm 251 women died, of whom 61 showed no evidence of systemic cancer and 25 had a fatal myocardial infarction. The incidence of fatal myocardial infarction in the two groups was significantly different (chi 2 = 6.88, p = 0.0087). CONCLUSION--Tamoxifen given for at least five years as adjuvant therapy for breast cancer seems to have a cardioprotective oestrogen-like effect in postmenopausal women.

Dual task performance and processing resources in normal subjects and patients with parkinson's disease

Abstract: In recent years, there has been a growing consensus among investigators that the presence or absence of external cues guiding behaviour and attention is an important factor in determining whether or not deficits are found in patients with Parkinson's disease (PD). In an earlier study, the authors suggested that the pattern of impaired and intact performance could be explained in terms of differential resource demands of the tasks, combined with depleted levels of central processing resources in PD patients. Two experiments are reported, both employing dual-task paradigms. The first assessed, in normal subjects, the relative processing demands of a cued and an uncued version of the Stroop task. The results supported the proposal that the noncued task made greater demands on the subject's limited processing resources. Further, performing a resource demanding secondary task concurrently with the Stroop test produced, in normal subjects, the same pattern of impaired performance as that reported previously in PD patients. In the second experiment the same dual-task paradigm was employed with a group of PD patients and normal agedmatched controls. Only the patients showed an increase in reaction time on the Stroop task when performing a resource demanding secondary task. The patients also showed an interfering effect with concurrent foot tapping but not with an articulatory suppression task. The results were taken to support the hypothesis that PD patients have depleted central processing resources. In considering the present data, alternative explanations for the results are considered, in particular the possibility that they represent a deficit in switching processing resources between two tasks as the combined demands outweigh available resources.

Introduction of a subtle mutation into the hox-2.6 locus in embryonic stem cells

Abstract: Gene targeting in embryonic stem (ES) cells is a powerful tool for generating mice with null alleles. Current methods of gene inactivation in ES cells introduce a neomycin gene (neo) cassette both as a mutagen and a selection marker for transfected cells. Although null alleles are valuable, changes at the nucleotide level of a gene are very important for functional analysis. One gene family in which subtle mutations would be particularly valuable are the clusters of Hox homeobox genes. Inactivation of gene in a cluster with a neo cassette that includes promoter/enhancer elements may deregulate transcription of neighbouring genes and generate a phenotype which is difficult to interpret. We describe here a highly efficient gene targeting method, termed the 'hit and run' procedure. This generates ES cells with subtle site-specific mutations with no selectable marker and may be useful for most genes. We have developed this procedure at the hypoxanthine phosphoribosyltransferase (hprt) locus and subsequently isolated ES cells with a premature stop codon in the homeobox of Hox-2.6 (ref. 14).

Growth and activities of sulphate-reducing bacteria in gut contents of healthy subjects and patients with ulcerative colitis

Abstract: During fermentation in the human large intestine, terminal oxidative processes may involve the activities of dissimilatory sulphate-reducing bacteria (SRB). Approximately 50% of healthy individuals harbour significant populations of SRB in faeces. In mixed culture, growth of SRB in vitro was modulated by sulphate availability, with sulphated polysaccharides such as mucin, chondroitin sulphate and carrageenan causing increased growth rates and sulphide production when compared with starch, pectin and arabino-galactan. Rates of H2S production were higher among SRB isolated from patients with ulccrative colitis in contrast to those present in healthy volunteers. The majority (up to 92%) of SRB in faecal samples belonged to the genus Desulfovibrio. In vitro studies demonstrated that compared to isolates from healthy subjects. Desulfovibrio desulfuricans from colitic individuals were better able to adapt to high dilution rates, which may be associated with the disease. These findings indicate that the metabolic capabilities of SRB isolated from the human large intestine are not uniform and may respond to the type of substrate available in the gut as well as the rate of passage of digesta.

Chemotherapy in advanced ovarian cancer: An overview of randomised clinical trials

Abstract: OBJECTIVES--To consider the role of platinum and the relative merits of single agent and combination chemotherapy in the treatment of advanced ovarian cancer. DESIGN--Formal quantitative overview using updated individual patient data from all available randomised trials (published and unpublished). SUBJECTS--8139 patients (6408 deaths) included in 45 different trials. RESULTS--No firm conclusions could be reached. Nevertheless, the results suggest that in terms of survival immediate platinum based treatment was better than non-platinum regimens (overall relative risk 0.93; 95% confidence interval 0.83 to 1.05); platinum in combination was better than single agent platinum when used in the same dose (overall relative risk 0.85; 0.72 to 1.00); and cisplatin and carboplatin were equally effective (overall relative risk 1.05; 0.94 to 1.18). CONCLUSIONS--In the past, randomised clinical trials of chemotherapy in advanced ovarian cancer have been much too small to detect the degree of benefit which this overview suggests is realistic for currently available chemotherapeutic regimens. Hence a new trial comparing cisplatin, doxorubicin, and cyclophosphamide (CAP) with carboplatin has been launched and plans to accrue 2000 patients.

Properties of the two terminal oxidases of Escherichia coli.

Abstract: Proton translocation coupled to oxidation of ubiquinol by O{sub 2} was studied in spheroplasts of two mutant strains of Escherichia coli, one of which expresses cytochrome d, but not cytochrome bo, and the other expressing only the latter. O{sub 2} pulse experiments revealed that cytochrome d catalyzes separation of the protons and electrons of ubiquinol oxidation but is not a proton pump. In contrast, cytochrome bo functions as a proton pump in addition to separating the charges of quinol oxidation. E. coli membranes and isolated cytochrome bo lack the Cu{sub A} center typical of cytochrome c oxidase, and the isolated enzyme contains only 1Cu/2Fe. Optical spectra indicate that high-spin heme o contributes < 10% to the reduced minus oxidized 560-nm band of the enzyme. Pyridine hemochrome spectra suggest that the hemes of cytochrome bo are not protohemes. Proteoliposomes with cytochrome bo exhibited good respiratory control, but H{sup +}/e{sup {minus}} during quinol oxidation was only 0.3-0.7. This was attributed to an inside out orientation of a significant fraction of the enzyme. Possible metabolic benefits of expressing both cytochromes bo and d in E. coli are discussed.

Limitations of the various methods for collecting dietary intake data.

Abstract: The dietary intake of population groups may be assessed using food consumption level estimates on a national basis and household food surveys. These may be useful for monitoring secular trends and geographical differences with stable and well-documented populations, but analytical studies of diet and health require data on individuals. Assessment techniques designed to assess the diet of individuals ranging from records with weights of food to questionnaires and biological markers have been critically reviewed in order to assess the accuracy of each. Quantitative estimates of the errors involved will be given. For example, the co-efficients of variation of differences incurred from asking subjects to estimate the weight of food portions, rather than weighing them, may regularly be in the 50% range for foods and 20% for nutrients. A variety of studies suggests that the co-efficients of differences in nutrient intake estimated over 1 day from the 24-hour recall method when compared with observed intakes ranged from 4 to 400%. These errors are believed to be random, and precision can be improved by increasing the numbers of observations on each individual or by increasing the numbers of individuals within each group. However, a substantial loss of power is incurred with errors of this magnitude. A more serious potential source of error is systematic bias due either to differences between different methods of dietary assessment or from deliberate over- or underreporting by the subjects themselves. Studies with the doubly labelled water technique have suggested that substantial underrecording of food intake can occur both in free-living individuals and in athletes. 24-hour urine nitrogen can be used to validate dietary assessments in individuals in nitrogen balance, and on-going studies in Cambridge show that within a group, underreporting occurs in specific individuals rather than in the group as a whole. Independent methods of validating dietary assessments, such as the doubly labelled water technique or the 24-hour urine nitrogen output, must be included in any study of free-living individuals.

The pathophysiology of acute optic neuritis. An association of gadolinium leakage with clinical and electrophysiological deficits.

Abstract: Eighteen patients with acute optic neuritis underwent optic nerve magnetic resonance imaging (MRI) before and after injection of gadolinium-diethylene triamine pentacetic acid (Gd-DTPA). Ten were re-examined 4 wks later. Leakage of Gd-DTPA across the blood-optic nerve barrier was a consistent finding in the acute lesion, and its presence was associated with abnormal visual acuity and colour vision, retro-ocular pain on eye movement, an afferent pupillary defect, and a reduced amplitude of the P100 component of the visual evoked potential. Gd-DTPA leakage had ceased in 9/11 nerves when restudied 4 wks later, and this evolution was associated with improved visual acuity and an increased P100 amplitude. Leakage is likely to reflect inflammation, and we conclude that the latter plays an important part in the production of conduction block and clinical deficit, and that its resolution is an important step in clinical remission from acute episodes of demyelination.

Maternal and fetal influences on blood pressure.

Abstract: To study maternal and fetal influences on blood pressure in childhood 405 children aged 4 years who were born and still resident in the Salisbury health district were visited at home for blood pressure and growth measurements. Information on the pregnancy, delivery, and baby was abstracted from the routine obstetric notes. Similar to recent findings in adults, the child's systolic pressure was inversely related to birth weight and positively related to placental weight. Systolic pressure at 4 years increased by 1.2 mm Hg for every SD decrease in the ratio of head circumference to length at birth, and by 1.1 mm Hg for every SD decrease in ponderal index at birth. Mothers whose haemoglobin concentrations fell below 100g/l during pregnancy had children whose systolic pressures were on average 2.9 mm Hg higher than the children of mothers with higher haemoglobin concentrations. Patterns of placental weight, birth weight, head circumference, and length that are associated with high blood pressure in adults are also associated with higher blood pressure in 4 year old children. Identification of the intrauterine influences that lead to these patterns of fetal growth could lead to the primary prevention of hypertension.

A phorbol ester/diacylglycerol-binding protein encoded by the unc-13 gene of Caenorhabditis elegans.

Abstract: Mutations in the unc-13 gene cause diverse defects in the nervous system of the nematode Caenorhabditis elegans. Molecular cloning of the gene and sequencing of the cDNA revealed that the product encodes a protein, 1734 amino acids in length, with a central domain with sequence similarity to the regulatory region of protein kinase C. The domain was expressed in Escherichia coli and shown to bind specifically to a phorbol ester in the presence of calcium; diacylglycerol inhibited the binding in a competitive manner. These findings confirm that the unc-13 gene product has binding sites similar to those of protein kinase C and may be a component of an alternative transduction pathway of the diacylglycerol signal to a different effector function in the nervous system.

Comparison of two simple methods for determining malaria parasite density.

Abstract: Two simple methods for determining malaria parasite density which require only the examination of a thick blood film were compared with a more accurate method which involves determination of the red blood cell count and measurement of the level of infection in red blood cells by examination of a thin blood film. In one method, the number of parasites present per white blood cell is counted and this figure multiplied by 8000 (an average white blood cell count per μl) to give the parasite density. In the other method, the number of parasites present per high power microscope field is determined and the parasite density calculated from this value and the assumed volume of blood present in one high power field. The latter method proved to be more accurate than that based on determination of the parasite/white blood cell ratio, probably because the variability in the volume of blood used in preparation of thick blood films was less than the variability of the white blood cell count in the population studied.

Interacting cognitive subsystems: A systemic approach to cognitive-affective interaction and change

Abstract: Interacting Cognitive Subsytems (ICS) is a comprehensive systemic model of the organisation and function of the resources underlying human cognition. in this paper we use ICS to provide a conceptua...

The relationship between Asperger's syndrome and Kanner's autism.

Abstract: A year before Asperger's first on ‘autistic psychopathy’ appeared, Kanner (1943) published his famous first account of eleven children with a pattern of abnormal behaviour that he decided to call ‘early infantile autism’. He began as follows: ‘Since 1938, there have come to our attention a number of children whose condition differs so markedly and uniquely from anything reported so far, that each case merits – and I hope will eventually receive – a detailed consideration of its fascinating peculiarities’. The characteristics of Kanner's syndrome Kanner pointed out that these children had a number of characteristics in common. Kanner and Eisenberg (1956) selected five diagnostic criteria from Kanner's descriptions. Kanner's own words are given in quotation marks. The expansions and examples are based on his descriptions and my clinical experience. ‘A profound lack of affective contact with other people’. When young, the children appear aloof and indifferent to other people, especially other children. Kanner wrote: ‘There is, from the start, an extreme autistic aloneness that, wherever possible, disregards, ignores, shuts out anything that comes to the child from outside’. Their parents describe them as ‘self-sufficient’, ‘like in a shell’, ‘acting as if people weren't there’, ‘happiest when alone’. […]

Asperger and his syndrome

Abstract: Introductory chapter to edited book on Autism and Asperger Syndrome. Sets historical context of Asperger's work.