Biomedical subjects

D H Wyllie

Publications and source records attributed to D H Wyllie.

10 recordsLinked to original sources

PubMed2005

Functional mapping of Toll/interleukin-1 signalling networks by expression cloning.

Multiple cellular proteins have been identified as participating in Toll/interleukin-1 receptor-mediated inflammatory gene expression. The continuing isolation of novel components, based on sequence similarities, protein-protein interactions and protein purification, suggests that many elements of this signalling network remain to be identified. We report here the development of a high-throughput functional screening platform and its application for the identification of components of inflammatory signalling networks. Our results enable us to estimate that 100-150 gene products are involved in controlling the transcription of the human interleukin 8 gene. The approach, which is simple and robust, constitutes a general method for mapping signal transduction systems and for rapid isolation of a large number of signalling components based on the control of pathways leading to regulation of gene expression.

Cloning, Molecular↗

PubMed2005

Bacteraemia prediction in emergency medical admissions: role of C reactive protein.

AIM: To define the contribution made by C reactive protein (CRP) measurement to bacteraemia prediction in adults with medical emergencies in the UK. METHODS: This two year cohort study involved 6234 patients admitted as emergency cases to the acute medical or infectious diseases services of the Oxford Radcliffe Hospitals, in whom blood cultures were taken on arrival. The main outcome measures were bacteraemia risk associated with admission CRP concentrations, lymphocyte counts, and neutrophil counts. RESULTS: The quantitative associations between CRP concentration, admission lymphocyte count, and neutrophil count were defined. Risk of bacteraemia rose continuously as the CRP increased: no "cutoff" value was evident. Models examining combinations of CRP, neutrophil count, and lymphocyte count were developed and validated using a split sample technique. CRP contributed to a model including lymphocyte and neutrophil counts, but its effect was small. CRP alone performed no better than either a model combining lymphopenia and neutrophilia, or than lymphopenia alone. CONCLUSIONS: In patients with acute medical emergencies who are suspected of bacteraemia clinically, CRP concentrations, although associated with bacteraemia, have a limited role in bacteraemia prediction.

Acute Disease↗

PubMed2004

Relation between lymphopenia and bacteraemia in UK adults with medical emergencies.

AIMS: To determine the relevance of lymphopenia to the diagnosis of bacteraemia in patients admitted with medical emergencies, relative to peripheral blood white cell count and neutrophilia. PATIENTS/METHODS: A two year cohort study carried out in a teaching hospital in Oxford, UK of 21,495 consecutive adult emergency admissions to general medical or infectious disease wards. Full blood data were available in 21,372 cases; 41 cases with extreme full blood count results (neutrophil count, > 75 x 10(9)/litre; lymphocyte count, > 10 x 10(9)/litre) were excluded, leaving 21,331 cases for analysis. The association between the admission lymphocyte and neutrophil counts and the risk of bacteraemia was assessed. RESULTS: Neutrophilia and lymphopenia were both associated with bacteraemia. Lymphopenia was the better predictor in this cohort. Both neutrophilia and lymphopenia were more predictive of bacteraemia than the total white blood cell count. CONCLUSIONS: Both lymphocyte and neutrophil counts, rather than total white blood cell count, should be considered in adult medical admissions with suspected bacteraemia.

Aged↗

PubMed2001

Regulation of Toll-like receptors in human monocytes and dendritic cells.

A number of pathogens induce immature dendritic cells (iDC) to migrate to lymphoid organs where, as mature DC (mDC), they serve as efficient APC. We hypothesized that pathogen recognition by iDC is mediated by Toll-like receptors (TLRs), and asked which TLRs are expressed during the progression of monocytes to mDC. We first measured mRNA levels for TLRs 1-5 and MD2 (a protein required for TLR4 function) by Northern analysis. For most TLRs, message expression decreased severalfold as monocytes differentiated into iDC, but opposing this trend, TLR3 and MD2 showed marked increases during iDC formation. When iDC were induced to mature with LPS or TNF-alpha, expression of most TLRs transiently increased and then nearly disappeared. Stimulation of iDC, but not mDC, with LPS resulted in the activation of IL-1 receptor-associated kinase, an early component in the TLR signaling pathway, strongly suggesting that LPS signals through a TLR. Surface expression of TLRs 1 and 4, as measured by mAb binding, was very low, corresponding to a few thousand molecules per cell in monocytes, and a few hundred or less in iDC. We conclude that TLRs are expressed in iDC and are involved in responses to at least one pathogen-derived substance, LPS. If TLR4 is solely responsible for LPS signaling in humans, as it is in mice, then its extremely low surface expression implies that it is a very efficient signal transducer in iDC.

Antigens, Surface↗

PubMed2001

Toll receptors and sepsis.

Toll-like receptors are a family of receptors that recognize components of bacteria and induce a proinflammatory response by cells, including macrophages and endothelial cells. Ten human Toll receptors differing in their specificity for microbial components have been cloned. They respond to various components, including lipopolysaccharide of Gram-negative bacteria, lipopeptides of Gram-positive cell walls, bacterial DNA, and flagella. Some Toll-like receptors require the cooperation of an adapter protein. Toll-like receptor 4 function requires the presence of the protein MD2. Recently, it has been shown that Toll-like receptors function cooperatively to increase the specificity of response to a given microbe. Human polymorphisms of Toll-like receptor genes have been discovered and are associated with hyporesponsiveness to bacterial components.

Animals↗

PubMed2000

Evidence for an accessory protein function for Toll-like receptor 1 in anti-bacterial responses.

Members of the Toll-like receptor (TLR) family are components of the mammalian anti-microbial response, signaling with a domain closely related to that of IL-1 receptors. In this report the expression and function of TLR1, a TLR of unknown function, are examined. TLR1 is expressed by monocytes, as demonstrated using a novel mAb. Monocytes also express TLR2. TLR1 transfection of HeLa cells, which express neither TLR1 nor TLR2, was not sufficient to confer responsiveness to several microbial extracts. However, cotransfection of TLR1 and TLR2 resulted in enhanced signaling by HeLa cells to soluble factors released from Neisseria meningitidis relative to the response with either TLR alone. This phenomenon was also seen with high concentrations of some preparations of LPS. The N. meningitidis factors recognized by TLR1/TLR2 were not released by N. meningitidis mutant in the LpxA gene. Although LpxA is required for LPS biosynthesis, because cooperation between TLR1 and TLR2 was not seen with all LPS preparations, the microbial component(s) TLR1/2 recognizes is likely to be a complex of LPS and other molecules or a compound metabolically and chemically related to LPS. The functional IL-1R consists of a heterodimer; this report suggests a similar mechanism for TLR1 and TLR2, for certain agonists. These data further suggest that mammalian responsiveness to some bacterial products may be mediated by combinations of TLRs, suggesting a mechanism for diversifying the repertoire of Toll-mediated responses.

Acyltransferases↗

PubMed2000

A lipopolysaccharide-deficient mutant of Neisseria meningitidis elicits attenuated cytokine release by human macrophages and signals via toll-like receptor (TLR) 2 but not via TLR4/MD2.

Meningococcal disease severity correlates with circulating concentrations of lipopolysaccharide (LPS) and proinflammatory cytokines. Disruption of the lpxA gene of Neisseria meningitidis generated a viable strain that was deficient of detectable LPS. The potency of wild-type N. meningitidis to elicit tumor necrosis factor (TNF)-alpha production by human monocyte-derived macrophages was approximately 10-fold greater than that of the lpxA mutant. Killed wild-type N. meningitidis and its soluble products induced interleukin (IL)-8 and TNF-alpha secretion by transfected HeLa cells expressing Toll-like receptor (TLR) 4/MD2, but the lpxA mutant was inactive via this pathway. In contrast, both strains induced IL-8 promoter activity in TLR2-transfected HeLa cells. These data provide evidence that N. meningitidis contains components other than LPS that can elicit biological responses via pathways that are independent of the TLR4/MD2 receptor system, and TLR2 is one of these alternate pathways. These findings have implications for future therapeutic strategies against meningococcal disease on the basis of the blockade of TLRs and the modulation of LPS activity.

Acyltransferases↗

PubMed2000

A novel mammalian expression screen exploiting green fluorescent protein-based transcription detection in single cells.

The accumulation of DNA sequence information from large-scale genomic and random library sequencing projects is leading to the rapid identification of many putative genes, virtual transcripts and ESTs of unknown function. There is therefore an increasing need for high throughput, sensitive and robust methods for identification and characterisation of genes, and/or their products, based on function. We describe a high throughput functional expression screen based on semi-quantitative analysis of enhanced green fluorescent protein expression in single cells by confocal microscopy. The assay was implemented in a micro-scale format, requiring around 10(4) cells/test. The system was validated by co-transfection of a series of cDNAs encoding pro-inflammatory cytokine intracellular signal mediators with a d2EGFP reporter containing a cytokine responsive promoter. The majority of the test plasmids gave a detectable signal above background at a pool size of 250-500. Replicate tests indicate that the assay is reproducible at this pool size. At this level we demonstrate that large (>10(6) transformants) libraries can be feasibly screened.

Adaptor Proteins, Signal Transducing↗

PubMed1992

Measuring change in surgical practice.

We reviewed the work of a single general surgical firm over six years--about 11,500 patient episodes. Although the workload remained approximately constant, a severe reduction in the number of available beds was accompanied by a marked change towards shorter duration of stay. This affected particularly the elective cases. Previous authors, when trying to predict requirement of hospital beds, have accepted that duration of stay and number of beds available are independent variables; this was not what we observed.

Bed Occupancy↗

PubMed1988

Pursuing efficiency in surgical practice.

To examine fluctuations in numbers of patients on surgical wards the dates of admission from January of each of the 5556 patients admitted from 1 January 1985 to 31 December 1987 were examined during computerised audit of a single surgical firm. The numbers of patients under the care of the firm fluctuated widely, often exceeding the 38 beds nominally available. Duration of stay varied from two days or less (3062 admissions) to more than a month (163 admissions). One patient was in hospital for 278 days. The patients admitted for more than a month (2.9% of the total) filled 28% of the beds; not all these patients were elderly. A further increase in throughput of patients undergoing elective operations might be achieved by always admitting patients on the day of operation, and perhaps by discharging patients even sooner than at present. Efficiency would increase but so would overall costs.

Aged↗