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Biomedical subjects

C Williams

Publications and source records attributed to C Williams.

At least 109 records · Page 6Linked to original sources
PubMed2000

Analysis of p53 mutations in single cells obtained from histological tissue sections.

We have previously reported on direct sequence analysis of the p53 gene in laser-dissected single cells from tissue sections, where each allele of two fragments (exons 7 and 8) could be accurately analyzed in only 14% of the cells due to the high frequency of exon and allele dropout. Here in an effort to minimize this problem, we have investigated various approaches for sample preparation and gene amplification. By pinpointing some critical steps in the procedure, we could increase the number of investigated exons and substantially improve the genetic analysis of single cells obtained from histochemically stained frozen tissue sections. The biggest improvement was achieved by minimizing DNA degradation using EDTA as a nuclease inhibitor in all sample preparation steps. Efforts to increase primer annealing, by increasing the concentration of template and primers, in addition to prolonging the annealing and extension times, also improved the amplification efficiency. With these measures we can now amplify six individual exons of the p53 gene (exons 4-9) in 70% of the cells and in 50% of these cells both alleles are amplified. This allows application of the method in various investigations such as within the field of tumor pathology.

Alleles↗
PubMed2000

Cold collision frequency shifts in a 87Rb atomic fountain.

We present measurements of cavity frequency pulling and collisional frequency shifts in a 87Rb fountain with a frequency resolution of 3x10(-16). Agreement with theory is found for the cavity pulling and the measured collisional shifts. The clock shift is found at least 50 times smaller than in 133Cs.

Journal Article↗
PubMed2000

Germline APC variants in patients with multiple colorectal adenomas, with evidence for the particular importance of E1317Q.

Mendelian tumour syndromes are caused by rare mutations, which usually lead to protein inactivation. Few studies have determined whether or not the same genes harbour other, more common variants, which might have a lower penetrance and/or cause mild disease, perhaps indistinguishable from sporadic disease and accounting for a considerable proportion of the unexplained inherited risk of tumours in the general population. Germline variants at the APC locus are excellent candidates for explaining why some individuals are predisposed to colorectal adenomas, but do not have the florid phenotype of familial adenomatous polyposis. We have screened 164 unrelated patients with 'multiple' (3-100) colorectal adenomas for germline variants throughout the APC gene, including promoter mutations. In addition to three Ashkenazi patients with I1307K, we found seven patients with the E1317Q variant. E1317Q is significantly associated with multiple colorectal adenomas (OR = 11. 17, 95% CI = 2.30-54.3, p < 0.001), accounting for approximately 4% of all patients with multiple colorectal adenomas. In addition, four patients with truncating APC variants in exon 9 or in the 3' part of the gene were identified. Germline APC variants account for approximately 10% of patients with multiple adenomas. Unidentified predisposition genes almost certainly exist. We argue that it is worthwhile to screen multiple adenoma patients for a restricted number of germline APC variants, namely the missense changes E1317Q and I1307K (if of Ashkenazi descent), and, if there is a family history of colorectal tumours, for truncating mutations 5' to exon 5, in exon 9 and 3' to codon 1580.

Adenoma↗
PubMed2000

Hanging gondola structure of the T1 domain in a voltage-gated K(+) channel.

The T1 domain is a approximately 100-residue sequence in the cytoplasmic N-terminal region of K(v)-type K(+) channels. The structure of the isolated domain is known, but it is uncertain whether the structure of this domain is maintained in the fully assembled, membrane-associated, homotetrameric channel protein. We use the structure of the isolated domain as a guide for designing disulfide bonds to cross-link Shaker K(+) channels through the T1 domain. Six pairs of residues with side chains closely apposed across the T1 subunit interface were selected for replacement by cysteine. Of these, three pairs formed cross-links upon air oxidation of cysteine-substituted Shaker channels expressed in Xenopus oocyte membranes. Two of these cross-linked channels were examined electrophysiologically and were found to have gating properties only slightly altered from wild-type. The results show that the structure of the isolated T1 domain exists in the mature ion channel. They also demand that this domain is attached to the membrane-embedded part of the protein as a cytoplasmic "hanging gondola", and that ions gain access to the pore through four "windows" formed by the linker connecting T1 to the channel's first transmembrane helix.

Animals↗
PubMed2000

Molecular portraits of human breast tumours.

Human breast tumours are diverse in their natural history and in their responsiveness to treatments. Variation in transcriptional programs accounts for much of the biological diversity of human cells and tumours. In each cell, signal transduction and regulatory systems transduce information from the cell's identity to its environmental status, thereby controlling the level of expression of every gene in the genome. Here we have characterized variation in gene expression patterns in a set of 65 surgical specimens of human breast tumours from 42 different individuals, using complementary DNA microarrays representing 8,102 human genes. These patterns provided a distinctive molecular portrait of each tumour. Twenty of the tumours were sampled twice, before and after a 16-week course of doxorubicin chemotherapy, and two tumours were paired with a lymph node metastasis from the same patient. Gene expression patterns in two tumour samples from the same individual were almost always more similar to each other than either was to any other sample. Sets of co-expressed genes were identified for which variation in messenger RNA levels could be related to specific features of physiological variation. The tumours could be classified into subtypes distinguished by pervasive differences in their gene expression patterns.

Breast Neoplasms↗
PubMed2000

Pharmacokinetics and microdistribution of polyethylene glycol-modified humanized A33 antibody targeting colon cancer xenografts.

Therapeutic proteins have been conjugated with polyethylene glycol (PEGylation) to reduce immunogenicity and enhance circulating dose. Here we have investigated the effect of PEGylation on immunogenicity, pharmacokinetics, and histologic microdistribution of tumor-targeting antibodies with humanized A33 antibody (huA33) as a model system. Conjugation of huA33 with methoxy-PEG of M(r) 5,000 (32%-34% of primary amines modified) or M(r) 20,000 (16%-18% modification) preserved >50% of native huA33 binding to SW1222 colon cancer cells. In mice, both PEGylated forms cleared from serum moderately slower than native huA33. After repeated immunization with PEG-huA33, antiantibody titers in immunocompetent mice were <5% of those in huA33-treated controls. Both PEG-huA33 forms reached approx. 75% of the maximum tumor dose of huA33 in SW1222-xenografted mice, but their tumor:blood ratios were considerably reduced. To demonstrate immunologic specificity of PEG-huA33 targeting in SW1222 tumor-bearing mice, antigenic sites were presaturated by injecting excess native huA33. This reduced subsequent uptake of PEG-huA33 by up to 80%, whereas presaturation with hu3S193 control antibody had no significant effect. To assess the microdistribution of antibody uptake in the same xenograft model, tumor tissue resected at different time points after antibody administration was examined for human IgG by immunohistochemistry. Both PEG preparations achieved the same peak staining intensity and homogeneity as native huA33 with a delay of several hours. Given the measured reduction in immunoreactivity in vitro, these results demonstrate that the tumor targeting potential of huA33 in vivo is preserved at PEGylation levels sufficient to suppress immunogenicity.

Adenocarcinoma↗
PubMed2000

An advanced glycation endproduct cross-link breaker can reverse age-related increases in myocardial stiffness.

Decreased elasticity of the cardiovascular system is one of the hallmarks of the normal aging process of mammals. A potential explanation for this decreased elasticity is that glucose can react nonenzymatically with long-lived proteins, such as collagen and lens crystallin, and link them together, producing advanced glycation endproducts (AGEs). Previous studies have shown that aminoguanidine, an AGE inhibitor, can prevent glucose cross-linking of proteins and the loss of elasticity associated with aging and diabetes. Recently, an AGE cross-link breaker (ALT-711) has been described, which we have evaluated in aged dogs. After 1 month of administration of ALT-711, a significant reduction ( approximately 40%) in age-related left ventricular stiffness was observed [(57.1 +/- 6.8 mmHg x m(2)/ml pretreatment and 33.1 +/- 4.6 mmHg x m(2)/ml posttreatment (1 mmHg = 133 Pa)]. This decrease was accompanied by improvement in cardiac function.

Aging↗
PubMed2000

Effects of oligomerization on the epitopes of the human immunodeficiency virus type 1 envelope glycoproteins.

To understand the differential expression of epitopes on monomeric and oligomeric forms of the envelope glycoproteins, nine human monoclonal antibodies (mAbs) were derived from the cells of human immunodeficiency virus-infected subjects by selection with soluble oligomeric gp140 (o.140). These nine mAbs and 12 human mAbs selected with V3 peptides, viral lysates, and rgp120, specific for the V2, V3, C5, CD4-binding domain (CD4bd), and gp41, were tested in a binding assay to compare the exposure of these regions on monomeric gp120 or gp41 and on o.140. None of the 21 mAbs were oligomer specific. However, mAbs to V3 and CD4bd were "oligomer sensitive," whereas mAbs to V2 and the distal epitope of C5 tended to be "monomer sensitive" (i.e., to react better with the oligomer or monomer, respectively). The majority of anti-gp41 mAbs reacted similarly with monomer and oligomer. Although the uncleaved o.140 used in this study differs from the cleaved gp120/41 oligomer found on the native virus particle, these results suggest that new epitopes are not introduced by oligomerization of viral envelope proteins, that such oligomer-specific epitopes, if they exist, are not highly immunogenic, and/or that they are not efficiently selected using soluble o.140.

Antibodies, Monoclonal↗
PubMed2000

Physiological and metabolic responses of female games and endurance athletes to prolonged, intermittent, high-intensity running at 30 degrees and 16 degrees C ambient temperatures.

Eight female games players (GP) and eight female endurance athletes (EA) ran intermittently at high-intensity and for prolonged periods in hot (30 degrees C) and moderate (16 degrees C) ambient temperatures. The subjects performed a two-part (A, B) test based on repeated 20-m shuttle runs. Part A comprised 60 m of walking, a maximal 15-m sprint, 60 m of cruising (90% maximal oxygen uptake, VO(2max)) and 60 m of jogging (45% VO(2max)) repeated for 75 min with a 3-min rest every 15 min. Part B involved an exercise and rest pattern of 60-s running at 100% VO(2max) and 60-s rest which was continued until fatigue. Although the GP and EA did not respond differently in terms of distances completed, performance was 25 (SEM 4)% less (main effect trial, P < 0.01) in the hot (HT) compared with the moderate trial (MT). Sprints of 15 m took longer to complete in the heat (main effect, trial, P < 0.01), and sprint performance declined during HT but not MT (interaction, trial x time, P < 0.01). A very high correlation was found between the rate of rise in rectal temperature in HT and the distance completed [GP, r =-0.94, P < 0. 01; EA (n = 7), r = -0.93, P < 0.01]. Blood lactate [La(-) ](b) and plasma ammonia [NH(3)](p1) concentrations were higher for GP than EA, but were similar in HT and MT [La(-) ](b), HT: GP vs EA, 8.0 (SEM 0. 9) vs 4.9 (SEM 1.1) mmol x l(-1); MT: GP vs EA, 8.0 (SEM 1.3) vs 4.4 (SEM 1.2) mmol x l(-1); interaction, group x time, P < 0.01; [NH(3)](p1), HT: GP vs EA, 70.1 (SEM 12.7) vs 43.2 (SEM 6.1) mmol x l(-1); MT: GP vs EA, 76.8 (SEM 8.8) vs 32.5 (SEM 3.8) micromol x l(-1); interaction, group x time, P < 0.01. Ad libitum water consumption was higher in HT [HT: GP vs EA, 18.9 (SEM 2.9) vs 13.5 (SEM 1.7) ml x kg(-1) x h(-1); MT: GP vs EA, 12.7 (SEM 3.7) vs 8.5 (SEM 1.5) ml x kg(-1) x h(-1); main effect, group, n.s.; main effect, trial, P < 0.01]. These results would suggest that elevated body temperature is probably the key factor limiting performance of prolonged, intermittent, high-intensity running when the ambient temperature is high, but not because of its effect on the metabolic responses to exercise.

Adult↗
PubMed2000

Short-term recovery from prolonged constant pace running in a warm environment: the effectiveness of a carbohydrate-electrolyte solution.

Recovery from prolonged exercise involves both rehydration and replenishment of endogenous carbohydrate stores. This study examined the influence of drinking a carbohydrate-electrolyte solution on short-term recovery and subsequent exercise capacity in a warm environment. Thirteen healthy male volunteers completed two trials, at least 7 days apart. On each occasion subjects performed an initial treadmill run at 60% of maximal oxygen uptake (VO2max), for 90 min or until volitional fatigue (T1), in a warm environment (35 degrees C, 40% relative humidity, RH). Volitional ingestion of water was permitted during each of the exercise trials. During a subsequent 4-h recovery period (REC) subjects consumed either a 6.9% carbohydrate-electrolyte solution (CES) or a sweetened placebo (P), in a volume equivalent to 140% of body mass loss. Following REC, subjects ran to exhaustion at the same % VO2max in order to assess their endurance capacity (T2). Mean (SEM) run times during T1 did not differ between the CES [74.8 (4.6) min] and P [72.5 (5.2) min] trials. Body mass was reduced (P < 0.01) by 1.9 (0.2)% (CES) and 1.7 (0.2)% (P), and plasma volume (P < 0.01) by 6.0 (0.9)% (CES) and 5.4 (1.0)% (P) during the T1 trials. During REC 2006 (176) ml and 1830 (165) ml of fluid was ingested, providing 138 (12) g and 0 g of carbohydrate in the CES and P trials, respectively. Prior to T2, plasma volume and net fluid balance were similarly restored [CES +58 (26) g; P -4 (68) g] in both trials. During T2 the exercise duration was longer (P < 0.01) in the CES compared to the P trial [CES 60.9 (5.5) min; P 44.9 (3.0) min]. Thus, provided that an adequate hydration status is maintained, inclusion of carbohydrate within an oral rehydration solution will delay the onset of fatigue during a subsequent bout of prolonged submaximal running in a warm environment.

Adult↗
PubMed2000

Increased prevalence of precancerous changes in relatives of gastric cancer patients: critical role of H. pylori.

BACKGROUND & AIMS: Helicobacter pylori is believed to predispose to gastric cancer by inducing gastric atrophy and hypochlorhydria. First-degree relatives of patients with gastric cancer have an increased risk of developing gastric cancer. The aim of this study was to determine the prevalence of atrophy and hypochlorhydria and their association with H. pylori infection in first-degree relatives of patients with gastric cancer. METHODS: H. pylori status, gastric secretory function, and gastric histology were studied in 100 first-degree relatives of patients with noncardia gastric cancer and compared with those of controls with no family history of this cancer. RESULTS: Compared with healthy controls, relatives of patients with gastric cancer had a higher prevalence of hypochlorhydria (27% vs. 3%) but a similar prevalence of H. pylori infection (63% vs. 64%). Relatives of cancer patients also had a higher prevalence of atrophy (34%) than patients with nonulcer dyspepsia (5%) matched for H. pylori prevalence. Among the relatives of cancer patients, the prevalence of atrophy and hypochlorhydria was increased only in those with evidence of H. pylori infection, was greater in relatives of patients with familial cancer than in relatives of sporadic cancer index patients, and increased with age. Eradication of H. pylori infection produced resolution of the gastric inflammation in each subject and resolution of hypochlorhydria and atrophy in 50% of the subjects. CONCLUSIONS: Relatives of patients with gastric cancer have an increased prevalence of precancerous gastric abnormalities, but this increase is confined to those with H. pylori infection. Consequently, prophylactic eradication of the infection should be offered to such subjects.

Achlorhydria↗
PubMed2000

The integration of SPR biosensors with mass spectrometry: possible applications for proteome analysis.

The successful integration of biospecific interaction analysis based on surface plasmon resonance and mass spectrometry produces a powerful technique that couples the benefits of sensitive affinity capture and characterization of binding events with the ability to characterize interacting molecules. A variety of biosensors has been used to capture proteins and peptides biospecifically on sensor surfaces, with subsequent analysis using mass spectrometry. Applying this type of analysis to proteomic studies could lead to ligand and protein-complex identification, and might provide clues leading to the identification of pathways.

Amino Acid Sequence↗
PubMed2000

Doing health, doing gender: teenagers, diabetes and asthma.

Although most research linking health disadvantage with gender has focused on women, recent work indicates that hegemonic masculinities can also place the health of men at risk. The importance of comparing the experiences of women and men has been emphasised and this paper focuses on the ways in which the social constructions of femininities and masculinities affect how teenagers live with asthma or diabetes. The majority of girls incorporated these conditions and the associated treatment regimens into their social and personal identities, showing a greater adaptability to living with asthma or diabetes. However, this could have detrimental effects in terms of control, as girls sometimes lowered expectations for themselves. In addition, two aspects of the treatment regimens, diet and exercise, were found to disadvantage girls and advantage boys, because of contemporary meanings of femininities and masculinities. The social construction of femininities meant that these conditions were not seen as the threat that they were by the majority of boys interviewed, who made every effort to keep both conditions outside their personal and social identities by passing. The majority of boys maintained a 'valued' identity by feeling in control of their body and their condition. However, for the small minority of boys who were no longer able to pass the impact of chronic illness led to a 'disparaged' identity. The interaction of gender and health is seen as a complex two-way process, with aspects of contemporary femininities and masculinities impacting on the management of these conditions, and aspects of these conditions impacting in gendered ways upon the constructions of gender.

Adaptation, Psychological↗
PubMed2000

Preliminary evaluation of an in vitro test for assessment of excitotoxicity by measurement of early gene (c-fos mRNA) levels.

Using primary cultures of mouse cerebellar granule cells as an in vitro model system, it has been demonstrated that different profiles of temporal expression of the c-fos proto-oncogene are observed under non-excitotoxic and excitotoxic conditions. A ratio has been derived previously for the steady-state level of c-fos mRNA after 30 min and 240 min which suggests that a 240 min/30 min ratio of greater than 1 correlates with excitotoxicity, whereas a ratio of less than 1 correlates with a non-excitotoxic outcome. Moreover, a positive correlation is seen with abrogation of excitotoxicity in response to selective excitatory amino acid receptor antagonists. This test, proposed as a specific biomarker for excitotoxicity is undergoing prevalidation. Excitotoxicity is defined as neuronal cell death mediated by hyperactivation of glutamate receptor subtypes and therefore might be expected to be prevented by selective glutamate receptor antagonists. In preliminary evaluation studies, we have conducted work under the direction of the European Center for Validation of Alternate Methods (ECVAM) using compounds specified by ECVAM that have been subdivided into four groups based on known or presumed actions. These groups comprise: Group 1-endogenous/synthetic excitotoxins; Group 2-environmental, putative excitotoxins; Group 3-neurotoxic but non-excitotoxic compounds, and Group 4-non-toxic compounds. The results of this study support the proposal that the c-fos mRNA time-ratio test is a specific biomarker of excitotoxicity. Just as importantly, this test has the potential for application in screening newly-designed EAA receptor antagonists in the search for clinically relevant drugs to treat a variety of neuropathologies.

Animals↗
PubMed2000

Biotechnology match making: screening orphan ligands and receptors.

To date there has been a considerable amount of interest and success in the pharmaceutical industry in the discovery of drug targets and diagnostics via genomic technologies, namely DNA sequencing, mutation/polymorphism detection and expression monitoring of mRNA. As the ultimate targets for the majority of these methods are actually proteins, more and more emphasis has been placed upon protein-based methods in an effort to define the function of proteins discovered by genomic technologies. One of the most challenging areas of drug target discovery facing researchers today is the search for novel receptor-ligand pairs. Database mining techniques in conjunction with other computational methods are able to identify many novel sequences of putative receptors, but the ability to similarly identify the receptor's natural ligand is not possible by these methods. The past few years have seen an increase in methodology and instrumentation focused on the ability to discover and characterize protein-protein interactions, as well as receptor-ligand pairs. Significant advances have been made in the areas of instrumentation (biosensors and fluorescent plate readers) as well as methodologies relating to phage/ribosome display and library construction.

Animals↗
PubMed2000

The use of zeolites as slow release anthelmintic carriers.

This work examines the ability of commercial zeolite Y to act as a slow release agent for a number of anthelmintic drugs. Administration to rats, dosed with Nippostrongylus brasiliensis, of pyrantel and/or fenbendazole and pigs, dosed with Ascaris and Oesophagostomum, of dichlorvos (DDVP) loaded onto zeolite Y was more successful in killing adult worms than administration of the pure drug alone. The zeolite Y was used as supplied for initial studies and then later dealuminated for further studies. The drug loadings were monitored by thermal analysis and the loaded zeolites were used in several field trials. The results indicate that zeolite Y is a suitable vehicle for the slow release of some anthelmintics. The slow release of drug from the zeolite matrix improved its efficacy.

Animals↗