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D Thomas

Publications and source records attributed to D Thomas.

At least 217 records · Page 12Linked to original sources
PubMed1999

Comparison of the intracellular signaling responses by three chimeric fibroblast growth factor receptors in PC12 cells.

Stably transfected PC12 cell lines expressing similar amounts of chimeric receptors composed of the extracellular domain of the human platelet-derived growth factor (PDGF)beta receptor and the transmembrane and intracellular domains of the fibroblast growth factor receptors (FGFRs) 1, 3, and 4 undergo ligand-induced differentiation. The FGFR1 chimera (PFR1) is the most potent of the three, and PFR4 requires more frequent (every 24 hr) addition of ligand to maintain the response. Both PFR1 and -3 also show significant ligand-independent autophosphorylation but PFR4 does not. All of the chimeras activated phospholipase Cgamma, Shc, FGFR substrate (FRS)2, and the mitogen-activated protein kinases, ERK1 and 2. PFR4 was moderately weaker in stimulating these effects as well; PFR1 and -3 were comparable. None of the chimeras induced Sos association or were coprecipitated with Shc. Cotransfection of a dominant-negative Shc derivative, with tyrosine at 239, 240, and 317 replaced with phenylalanine, in the PFR-expressing cells was without effect on PDGF-induced neurite outgrowth. The same derivative substantially inhibited the response of these cells to NGF. These results indicate that FGFR1, 3, and 4 (i) are capable of signaling in a similar fashion; (ii) primarily use FRS2 and, perhaps, PLCgamma; and (iii) do not utilize Shc. The results also suggest that the principal difference between FGFR1, 3, and 4 is in the strength of the tyrosine kinase activity and that qualitative differences in signaling capacity are likely to be less important.

Animals↗
PubMed1999

Relation between direct detection of Chlamydia pneumoniae DNA in human coronary arteries at postmortem examination and histological severity (Stary grading) of associated atherosclerotic plaque.

BACKGROUND: Numerous studies have suggested a link between Chlamydia pneumoniae infection, atherosclerosis, and coronary artery disease. However, it is still unclear whether C pneumoniae plays a causal role in the pathogenesis of these conditions. Accordingly, we have performed a systematic dissection of the 3 coronary arteries on 33 postmortem subjects and studied the relationship in individual artery segments between the presence of C pneumoniae DNA and the severity of associated atherosclerosis. METHODS AND RESULTS: The prevalence of C pneumoniae DNA in arterial segments was determined by polymerase chain reaction (PCR) after controlling for the presence of PCR inhibitors. Atherosclerosis in each arterial segment was graded histologically with the Stary classification. C pneumoniae was detected by PCR in 78.8% of subjects, but there was no association between the presence of this DNA and cause of death or grade of atherosclerosis. When paired mild and severe atherosclerotic lesions within subjects were compared, mild lesions were as likely to be positive for C pneumoniae as severe lesions. CONCLUSIONS: This study demonstrates that C pneumoniae can frequently be detected in atheromatous plaques in coronary arteries. However, its distribution did not correlate with severity or extent of disease.

Aged↗
PubMed1999

In vitro assembly properties of wild-type and cyclophilin-binding defective human immunodeficiency virus capsid proteins in the presence and absence of cyclophilin A.

The cellular protein cyclophilin A (CypA) binds specifically to the human immunodeficiency virus type 1 (HIV-1) capsid (CA) protein and is incorporated into HIV-1 particles at a molar ratio of 1:10 (CypA/CA). Structural analysis of a CA-CypA complex suggested that CypA may destabilize interactions in the viral capsid and thus promote uncoating. We analyzed the influence of CypA on the in vitro assembly properties of wild-type (WT) CA and derivatives containing substitutions of Gly89 in the Cyp-binding loop. All variant proteins were significantly impaired in CypA binding. In the presence of CypA at a molar ratio of 1:10 (CypA/CA), WT CA assembled into hollow cylinders that were similar to those observed in the absence of CypA but slightly longer. Higher CypA concentrations inhibited cylinder formation. Variant CA proteins G89L and G89F yielded similar cylinders as the WT protein but were significantly more resistant to CypA. Cryoelectron microscopic analysis of WT cylinders assembled in the presence of CypA revealed direct binding of CypA to the outer surface. Electron diffraction patterns generated from these cylinders indicated that CypA causes local disorder. The addition of CypA to preassembled cylinders had little effect, however, and cylinders were only disrupted when incubated with a threefold molar excess of CypA for several hours. These results suggest that CypA does not efficiently destabilize CA interactions at the molar ratio observed in the virion and therefore is unlikely to serve as an uncoating factor.

Capsid↗
PubMed1999

Mercuric chloride-induced apoptosis is dependent on protein synthesis.

Apoptosis is a mode of cell death with morphologic and biochemical features that distinguish it from necrosis. Recent studies demonstrating that mercury compounds initiate apoptosis in cultured cells did not elucidate if the biochemical mechanism of apoptosis involved a dependence on macromolecular synthesis post-insult, i.e. programmed cell death. The objectives of this in vitro study were (1) to determine if HgCl2 cytotoxicity includes an apoptotic component, and (2) to determine if apoptosis is dependent on protein synthesis, i.e. proceeds by an inducible mechanism. Suspensions of mouse lymphoma (L5178Y-R) cells were exposed to 0, 1, 5, or 10 microM HgCl2 and apoptosis was evaluated utilizing qualitative and quantitative methods. At 24 h after exposure, transmission electron microscopy revealed a concentration-related increase in morphologic changes typical of apoptosis: margination of condensed chromatin to the nuclear membrane, dilation of the rough endoplasmic reticulum, cytoplasmic condensation and vacuolation, nuclear dissolution, and plasma membrane blebbing. An increase in Hg-induced DNA fragmentation (DNA 'ladder') was observed using agarose gel electrophoresis. Time- and concentration-dependent increases in the percent of apoptotic cells were observed at 1, 6, 12, and 24 h after HgCl2 exposure using a flow cytometric method that discriminates between cells according to size and granularity. Pretreatment of cells with cycloheximide (CHX), an inhibitor of translation, prior to HgCl2 exposure resulted in a 25-50% reduction in apoptotic cells 24 h after exposure to 10 and 20 microM HgCl2, and concomitantly reduced the overall cytotoxicity compared to HgCl2 alone. These results, although limited to a single cell line, support the hypothesis that HgCl2 induces apoptosis that is dependent, at least in part, upon protein synthesis.

Animals↗
PubMed1999

The repressor which binds the -75 GATA motif of the GPB promoter contains Ku70 as the DNA binding subunit.

Glycophorin B (GPB) is an abundant cell surface glycoprotein which is only expressed in human erythroid cells. Previous functional analysis demonstrated that the repression of the GPB promoter is determined by the binding of a ubiquitous factor which recognizes a GATA motif centered at position -75. In erythroid cells this ubiquitous factor is displaced by the binding of the erythroid-specific factor hGATA1. Here, we have identified the Ku70 protein as a candidate GPB repressor DNA binding subunit through the screening of a human HeLa expression library using the -75 GATA sequence as bait (one-hybrid method). Electrophoretic mobility shift assays demonstrated that the ubiquitous factor that binds the -75 GATA sequence was the Ku70-Ku80 (Ku) heterodimer. Co-transfection experiments demonstrated that overexpression of Ku70 in the K562 erythroleukeamic cell line resulted in transcriptional repression of the chloramphenicol acetyltransferase reporter gene when placed under the control of the wild-type GPB promoter. Conversely, no repression was observed when a mutation that abolished the binding of Ku was introduced in the GPB promoter construct. Altogether, these results indicate that Ku binds in vivo to the -75 WGATAR motif and is involved in negative regulation of the GPB promoter. These findings suggest that, besides its role in many functions, Ku is also involved in transcriptional regulation of erythroid genes.

Antigens, Nuclear↗
PubMed1999

[Impact of an information campaign on cardiovascular risk factors. 5-year results at the study town Epernon].

OBJECTIVES: Information campaigns are implemented to improve knowledge of cardiovascular disease and risk factors. The impact of these programs must be evaluated to determine whether they truly contribute to modifying risk factors in the population. METHODS: A 5-year information campaign on cardiovascular disease and risk factors was conducted in Epernon, France. The main objectives of the campaign focused on stopping smoking, regular physical exercise and balanced nutrition. Data were collected from a representative sample of the female and male population aged 20 to 65 years selected from the study town Epernon (500 subjects), and in control towns, Magny-en-Vexin and Moret-sur-Loing (200 subjects). The study town and control towns were comparable for population, demographic characteristics and geographic localization (distance from Paris). The subjects were invited to respond to a questionnaire on demographic data, attitudes toward health, risk factors and diet and underwent a clinical examination with blood tests. RESULTS: The initial sample included 961 subjects and 68.5% participated in the final survey. We were unable to evidence any significant difference in risk factors or the 10-year risk score calculated from the Framingham equation adapted to France. The information campaign was well accepted, the population not expression a feeling of lassitude. The campaign had a minimal effect on the way individuals relate to health. There was a fall in mean energy intake, mainly fat calories, which was similar in all three towns. CONCLUSION: No major modification in cardiovascular risk factors was observed in this low-risk population, suggesting that future information campaigns should be aimed at targeted populations with a higher risk profile using simple and selected messages.

Adult↗
PubMed1999

Switch from an aquaporin to a glycerol channel by two amino acids substitution.

The MIP (major intrinsic protein) proteins constitute a channel family of currently 150 members that have been identified in cell membranes of organisms ranging from bacteria to man. Among these proteins, two functionally distinct subgroups are characterized: aquaporins that allow specific water transfer and glycerol channels that are involved in glycerol and small neutral solutes transport. Since the flow of small molecules across cell membranes is vital for every living organism, the study of such proteins is of particular interest. For instance, aquaporins located in kidney cell membranes are responsible for reabsorption of 150 liters of water/day in adult human. To understand the molecular mechanisms of solute transport specificity, we analyzed mutant aquaporins in which highly conserved residues have been substituted by amino acids located at the same positions in glycerol channels. Here, we show that substitution of a tyrosine and a tryptophan by a proline and a leucine, respectively, in the sixth transmembrane helix of an aquaporin leads to a switch in the selectivity of the channel, from water to glycerol.

Adult↗
PubMed1999

Latency analysis in epidemiologic studies of occupational exposures: application to the Colorado Plateau uranium miners cohort.

BACKGROUND: Latency effects are an important factor in assessing the public health implications of an occupational or environmental exposure. Usually, however, latency results as described in the literature are insufficient to answer public health related questions. Alternative approaches to the analysis of latency effects are warranted. METHODS: A general statistical framework for modeling latency effects is described. We then propose bilinear and exponential decay latency models for analyzing latency effects as they have parameters that address questions of public health interest. Methods are described for fitting these models to cohort or case-control data; statistical inference is based on standard likelihood methods. APPLICATION: A latency analysis of radon exposure and lung cancer in the Colorado Plateau uranium miners cohort was performed. We first analyzed the entire cohort and found that the relative risk associated with exposure increases for about 8.5 years and thereafter decreases until it reaches background levels after about 34 years. The hypothesis that the relative risk remains at its peak level is strongly rejected (P < 0.001). Next, we investigated the variation in the latency effects over subsets of the cohort based on attained age, level and rate of exposure, and smoking. Age was the only factor for which effect modification was demonstrated (P = 0.014). We found that the decline in effect is much steeper at older ages (60+ years) than younger. CONCLUSION: The proposed methods can provide much more information about the exposure-disease latency effects than those generally used.

Air Pollutants, Radioactive↗
PubMed1999

The role of familial values in understanding the impact of social class on weight concern.

OBJECTIVE: To examine the role of social class on aspects of weight concern and to assess the possible impact of values on mediating this association. METHOD: Two hundred fifty-seven girls ranging in age from 13 to 16, from either a fee paying inner city independent girls school (higher class school, n = 135) or a state comprehensive, inner city girls school (lower class school, n = 122) completed a questionnaire concerning their profile characteristics (age, social class), aspects of their weight concern, and their own and their perceptions of their significant others' values (achievement, family life, and physical appearance). RESULTS: The results showed consistent effects of class on weight concern, with the higher class subjects reporting higher levels of restrained eating, greater body dissatisfaction, and body distortion than their lower class counterparts. The results also showed an effect of class on values, with the lower class subjects placing more importance on family life from both their own perspective and that of their parents and friends, and rating their friends as valuing achievement and physical appearance more than the higher class subjects. In terms of the best predictors of weight concern, the results showed that higher levels of restrained eating were related to being from a higher class, placing greater importance upon physical appearance, preferring a thinner ideal female body, and placing less importance upon family life; greater body dissatisfaction was related to being from a higher social class, placing greater importance upon physical appearance and a lower importance upon achievement, and greater body distortion was related to being from a higher social class and a high value placed upon physical appearance. CONCLUSION: The results indicate both a direct social class/weight concern link and a relationship which is mediated by values. The results are discussed in terms of developing an improved measure of class values and the relatively stable nature of class boundaries.

Adolescent↗
PubMed1999

Inhibitory effects of the class III antiarrhythmic drug amiodarone on cloned HERG potassium channels.

The human ether-a-go-go-related gene (HERG) encodes a K+ channel with biophysical properties nearly identical to the rapid component of the cardiac-delayed rectifier K+ current (I(Kr)). HERG channels are one primary target for the pharmacological management of arrhythmias. In this study, we investigated the acute effects of the class III antiarrhythmic drug amiodarone on HERG channels expressed heterologously in Xenopus oocytes by use of the two-microelectrode voltage clamp technique. Amiodarone blocked HERG channels with an IC50 of 9.8 microM with a maximum outward tail current reduction of 62.8%. The block consisted of two main components, a closed channel block that could not be reversed within the time of experiments and an open channel block with a slow unblock, having a recovery time constant of 73 s at -80 mV. Inactivation of the HERG channel at very positive potentials could not prevent amiodarone block. These results indicate that HERG channels can be blocked by amiodarone in closed, open and inactivated states. The block of open channels was cumulative, use-dependent and voltage-dependent. In summary, our data suggest that the strong class III antiarrhythmic action of amiodarone is at least partially based upon its acute inhibitory effects on HERG potassium channels.

Amiodarone↗
PubMed1999

Desiccation and osmotic stress increase the abundance of mRNA of the tonoplast aquaporin BobTIP26-1 in cauliflower cells.

Changes in vacuolar structure and the expression at the RNA level of a tonoplast aquaporin (BobTIP26-1) were examined in cauliflower (Brassicaoleracea L. var. botrytis) under water-stress conditions. Gradual drying out of slices of cauliflower floret tissue caused its collapse, with a shrinkage in tissue and cell volumes and an apparent vesiculation of the central vacuole, whereas osmotic stress resulted in plasmolysis with a collapse of the cytoplasm and the central vacuole within. Osmotic stress caused a rapid and substantial increase in BobTIP26 mRNA in slices of floret tissue. Exposure of tissue slices to a regime of desiccation showed a slower but equally large rise in BobTIP26 mRNA followed by a rapid decline upon rehydration. In situ hybridization showed that BobTIP26-2 mRNA is expressed most highly in meristematic and expanding cells of the cauliflower florets and that desiccation strongly increased the expression in those cells and in differentiated cells near the xylem vessels. These data indicate that under water-deficit conditions, expression of the tonoplast aquaporin gene in cauliflower is subject to a precise regulation that can be correlated with important cytological changes in the cells.

Animals↗
PubMed1999

Epidermal growth factor- and hepatocyte growth factor-receptor activity in serum-free cultures of human hepatocytes.

BACKGROUND/AIMS: Serum-free primary cultures of hepatocytes are a useful tool to study factors triggering hepatocyte proliferation and regeneration. We have developed a chemically defined serum-free system that allows human hepatocyte proliferation in the presence of epidermal growth factor and hepatocyte growth factor. METHODS: DNA synthesis and accumulation were determined by [3H]thymidine incorporation and fluorometry, respectively. Western blot analyses and co-immunoprecipitations were used to investigate the association of proteins involved in epidermal growth factor and hepatocyte growth factor activation and signaling: epidermal growth factor receptor, hepatocyte growth factor receptor (MET), urokinase-type plasminogen activator and its receptor, and a member of the signal transducer and activator of transcription family, STAT-3. RESULTS: Primary human hepatocytes proliferated under serum-free conditions in a chemically defined medium for up to 12 days. Epidermal growth factor-receptor and MET were present and functional, decreasing over time. MET, urokinase-type plasminogen activator and urokinase-type plasminogen activator receptor co-precipitated to varying degrees during the culture period. STAT-3 co-precipitated with epidermal growth factor-receptor and MET to varying degrees. CONCLUSIONS: Proliferation of human hepatocytes can improve by modification of a chemically defined medium originally used for rat hepatocyte cultures. In these long-term cultures of human hepatocytes, hepatocyte growth factor and epidermal growth factor can stimulate growth and differentiation by interacting with their receptors and initiating downstream signaling. This involves complex formation of the receptors with other plasma membrane components for MET (urokinase-type plasminogen activator in context of its receptor) and activation of STAT-3 for both receptors.

Adolescent↗
PubMed1999

The secondary fungal metabolite gliotoxin targets proteolytic activities of the proteasome.

BACKGROUND: The fungal epipolythiodioxopiperazine metabolite gliotoxin has a variety of toxic effects such as suppression of antigen processing, induction of macrophagocytic apoptosis and inhibition of transcription factor NF-kappaB activation. How gliotoxin acts remains poorly understood except that the molecule's characteristic disulfide bridge is important for immunomodulation. As this fungal metabolite stabilizes the NF-kappaB inhibitor IkappaBalpha in the cytoplasm, we decided to investigate its molecular mechanism of action. RESULTS: We show that gliotoxin is an efficient, noncompetitive inhibitor of the chymotrypsin-like activity of the 20S proteasome in vitro. Proteasome inhibition can be reversed by dithiothreitol, which reduces gliotoxin to the dithiol compound. In intact cells, gliotoxin inhibits NF-kappaB induction through inhibition of proteasome-mediated degradation of IkappaBalpha. CONCLUSIONS: Gliotoxin targets catalytic activities of the proteasome efficiently. Inhibition by gliotoxin may be countered by reducing agents, which are able to inactivate the disulfide bridge responsible for the inhibitory capacity of gliotoxin.

Cells, Cultured↗
PubMed1999

Seroprevalence of human herpesvirus-8 (HHV-8) in countries of Southeast Asia compared to the USA, the Caribbean and Africa.

Seroprevalence of HHV-8 has been studied in Malaysia, India, Sri Lanka, Thailand, Trinidad, Jamaica and the USA, in both healthy individuals and those infected with HIV. Seroprevalence was found to be low in these countries in both the healthy and the HIV-infected populations. This correlates with the fact that hardly any AIDS-related Kaposi's sarcoma has been reported in these countries. In contrast, the African countries of Ghana, Uganda and Zambia showed high seroprevalences in both healthy and HIV-infected populations. This suggests that human herpes virus-8 (HHV-8) may be either a recently introduced virus or one that has extremely low infectivity. Nasopharyngeal and oral carcinoma patients from Malaysia, Hong Kong and Sri Lanka who have very high EBV titres show that only 3/82 (3.7%) have antibody to HHV-8, demonstrating that there is little, if any, cross-reactivity between antibodies to these two gamma viruses.

Adolescent↗
PubMed1999

Deletions and losses in chromosomes 5 or 7 in adult acute lymphocytic leukemia: incidence, associations and implications.

Deletions or losses in chromosomes 5 or 7 are recurrent non-random abnormalities in acute myeloid leukemia (AML), and are associated with prior exposure to carcinogens or leukemogenic agents, and with poor prognosis. Their occurrence and significance in adult acute lymphocytic leukemia (ALL) is not well described. The aim of the study was to evaluate the incidence, associations and implications of chromosome 5 or 7 abnormalities in adult ALL. Patients with newly diagnosed ALL referred to MD Anderson Cancer Center between 1980 and 1996 were analyzed. Characteristics and outcome of patients with or without chromosome 5 or 7 abnormalities were compared by standard statistical methods. Thirty-one of 468 patients (6.6%) had chromosome 5 or 7 abnormalities. Loss of chromosome 5 occurred in six cases, three of them had both chromosome 5 and 7 abnormalities. Deletion or loss of chromosome 7 occurred as a single abnormality in three patients; in 28 patients it was associated with other abnormalities. The most significant cytogenetic association was with the Philadelphia chromosome (Ph) abnormality occurring in nine patients (29%). Compared with patients without the abnormalities, patients with chromosome 5 or 7 abnormalities tended to express CD34 more frequently (74% vs 54% P = 0.07), to be older (age >60 years 29% vs 18% P = 0.14), and to be associated with Ph (29% vs 11% P = 0.004). With therapy, the complete response (CR) rate with chromosome 5 or 7 abnormalities was lower (64% vs 79% P = 0.038) but the survival rate was similar (3-year survival rate 32% vs 36% P = 0.14). When the 22 patients without Ph were considered separately, the CR and survival rates were similar among patients with or without chromosome 5 or 7 abnormalities. Abnormalities in chromosome 5 or 7 are not specific for AML, and may occur in ALL. Unlike in AML, chromosome 5 or 7 abnormalities in ALL were not predictive of worse prognosis, which is accounted for mostly by the association with Ph.

Adolescent↗
PubMed1999

Vascular surgical society of great britain and ireland: transcranial doppler ultrasonography as a predictor of haemodynamically significant carotid stenosis

BACKGROUND: Transcranial Doppler (TCD) ultrasonography can detect evidence of collateral flow across the anterior communicating artery and/or the posterior communicating artery, which occurs when there is significant alteration of 'inflow' to the brain. The aim of the study was to determine the blood flow velocity produced by a carotid stenosis which produces this haemodynamic effect on the cerebral circulation and evokes collateral circulation. METHODS: Forty-eight patients with varying degrees of carotid stenosis (10 per cent to occlusion) who underwent both carotid duplex and TCD examination were reviewed. An ATL HDI 3000 ultrasound system was used for the carotid and TCD studies. The carotid examination recorded peak-systolic velocity (PSV) and end-diastolic velocity (EDV) in the carotid systems bilaterally. TCD recorded Doppler spectra from the bilateral middle cerebral, anterior cerebral, posterior cerebral, intracranial vertebral and basilar arteries. Collateral flow was assessed in two ways: 'intracranial crossover' collateral and 'posterior to anterior' collateral. Each internal carotid artery (ICA), together with the ipsilateral hemisphere, was analysed for the presence or absence of collateral flow. Data were expressed as mean(s.e.m.). RESULTS: The PSV of the group with collateral circulation was 472(14) cm s-1 and that of the group without collateral flow was 164(3) cm s-1 (P < 0.0001, Mann-Whitney test). The respective EDVs were 158(13) and 58(7) cm s-1 (P < 0.0001). CONCLUSION: PSVs and EDVs in the ICA, in conjunction with collateral flow measured by TCD, are indicators of a haemodynamically significant carotid lesion, and provide more information than two-dimensional imaging studies. In the future, parameters set by combining carotid duplex and TCD investigations may represent the 'gold standard' for evaluation of cerebral blood flow.

Journal Article↗