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R Newton

Publications and source records attributed to R Newton.

At least 91 records · Page 5Linked to original sources
PubMed1998

Effect of dexamethasone on interleukin-1beta-(IL-1beta)-induced nuclear factor-kappaB (NF-kappaB) and kappaB-dependent transcription in epithelial cells.

The production of inflammatory mediators by epithelial cells in inflammatory lung diseases may represent an important target for the anti-inflammatory effects of glucocorticoids. Nuclear factor-kappaB (NF-kappaB) is a major activator of inflammatory genes and has been proposed as a target for inhibition by glucocorticoids. We have used human pulmonary type-II A549 and airway epithelial BEAS-2B cells to investigate the effect of glucocorticoids on NF-kappaB regulation and kappaB-dependent transcription. In A549 cells following interleukin-1beta (IL-1beta) treatment, there was no effect of dexamethasone on the disappearance of I kappaB alpha protein, its subsequent reappearance 90-min later or the rapid induction of I kappaB alpha mRNA and transcription rate. Expression of p65 and p50/p105 proteins were also unaffected by dexamethasone. In addition, the rapid IL-1beta-induction of NF-kappaB DNA binding and p65 nuclear localisation was unaffected by short (1-6 hours) dexamethasone pre-treatments. Similarly, BEAS-2B cells showed no effect of dexamethasone on IL-1beta-induced NF-kappaB (p50/p65). Stable transfection of a kappaB-dependent reporter in A549 cells resulted in an 8-9-fold activation by IL-1beta or phorbol ester, that was repressed 30-40% by dexamethasone. However, in these cells, IL-1beta induction of inducible nitric oxide synthase, granulocyte-macrophage colony stimulating factor and cyclooxygenase-2 mRNA showed 70-90% repression by dexamethsone. We, therefore, conclude that in these epithelial cells, the repressive effects of glucocorticoids are not mediated by up-regulation of I kappaB alpha, decreased p50/p65 gene expression or inhibition of NF-kappaB DNA binding. Furthermore, since the maximal repression of IL-1beta or phorbol-ester-induced kappaB-dependent transcription by dexamethasone was less than 40%, simple inhibition of kappaB-dependent transcription cannot by itself account for the full repressive effects of glucocorticoids observed in these cells.

Cell Line↗
PubMed1998

IkappaBalpha degradation and nuclear factor-kappaB DNA binding are insufficient for interleukin-1beta and tumor necrosis factor-alpha-induced kappaB-dependent transcription. Requirement for an additional activation pathway.

Two closely related IkappaBalpha kinases as well as the upstream kinase, NIK, which integrates interleukin-1beta (IL-1beta)- and tumor necrosis factor (TNF)-alpha-dependent activation of the transcription factor NF-kappaB have recently been described. However, in this emerging pathway the role of previously identified components of cytokine-induced NF-kappaB activation, namely phosphatidylcholine-specific phospholipase C and protein kinase C, remains unclear. We now show that, in A549 human alveolar epithelial cells, the activation of a stably transfected NF-kappaB-dependent reporter gene by TNF-alpha and IL-1beta is completely blocked by the phosphatidylcholine-specific phospholipase C inhibitor D609 and the protein kinase C inhibitor RO31-8220. However, IL-1beta-induced IkappaBalpha degradation as well as NF-kappaB nuclear translocation and DNA binding, as determined by Western blot and electro-mobility shift assay, respectively, are not affected by these inhibitors. A similar effect, although less pronounced, is observed with the p38 mitogen-activated protein kinase inhibitor SB 203580. On the basis of these data we propose the existence of a second signaling pathway induced by IL-1beta and TNF-alpha that is activated in parallel to the cascade leading to IkappaBalpha degradation and is specifically required for NF-kappaB-dependent transcriptional competency.

Cell Line↗
PubMed1998

Test-retest reliability of the Fukuda Stepping Test.

BACKGROUND AND PURPOSE: Although the Fukuda Stepping Test (FST) is commonly used to assess individuals with peripheral vestibular dysfunction or balance instability, test-retest reliability has not been examined. METHODS: Sixteen males and 14 females, ranging in age from 23 to 56 years (mean 32.8 years), with no known vestibular dysfunction were tested. The subjects stood in the center of two concentric circles with eyes blindfolded. They stepped in place with outstretched arms for the prescribed number of steps. Subjects were tested on two consecutive days. Their angle of rotation, angle of displacement and distance of displacement were measured. RESULTS: Inter-class correlation coefficients were moderate for the 50-step protocol but lower for the 100-step protocol. CONCLUSIONS: If practitioners use the FST they should use the 50-step protocol. In addition, practitioners should interpret the results of the FST with caution if it is used as a screening tool. The test should be used in conjunction with other tests for vestibular function.

Adult↗
PubMed1998

The geographical distribution of Kaposi's sarcoma and of lymphomas in Africa before the AIDS epidemic.

Estimated incidence rates are presented for three human immunodeficiency virus (HIV)-associated cancers [Kaposi's sarcoma (KS), Burkitt's lymphoma (BL) and other non-Hodgkin's lymphomas (NHLs)] from across the African continent, based on data collected before the HIV epidemic. Mapping of the rates and comparisons with a range of geographical variables indicate completely different distributions for KS and BL but a degree of similarity in the occurrence of Burkitt's lymphoma and other NHLs. Comparisons with rates elsewhere in the world suggest, most notably, that KS was as common in some regions of sub-Saharan Africa as was cancer of the colon in much of Western Europe. Comparison with data from the era of AIDS indicates 20-fold increases in the occurrence of Kaposi's sarcoma in Uganda and Zimbabwe. The highest rates for BL were three to four times the rates for leukaemia at young ages in Western populations, but the general incidence of other NHL was no higher than in the West and very low rates were indicated for much of southern Africa.

Adolescent↗
PubMed1998

Overview of the epidemiology of immunodeficiency-associated cancers.

Immunodeficiency, be it congenital, therapeutic, or infectious in origin, increases the risk of certain, but not all, types of cancer. A common feature of these cancers is that specific infectious agents appear to be important in their etiology, not only in immunodeficient subjects but also in the general population. People with acquired immunodeficiency syndrome (AIDS) are at an increased risk of Kaposi's sarcoma, non-Hodgkin's lymphoma, Hodgkin's disease, squamous cell carcinoma of the conjunctiva, and childhood leiomyosarcoma. It is striking that most of these cancers have been associated with specific human herpesvirus (HHV) infections: HHV-8 with Kaposi's sarcoma and the closely related Epstein-Barr virus with non-Hodgkin's lymphoma, Hodgkin's disease, and possibly also with childhood leiomyosarcoma. Moreover, similar associations between these viruses and cancer have been found, albeit inconsistently, in people who are not immunosuppressed. Further research is needed to establish whether the risk of other cancers is also increased in people with AIDS, although, if so, the cancers are likely to be rare or to have comparatively small associated relative risks. Existing evidence suggests that there may be no marked increase in the risk of two common cancers that are known to be caused by infectious agents--hepatocellular carcinoma and invasive carcinoma of the uterine cervix. The apparent lack of an increase in invasive cervical cancer is unexpected and needs further investigation, especially since the prevalence of cervical infection with human papillomaviruses and of low-grade preneoplastic changes in the cervical epithelium is increased in women with AIDS. With the prospect of improved survival in people with AIDS, the effect of immunosuppression on cancer is likely to become an increasingly important issue.

Acquired Immunodeficiency Syndrome↗
PubMed1998

Paraoxonase active site required for protection against LDL oxidation involves its free sulfhydryl group and is different from that required for its arylesterase/paraoxonase activities: selective action of human paraoxonase allozymes Q and R.

Human serum paraoxonase (PON 1) exists in 2 major polymorphic forms (Q and R), which differ in the amino acid at position 191 (glutamine and arginine, respectively). These PON allozymes hydrolyze organophosphates and aromatic esters, and both also protect LDL from copper ion-induced oxidation. We have compared purified serum PONs of both forms and evaluated their effects on LDL oxidation, in respect to their arylesterase/paraoxonase activities. Copper ion-induced LDL oxidation, measured by the production of peroxides and aldehydes after 4 hours of incubation, were reduced up to 61% and 58%, respectively, by PON Q, but only up to 46% and 38%, respectively, by an equivalent concentration of PON R. These phenomena were PON-concentration dependent. Recombinant PON Q and PON R demonstrated similar patterns to that shown for the purified serum allozymes. PON Q and PON R differences in protection of LDL against oxidation were further evaluated in the presence of glutathione peroxidase (GPx). GPx (0.1 U/mL) alone reduced copper ion-induced LDL oxidation by 20% after 4 hours of incubation. The addition of PON R to the above system resulted in an additive inhibitory effect on LDL oxidation, whereas PON Q had no such additive effect. The 2 PON allozymes also differed by their ability to inhibit initiation, as well as propagation, of LDL oxidation. PON Q was more efficient in blocking LDL oxidation if added when oxidation was initiated, whereas PON R was more potent when added 1 hour after the initiation of LDL oxidation. These data suggest that the 2 allozymes act on different substrates. Both PON allozymes were also able to reduce the oxidation of phospholipids and cholesteryl ester. PON Q arylesterase activity was reduced after 4 hours of LDL oxidation by only 28%, whereas the arylesterase activity of PON R was reduced by up to 55%. Inactivation of the calcium-dependent PON arylesterase activity by using the metal chelator EDTA, or by calcium ion removal on a Chelex column, did not alter PON's ability to inhibit LDL oxidation. However, blockage of the PON free sulfhydryl group at position 283 with p-hydroxymercuribenzoate inhibited both its arylesterase activity and its protection of LDL from oxidation. Recombinant PON mutants in which the PON free sulfhydryl group was replaced by either alanine or serine were no longer able to protect against LDL oxidation, even though they retained paraoxonase and arylesterase activities. Overall, these studies demonstrate that PON's arylesterase/paraoxonase activities and the protection against LDL oxidation do not involve the active site on the enzyme in exactly the same way, and PON's ability to protect LDL from oxidation requires the cysteine residue at position 283.

Aryldialkylphosphatase↗
PubMed1998

Effects of atorvastatin on the intracellular stability and secretion of apolipoprotein B in HepG2 cells.

We investigated the effects of atorvastatin, a new 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, on the biogenesis of apolipoprotein B (apoB) in intact and permeabilized HepG2 cells. Intact cells were pretreated either with single or multiple doses of atorvastatin (0.1 to 20 micromol/L) for periods of 6 to 20 hours and pulsed with [35S]methionine. In some cases the cells were permeabilized with digitonin. Experiments were performed to investigate the effects of atorvastatin on (1) the rates of lipid synthesis and secretion, (2) the synthesis and accumulation of apoB, (3) the intracellular stability of apoB, (4) the amount of apoB-containing lipoprotein particles assembled in HepG2 microsomes, and (5) the secretion and accumulation of apoB into the culture medium. ApoB synthesis, degradation, and secretion were measured by pulse-chase experiments with [35S]methionine in both intact and permeabilized HepG2 cells. Lipid synthesis was assessed by pulse-labeling experiments with [3H]acetate or [3H]oleate bound to bovine serum albumin. Comparisons were made under basal conditions and in the presence of oleate (0.36 micromol/L). Atorvastatin acutely inhibited the synthesis of cholesterol and cholesterol ester but did not have a significant effect on triglyceride or phospholipid synthesis. Atorvastatin did not affect the uptake of [35S]methionine by the cells nor did it influence the synthesis of apoB or a control protein, albumin. However, atorvastatin reduced the secretion of apoB into the culture medium, apparently by enhancing the degradation of apoB in the cell under basal and induced conditions with oleate. The stability of apoB associated with the lipoprotein particles was also significantly lowered by atorvastatin. The stimulated degradation of apoB in atorvastatin-treated cells was sensitive to MG132, a proteasome inhibitor. The net effect of atorvastatin was a reduction in the number of apoB-containing lipoprotein particles of different sizes isolated from microsomes and a reduction in apoB secretion into the culture medium. The data suggest that atorvastatin may impair the translocation of apoB into the lumen of the endoplasmic reticulum, thus increasing the amount of apoB degraded intracellularly. It is hypothesized that atorvastatin alters these parameters primarily as a result of inhibiting cholesterol synthesis and limiting the availability of cholesterol and/or cholesterol ester for the normal assembly of apoB-containing lipoprotein particles.

Albumins↗
PubMed1998

Effect of inhibition of nitric oxide synthase on Pseudomonas aeruginosa infection of respiratory mucosa in vitro.

We studied the effect of the nitric oxide synthase (NOS) inhibitor asymmetric dimethyl arginine (ADMA) and the inactive enantiomer N G-methyl-D-arginine (D-NMMA) on Pseudomonas aeruginosa infection of the respiratory mucosa in nasal turbinate organ cultures. We also investigated the effect of P. aeruginosa culture filtrate on the expression of inducible NOS (iNOS) messenger RNA (mRNA) by an epithelial cell line (A549). Organ cultures were preincubated with ADMA (0.1 to 4 x 10(-4) M) or D-NMMA (2 x 10(-4) M) for 30 min prior to bacterial infection. Infected organ cultures (8 h) had significantly (P <= 0.05) greater epithelial damage and fewer ciliated and unciliated cells than did control cultures. There was an increased level of nitrite in the medium feeding infected organ cultures as compared with control cultures. ADMA significantly (P <= 0.05) reduced both bacterially induced epithelial damage and loss of ciliated cells in a concentration-dependent manner. D-NMMA did not influence the effect of P. aeruginosa infection of the mucosa. ADMA, but not D-NMMA, significantly (P <= 0.04) reduced total bacterial numbers adherent to the respiratory mucosa. P. aeruginosa culture filtrates (24 h and 36 h) significantly (P = 0.02) increased iNOS with respect to glyceraldehyde-3-phosphate dehydrogenase mRNA expression. These results show that P. aeruginosa stimulates iNOS expression by a cell line and NO production by an organ culture. ADMA reduces mucosal damage and loss of ciliated cells, which suggests that NO may be a mediator of epithelial damage caused by P. aeruginosa.

Arginine↗
PubMed1997

Superinduction of COX-2 mRNA by cycloheximide and interleukin-1beta involves increased transcription and correlates with increased NF-kappaB and JNK activation.

Many primary response genes, including cyclooxygenase-2 (COX-2), exhibit mRNA superinduction following agonist stimulation in the presence of translational blockers such as cycloheximide. This is widely assumed to result from mRNA stabilisation. However, superinduction of IL-1beta-induced COX-2 mRNA levels by cycloheximide in pulmonary type II A549 cells occurred by increased transcription and not by mRNA stabilisation. Furthermore, equivalent effects were observed on NF-kappaB binding to COX-2 promoter kappaB sites and activation of the Jun N-terminal kinases (JNK), p54 and p46. These signalling pathways play important roles in COX-2 induction and may therefore account for the observed increases in COX-2 transcription. These data are consistent with negative feed-back involving down-regulation of NF-kappaB by de novo IkappaB alpha synthesis and suggest that JNK activation may also be down-regulated by a cycloheximide sensitive process.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1997

The epidemiology of non-Hodgkin's lymphoma: comparison of nodal and extra-nodal sites.

International population-based cancer incidence data, coded according to the International Classification of Diseases for Oncology (WHO, 1990), were used to describe geographical patterns of incidence of extra-nodal non-Hodgkin's lymphomas. Incidence data from the USA were also used to describe age and sex distribution of lymphomas at different extranodal sites. The percentage of all non-Hodgkin's lymphomas coded as being of extra-nodal origin is between 25% and 35% in most countries, with the stomach, skin and small intestine being the most common extra-nodal sites. In general, the pattern of incidence rates for extra-nodal lymphomas tends to reflect that of other lymphomas. For example, the age incidence curve of each site-specific extra-nodal lymphoma is similar to that of nodal lymphomas, and in countries where total lymphoma incidence is high the incidence of lymphomas at each extra-nodal site also tends to be relatively high. Although specific factors are known to increase the risk of lymphomas at certain anatomical sites, these data suggest that the aetiology of extra-nodal lymphomas is not entirely independent from that of nodal lymphomas.

Adult↗
PubMed1997

Evidence for involvement of NF-kappaB in the transcriptional control of COX-2 gene expression by IL-1beta.

The cyclooxygenase (COX) isoforms COX-1 and COX-2 convert arachidonic acid to prostaglandin (PG) precursors and are a limiting step in PG production. Interleukin-1beta (IL-1beta) treatment of type II A549 cells increases PGE2 synthesis via transcription- and translation-dependent induction of COX-2. IL-1beta produces a 10-fold induction of COX-2 mRNA and an 8-fold increase in COX-2 transcription that was temporally preceded by activation of the transcription factor nuclear factor-kappaB (NF-kappaB). The protein-tyrosine phosphatase inhibitor phenylarsine oxide (PAO) prevented both NF-kappaB activation and induction of COX-2 mRNA. We show that two putative NF-kappaB motifs, kappaBu (-447/-438) and kappaBd (-224/-214), from the COX-2 promoter bind p50/p65 NF-kappaB heterodimers in an IL-1beta-dependent manner and that the upstream element has the greater affinity. Finally, we demonstrate that the two NF-kappaB subunits, p50 and p65, synergistically activate a -917/+49 COX-2 promoter construct. We conclude that IL-1beta stimulates PG production via transcriptional activation of COX-2 and provide evidence that this may involve NF-kappaB.

Arsenicals↗
PubMed1997

Alternate COX-2 transcripts are differentially regulated: implications for post-transcriptional control.

Prostaglandin (PG) synthesis during inflammation occurs mainly via the transcriptionally regulated cyclooxygenase, COX-2. In pulmonary type II A549 cells, Northern analysis identified multiple IL-1 beta-inducible COX-2 mRNA transcripts. Amplification of 3'-cDNA ends by anchored PCR revealed products corresponding to the predominant 4.5 and 2.7 kb transcripts. Sequence analysis of amplification products indicated that these transcripts arose by alternate consensus and non-consensus polyadenylation site usage. The predominant 4.5 kb transcript showed a half-life in excess of two hours that was further stabilized by IL-1 beta. In addition, the COX-2 3'-untranslated region (UTR), which contains 22 copies of the putative RNA instability motif, AUUUA, when cloned downstream of a constitutively expressed luciferase gene, was found to confer partial IL-1 beta responsiveness in LA-4 cells. Finally, in vivo in LPS-treated rats, differential expression of similar COX-2 mRNA isoforms was also observed. Taken together these data suggest a functional role for post-transcriptional mechanisms, including alternate polyadenylation, in the control of COX-2.

Animals↗
PubMed1997

Cytokine induction of cytosolic phospholipase A2 and cyclooxygenase-2 mRNA is suppressed by glucocorticoids in human epithelial cells.

Prostaglandin (PG) release, which is increased in vivo by inflammatory conditions and in vitro by pro-inflammatory cytokines, is decreased by glucocorticoids. Two phospholipase A2 isoforms, secretory (sPLA2) and cytosolic (cPLA2,), have been implicated in inflammation. These enzymes catalyse the release of arachidonic acid which is then converted to prostaglandins by the cyclooxygenases (COX-1 and COX-2). Regulation of these events at the mRNA level is poorly characterised in epithelial cells. We have used a human epithelial-like cell line (A549) as a model system to study mRNA expression of sPLA2, cPLA2, COX-1 and COX-2. Following treatment of cells and extraction of RNA, semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) was used to examine expression of these genes. We show a coordinate induction of both cPLA2 and COX-2 mRNA by pro-inflammatory cytokines which correlated with increased PGE2 release. By contrast, sPLA2 mRNA was undetectable and COX-1 was found to be expressed at a constant low level. In addition dexamethasone pretreatment significantly reduced both cPLA2 and COX-2 mRNA levels as well as PGE2 release following cytokine stimulation. These data indicate a major role for control of prostaglandin synthesis at the mRNA level of key synthetic genes in epithelial cells. Furthermore we show that a major mechanism of glucocorticoid action in preventing prostaglandin release occurs by suppression of cPLA2 and COX-2 mRNA levels.

Cycloheximide↗
PubMed1997

Risk factors for Kaposi's sarcoma in HIV-positive subjects in Uganda.

BACKGROUND: Kaposi's sarcoma (KS) is associated epidemiologically with HIV infection and with human herpesvirus 8 (HHV-8 or KSHV). Both KS and HIV infection are common in Uganda. We conducted a case-control study of 458 HIV-seropositive. Ugandan adults with KS and 568 HIV-seropositive subjects without KS to examine risk factors for HIV-associated KS. METHODS: We recruited newly diagnosed adult KS cases from five hospitals in Kampala, Uganda and controls from a large referral clinic for HIV infection at Mulago Hospital. All cases and controls were counselled and tested for HIV and answered an interviewer-administered questionnaire about their home, socio-economic conditions, lifestyle and sexual behaviour before they became ill. Only HIV-seropositive subjects were included in the analysis. RESULTS: There were 295 males and 163 females with KS and 227 male and 341 female controls. Age distribution was similar but there was a higher proportion of cases (45%) than controls (29%) residing in rural regions of Uganda. KS cases were more likely than controls to have a higher level of education (X2 for trend, 4.8; P = 0.03), to have occupations associated with affluence [chi 2 for heterogeneity, 17.3 on 5 degrees of freedom (df); P = 0.004] and to come from larger settlements [adjusted odds ratio (OR) for settlements of > 1000 versus 10-99 houses, 1.8; 95% confidence interval (CI), 1.1-3.0]. Cases were more likely than controls to have high household income (chi 2 for trend, 32.6; P < 0.001) and other markers of urban or rural wealth such as owning several cows (chi 2 for trend, 9.5; P = 0.002). Cases were more likely to travel away from home (adjusted OR, 1.6; 95% CI, 1.1-2.3) and more likely to have spent increasing time in contact with water (chi 2 for trend, 12.3; P < 0.001). Few indices of sexual behaviour were related to risk of KS, including reported number of sexual partners. Cases were more likely than controls to be married to one rather than several spouses (adjusted OR, 1.6; 95% CI, 1.2-2.2) and to have reported a history of sexually transmitted diseases (STD) (adjusted OR, 1.6; 95% CI, 1.2-2.3). CONCLUSIONS: Among HIV-infected subjects, KS cases are characterized by better education and greater affluence, compared with controls. Urban address, travel away from home, exposure to water, monogamous marriage and self-reported STD were also more frequent among KS cases than controls. The higher socio-economic status of persons with HIV and KS may be a marker for enhanced exposure to a possibly sexually transmitted agent, or for a delayed exposure to a childhood infection. The risk posed by exposure to water among KS cases requires further study.

AIDS-Related Opportunistic Infections↗
PubMed1997

Sodium salicylate inhibits cyclo-oxygenase-2 activity independently of transcription factor (nuclear factor kappaB) activation: role of arachidonic acid.

Acetylsalicylic acid (aspirin) is the drug most commonly self-administered to reduce inflammation, swelling, and pain. The established mechanism of action of aspirin is inhibition of the enzyme cyclo-oxygenase (COX). Once taken, aspirin is rapidly deacetylated to form salicylic acid, which may account, at least in part, for the therapeutic actions of aspirin. However, where tested, salicylic acid has been found to be a relatively inactive inhibitor of COX activity in vitro, despite being an effective inhibitor of prostanoids formed at the site of inflammation in vivo. Recently, the identification of a cytokine-inducible isoform of COX, COX-2, has led to the suggestion that salicylate produces its anti-inflammatory actions by inhibiting COX-2 induction through actions on nuclear factor kappaB (NF-kappaB). We have used interleukin 1beta-induced COX-2 in human A549 cells to investigate the mechanism of action of salicylate on COX-2 activity. Sodium salicylate inhibited prostaglandin E2 release when added together with interleukin 1beta for 24 hr with an IC50 value of 5 microg/ml, an effect that was independent of NF-kappaB activation or COX-2 transcription or translation. Sodium salicylate acutely (30 min) also caused a concentration-dependent inhibition of COX-2 activity measured in the presence of 0, 1, or 10 microM exogenous arachidonic acid. In contrast, when exogenous arachidonic acid was increased to 30 microM, sodium salicylate was a very weak inhibitor of COX-2 activity with an IC50 of >100 microg/ml. Thus, sodium salicylate is an effective inhibitor of COX-2 activity at concentrations far below those required to inhibit NF-kappaB (20 mg/ml) activation and is easily displaced by arachidonic acid.

Arachidonic Acid↗