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Biomedical subjects

D R Smith

Publications and source records attributed to D R Smith.

At least 127 records · Page 7Linked to original sources
PubMed1996

Microbial pathogen genomes--new strategies for identifying therapeutics and vaccine targets.

Advances in high-throughput DNA-sequencing techniques have given us the unprecedented ability to rapidly determine the nucleotide sequences of entire bacterial genomes. The application of these methods to the genomes of microbial pathogens, combined with efficient analytical tools and genome-scale approaches for studying gene expression, is revolutionizing our approach to the selection of targets for drug screening and vaccine development. This is bringing new life to this important, but long-neglected, field of research.

Animals↗
PubMed1996

C-KI-RAS activation and the biological behaviour of proximal and distal colonic adenocarcinomas.

One hundred and forty colonic adenocarcinomas originating on the left side of the colorectum and 70 colorectal carcinomas originating on right side of the colorectum were examined for activating mutations of codons 12 and 13 of the C-KI-RAS proto-oncogene. Rates of mutation were significantly different (right colon 43%, 30/70 versus left colon 23%, 32/140; P = 0.0025). Adenocarcinomas from the left side of the colorectum showed a significant association between C-KI-RAS activation and tumour progression, including the presence of distant organ metastasis at the time of surgery (P = 0.0039), and during patient follow-up (P = 0.00027), whereas those from the right of the colorectum did not (P = 0.4 and P = 0.5, respectively). Mutation of the C-KI-RAS proto-oncogene was found to be associated with a significantly poorer patient prognosis on the left of the colorectum (P = 0.0001 by log rank analysis of Kaplan-Meier plots) but not on the right (P = 0.7). These results demonstrate that, not only is the timing and frequency of C-KI-RAS activation different between carcinomas originating on the left or right of the colorectum, but also that the biological consequences of such mutations may differ.

Adenocarcinoma↗
PubMed1996

Development and validation of analytical methodology for near-infrared conformance testing of pharmaceutical intermediates.

An approach was developed to create and validate analytical methods to perform near-infrared (NIR) conformance testing on two isolated intermediates used in the manufacture of loracarbef monohydrate bulk drug substance. Method calibration sets were developed from second-derivative NIR reflectance spectra for 30 representative batches of each intermediate. In conformance testing, second-derivative NIR spectra for samples from newly manufactured batches are compared with the calibration set. If the new spectrum is not statistically different to the average of the calibration set, the sample passes the conformance test. Using authentic batch samples of typical and low-potency lots, the methods were validated for accuracy, selectivity, ruggedness and repeatability of the methods.

Calibration↗
PubMed1996

Prognostic significance of p53 overexpression and mutation in colorectal adenocarcinomas.

The p53 tumour-suppressor gene is found altered in the majority of colorectal cancers. Lesions include allelic loss, mutation of the gene and overexpression of the p53 protein. All of these lesions have been analysed for prognostic significance, and whereas both mutation and allelic loss have been shown to be reasonably useful markers of prognosis, the utility of overexpression of the p53 protein is more ambiguous. Given that many authors use p53 overexpression as a marker for point mutation this issue is of some importance. We have therefore examined 100 colorectal carcinomas for mutation of the p53 gene, as well as overexpression of the p53 protein. Results show that whereas mutation of the p53 gene is associated with p53 overexpression, the degree of association depends, at least in part, upon the particular antibody used. Moreover, although mutation of the p53 gene does provide prognostic information, overexpression of the p53 protein, as detected with two antibodies, does not. These results suggest that immunohistochemistry is not a suitable alternative to direct detection of mutation in assessing prognosis in colorectal cancer patients.

Adenocarcinoma↗
PubMed1996

Use of endogenous, stable lead isotopes to determine release of lead from the skeleton.

The stable lead isotope methodology can be used to study the release of lead from bone into the circulation because of its potential to distinguish circulatory lead from "older" and isotopically different skeletal lead that may have been accumulated years or decades earlier. Here we report the initial results from a larger ongoing study that evaluates the skeleton as a source of lead to the circulation in environmentally exposed human subjects. Lead concentrations and stable lead isotopic compositions were measured in blood and trabecular bone samples obtained from five patients who underwent total hip or knee joint replacement. All subjects contained low blood (1-6 micrograms/dl) and bone (0.6-7 micrograms/g dry weight) lead concentrations typical of environmentally exposed individuals. There were relatively large differences in the lead isotopic compositions between the paired blood and bone samples from each subject. These isotopic differences are attributed to differences in the lead isotopic compositions of past versus current lead exposures and to the long elimination half-life of lead in the skeleton compared to lead in the circulation. Based on these data, we determined that the skeleton contributed 40-70% of the lead in the blood of these subjects. This initial study demonstrates the utility of the stable lead isotope methodology for investigating the release of lead from the skeleton. It also shows that the skeleton can be an important endogenous source of lead exposure in environmentally exposed humans.

Aged↗
PubMed1996

Depression of lipogenesis in swine adipose tissue by specific dietary fatty acids.

The objective of this study was to document the influence of specific dietary fatty acids on rates of lipid synthesis and sensitivity to insulin in porcine adipose tissue. Weanling pigs were assigned to one of six groups, and each group was fed diets containing 10 g/100 g of added cornstarch or 10 g/100 g of added fatty acid. The fatty acid-enriched diets contained either a combination of 14:1 plus 16:1 (14:1/16:1 diet), 16:0, 18:0, 18:1, or 18:2 (n-6). With the exception of the cornstarch diet, all diets contained approximately 35% 14:0. Subcutaneous adipose tissue samples were collected at slaughter from the area overlying the first cranial vertebra. Fresh samples were incubated for 2 h in 20 mM glucose and 0, 10, 100 or 1,000 microU/mL of porcine insulin. The smallest adipocytes were observed in adipose tissue from pigs fed the 16:0 or 18:2 diets. Glucose incorporation into lipids was greater (P < .05) in adipose tissue from cornstarch-fed pigs than in adipose tissue from the other treatment groups. Lipogenesis was 67, 53, 35, 32, and 20% lower (P < .05) in adipose tissue from 16:0-, 14:1/16:1-, 18:0-, 18:2-, and 18:1-fed pigs, respectively, than in adipose tissue from the cornstarch-fed pigs. Insulin increased lipogenesis by 19% (P < .05) in adipose tissue from the cornstarch-fed pigs and by 15 to 40% (P < .05) in adipose tissue from the 14:1/16:1-fed pigs. Insulin did not stimulate lipogenesis (P > .4) in adipose tissue from pigs fed the 16:0, 18:0, or 18:1 diets. The data suggest that fatty acid chain length and unsaturation are determinants in the effects of dietary fat and insulin on de novo lipogenesis.

Adipocytes↗
PubMed1996

Stimulation of mitochondrial oxygen consumption in isolated cardiomyocytes after hypoxia-reoxygenation.

An increase in mitochondrial matrix free calcium has been shown to occur during oxygen and substrate deprivation of the perfused heart which predisposes to calcium overload and inhibition of mitochondrial function on reoxygenation. In the current study we have assessed the effect of substrate free hypoxia on mitochondrial oxygen consumption and ATP synthesis in isolated rat cardiomyocytes. Myocytes were subjected to 40 min of substrate-free hypoxia and the oxygen consumption measured together with the effects on ATP and PCr synthesis. After hypoxia myocytes showed a fall in ATP to 10% of the control value. Within 5 sec of reoxygenation the ATP level recovered to a new steady state level of 30% of the original value. The rate of oxygen consumption of hypoxic/reoxygenated cells was 3-4 fold higher than that of cells maintained under normoxic controls but in the presence of oligomycin the difference was only 1.5-fold higher, indicating a greater requirement for mitochondrial synthesis of ATP following reoxygenation. Reoxygenation in the absence of extracellular Ca2+ resulted in a lower rate of oxygen consumption (50% of the rate measured in the presence of 1 mM-Ca2+) but did not affect the steady state concentration of ATP attained 5 min after reoxygenation. These results support the idea that the increased O2 consumption of myocytes following hypoxia/reoxygenation is due to an increased demand for ATP synthesis by mitochondria and is a response to the NA+ and Ca2+ loading of the cells which occurs under these conditions. This increased demand is likely to result in a greater generation of free radicals such as superoxide by the respiratory chain which could impair cellular function over the long term.

Adenine Nucleotides↗
PubMed1996

Infection and cross-protection studies of winter dysentery and calf diarrhea bovine coronavirus strains in colostrum-deprived and gnotobiotic calves.

OBJECTIVE: To investigate in vitro antigenic relations, in vivo cross-protection, and isotype antibody responses to a winter dysentery (WD) and calf diarrhea strain of bovine coronavirus (BCV). DESIGN AND ANIMALS: Gnotobiotic and colostrum-deprived calves were inoculated oronasally with a WD (DBA) or a calf diarrhea (DB2) BCV, and were challenge exposed with the heterologous BCV. PROCEDURE: Nasal swab and feces specimens and blood samples were collected. Fecal and nasal specimens were assayed for BCV shedding by antigen-capture ELISA or immune electron microscopy. Bovine coronavirus antigens were detected in nasal epithelial cells by immunofluorescence. Antibody titers to BCV in serum were assayed by virus neutralization (VN), and BCV antibody isotype titers in feces and sera were quantitated by ELISA. RESULTS: All calves developed diarrhea and shed BCV nasally and in feces, then recovered and were protected from BCV-associated diarrhea after challenge exposure with the heterologous BCV. After challenge exposure with either strain, fecal shedding of DBA was detected in 1 of 4 calves and nasal shedding of DB2 was detected in 2 of 4 calves. Immunoglobulin M was the principal coproantibody to BCV early, followed predominantly by IgA. Immunoglobulin G1 coproantibody titers to BCV were low, but increased after challenge exposure. Immunoglobulin G1 antibodies were predominant in serum. After challenge exposure, all serum antibody isotype titers increased except IgG2. The VN antibody responses paralleled serum IgG1 antibody responses. CONCLUSIONS AND CLINICAL RELEVANCE: Immunoglobulin A coproantibodies at challenge exposure were associated with protection against diarrhea. Nasal shedding of BCV after challenge exposure confirmed field data documenting reinfection of the respiratory tract of cattle, suggesting that, in closed herds, respiratory tract infections constitute a source of BCV transmission to cows (WD) or young calves.

Animals↗
PubMed1996

p53 and prognosis in colorectal cancer.

The p53 tumour suppressor gene is found altered in the majority of colorectal adenocarcinomas. While these changes are believed to reflect underlying mechanisms of tumour development and progression, it is becoming increasingly clear that such changes may also reflect the macroscopic biological behaviour of the course of the disease. Knowledge of the p53 tumour suppressor gene status may therefore provide important prognostic information. This review examines some of the data that are beginning to show that p53 tumour suppressor gene status may be an important prognostic indicator.

Adenocarcinoma↗
PubMed1996

Molecular changes of colorectal cancer in Singapore.

Improved success in the management of colorectal cancer requires a better understanding of its development and biological behaviour. The key for this is molecular genetics. Gene changes parallel the multi-step changes in the adenoma-carcinoma sequence. Cancer results from a variable combination of defects in oncogenes, tumour suppressor, mutator and apoptotic genes. These changes are similar whether they occur in inherited disorders like adenomatous polyposis coli (APC) and hereditary non-polyposis colorectal cancer (HNPCC) or acquired cancer in the elderly. In Singapore, the c-myc and c-Ki-ras proto-oncogenes are found to be activated in 70% and 29% of tumours respectively. Allelic loss of chromosome 5q and 17p occurs in 25% and 70% of tumours respectively, while point mutation of the p53 tumour suppressor gene occurs in 50% of colorectal cancers. Both the frequency and the nature of the lesion occurring are compatible to the changes detected in Caucasian patients, suggesting common aetiological factors. The biological behaviour of colorectal adenocarcinomas is determined by the nature of defects or mutations in key genes such as the p53 tumour suppressor gene. Lymphatic spread is associated with the presence of point mutations and haematogenous spread is associated with loss of heterozygosity of p53. Survival is worse when conserved regions of the gene are mutated compared with those outside, and worst when codon 175 is mutated. Sensitivity to radiotherapy and chemotherapy is also determined by p53 mutation which controls apoptosis. Prognosis could now be individualised and with the prospect of gene therapy, molecular genetics will have a major impact on the management of colorectal cancer.

Adenocarcinoma↗
PubMed1996

Use of molecular methods in the early diagnosis of familial adenomatous polyposis and hereditary nonpolyposis colorectal cancer.

Familial adenomatous polyposis (FAP) and hereditary nonpolyposis colorectal cancer (HNPCC) are the most well-defined heritable conditions which predispose to colorectal cancer at a young age. Significant progress in understanding the pathogenesis of FAP and HNPCC has led to the development of techniques which can be used for the diagnosis of these two conditions. In this article, the technical aspects and clinical applications of molecular methods such as linkage analysis, mutational analysis and in vitro synthesised-protein assay for FAP and microsatellite instability or replication error (RER) assay for HNPCC are described and discussed. The potential of such molecular tests is the identification of affected individuals for proper surveillance and management as well as the identification of non-affected individuals to free them from the trauma of uncertainty and repeated unnecessary colonoscopies.

Adenomatous Polyposis Coli↗