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Biomedical subjects

A Lloyd

Publications and source records attributed to A Lloyd.

At least 37 records · Page 2Linked to original sources
PubMed2001

Regulatory role of IL-1beta in the expression of IL-6 and IL-8 in human corneal epithelial cells during Pseudomonas aeruginosa colonization.

Pseudomonas aeruginosa is a virulent pathogen and is frequently associated with bacterial keratitis. Recent studies have shown that high levels of interleukin (IL)-1beta and macrophage inflammatory protein-2 are associated with the severity of corneal infection. Interleukin-1beta is a principal inflammatory mediator. Understanding the regulatory role of IL-1beta would provide better understanding of host responses during P. aeruginosa corneal infection. A human corneal epithelial (HCE) cell line and three P. aeruginosa strains were used in this experiment. Confluent HCE cells were challenged with P. aeruginosa and monoclonal antihuman IL-1beta antibody (IL-1beta mAb). The culture supernatants were collected for measuring cytotoxicity and protein levels of IL-1beta, IL-8 and IL-6 by enzyme-linked immunosorbent assay. Results showed that HCE cells expressed low levels of IL-1beta and high levels of IL-6 and IL-8 during P. aeruginosa colonization. Paer1-colonized HCE cells produced higher levels of IL-1beta, IL-6 and IL-8 protein compared to those produced by 6206- and 6294-colonized HCE cells. Administration of IL-1beta mAb decreased the production of IL-8 and IL-6. In conclusion, P. aeruginosa-colonized HCE cells produced low levels of IL-1beta and high levels of IL-6 and IL-8. Neutralizing IL-1beta protein significantly downregulated the production of IL-8 and IL-6.

Cell Line↗
PubMed2001

The role of IL-1beta in the regulation of IL-8 and IL-6 in human corneal epithelial cells during Pseudomonas aeruginosa colonization.

PURPOSE: Recent studies have shown that the levels of interleukin (IL)-1beta, IL-6 and IL-8 are associated with the severity of infectious diseases. The purpose of this study was to investigate whether IL-1beta regulates the expression of IL-6 and IL-8 in human corneal epithelial cells during Pseudomonas aeruginosa colonization. METHODS: Confluent immortalized human corneal epithelial cells were challenged with P. aeruginosa 6294 in the presence of anti-human IL-1beta antibody or matched control antibody. The cells were also challenged with recombinant IL-1beta protein without bacterial colonization. Expression of IL-1beta, IL-6 and IL-8 mRNA and protein was detected by reverse transcription (RT)-PCR and by enzyme-linked immunosorbent assay (ELISA), respectively. IL-1beta localization was determined by immunohistochemistry. RESULTS: Human corneal epithelial cells expressed low levels of IL-1beta and high levels of IL-6 and IL-8 during P. aeruginosa colonization. Addition of IL-1beta Ab resulted in a significant decrease (P < 0.05) in IL-8 protein expression at 4 h, 8 h and 12 h. Addition of IL-1beta Ab reduced IL-6 protein expression at 8 h and increased IL-6 protein expression at 12 h. Addition of recombinant IL-1beta protein alone strongly stimulated the expression of IL-8 and IL-6. Immunohistochemical staining showed that IL-1beta protein was present both intracellularly and extracellularly in P. aeruginosa colonized cells. CONCLUSIONS: IL-1beta is able to modulate expression of both IL-6 and IL-8 at transcriptional and post-transcriptional levels in human corneal epithelial cells.

Cells, Cultured↗
PubMed2001

Antisense expression of an Arabidopsis ran binding protein renders transgenic roots hypersensitive to auxin and alters auxin-induced root growth and development by arresting mitotic progress.

We cloned a cDNA encoding an Arabidopsis Ran binding protein, AtRanBP1c, and generated transgenic Arabidopsis expressing the antisense strand of the AtRanBP1c gene to understand the in vivo functions of the Ran/RanBP signal pathway. The transgenic plants showed enhanced primary root growth but suppressed growth of lateral roots. Auxin significantly increased lateral root initiation and inhibited primary root growth in the transformants at 10 pM, several orders of magnitude lower than required to induce these responses in wild-type roots. This induction was followed by a blockage of mitosis in both newly emerged lateral roots and in the primary root, ultimately resulting in the selective death of cells in the tips of both lateral and primary roots. Given the established role of Ran binding proteins in the transport of proteins into the nucleus, these findings are consistent with a model in which AtRanBP1c plays a key role in the nuclear delivery of proteins that suppress auxin action and that regulate mitotic progress in root tips.

Amino Acid Sequence↗
PubMed2001

The MCP silencer of the Drosophila Abd-B gene requires both Pleiohomeotic and GAGA factor for the maintenance of repression.

Silencing of homeotic gene expression requires the function of cis-regulatory elements known as Polycomb Response Elements (PREs). The MCP silencer element of the Drosophila homeotic gene Abdominal-B has been shown to behave as a PRE and to be required for silencing throughout development. Using deletion analysis and reporter gene assays, we defined a 138 bp sequence within the MCP silencer that is sufficient for silencing of a reporter gene in the imaginal discs. Within the MCP138 fragment, there are four binding sites for the Pleiohomeotic protein (PHO) and two binding sites for the GAGA factor (GAF), encoded by the Trithorax-like gene. PHO and the GAF proteins bind to these sites in vitro. Mutational analysis of PHO and GAF binding sequences indicate that these sites are necessary for silencing in vivo. Moreover, silencing by MCP138 depends on the function of the Trithorax-like gene, and on the function of the PcG genes, including pleiohomeotic. Deletion and mutational analyses show that, individually, either PHO or GAF binding sites retain only weak silencing activity. However, when both PHO and GAF binding sites are present, they achieve strong silencing. We present a model in which robust silencing is achieved by sequential and facilitated binding of PHO and GAF.

Animals↗
PubMed2001

Reliability and sensitivity of joint space measurements in hand radiographs using computerized image analysis.

OBJECTIVE: To establish the sensitivity and reliability of proximal interphalangeal (PIP) and metacarpophalangeal (MCP) mean joint space measurements using standard clinical radiographs of healthy subjects, in order to determine the limits at which a change in radiographic joint space could indicate a change in actual joint size. METHODS: Repeat hand radiographs of healthy subjects were taken using standard techniques at 3-5 day intervals with the hands flat (5 posteroanterior radiographs in 8 subjects) or in 6 different flexed positions on a single occasion (8 subjects). The mean joint space was determined using custom soft ware and was validated manually. Measurement reproducibility within subjects, within films, and between hand positions was assessed by analysis of variance. RESULTS: In repeat radiographs taken in the standard clinical position, the precision of individual join space measurements indicates that changes > 0.11 mm (approximately 7%) would represent an actual physical change in joint space width (with 95% probability). Averaging measurements across fingers for a single subject decreases the detectable change to 0.05 mm (approximately 3%). With increasing flexure, radiographic joint space tended to increase in MCP and decrease in PIP. CONCLUSION: Mean finger joint space measured from standard clinical radiographs is a reliable and sensitive measurement in healthy subjects even with some change in hand position. Work is required to establish whether the joint space change measured from serial radiographs of patients with arthritis over a period of 6-12 mo exceeds the detectable limits of change derived in this study.

Adult↗
PubMed2000

Membrane impermeant antioestrogens discriminate between ligand- and voltage-gated cation channels in NG108-15 cells.

Native 5-HT(3) and AChR ligand-gated cation channels can be inhibited (blocked) by the non-steroidal antioestrogen tamoxifen. However, the exact site and mechanism of inhibition by tamoxifen on these channels remain unclear. We have investigated the action of the membrane impermeant quaternary derivative, ethylbromide tamoxifen (EBT), on native ligand-gated 5-HT(3) receptor channels and voltage-gated K(+) channels in NG108-15 cells using whole cell patch clamp. Extracellular EBT inhibited whole cell cationic currents of 5-HT(3) receptors with IC(50) of 0.22+/-0.4 microM (n(H)=1.05+/-0.2). The channel block was characterised by voltage independent and use independent behaviour (similar to that of tamoxifen). EBT was unable to inhibit voltage-gated K(+) currents in NG108-15 cells. This was in contrast to the inhibition by tamoxifen which, at similar concentrations, accelerated the apparent inactivation of these outward K(+) currents. The inhibition of 5-HT(3) receptors by a membrane impermeant derivative of tamoxifen supports the view that the binding site for antioestrogens is extracellular and the inhibition is not mediated through genomic/transcriptional activity.

Animals↗
PubMed2000

Monitor: molecules and profiles.

Monitor provides an insight into the latest developments in drug discovery through brief synopses of recent presentations and publications together with expert commentaries on the latest technologies. There are two sections: Molecules summarizes the chemistry and the pharmacological significance and biological relevance of new molecules reported in the literature and on the conference scene; Profiles offers commentary on promising lines of research, emerging molecular targets, novel technology, advances in synthetic and separation techniques and legislative issues.

Journal Article↗
PubMed2000

Farewell.

Explore the source record for details and available documents.

Journal Article↗
PubMed2000

Monitor: molecules and profiles.

Monitor provides an insight into the latest developments in drug discovery through brief synopses of recent presentations and publications together with expert commentaries on the latest technologies. There are two sections: Molecules summarizes the chemistry and the pharmacological significance and biological relevance of new molecules reported in the literature and on the conference scene; Profiles offers commentary on promising lines of research, emerging molecular targets, novel technology, advances in synthetic and separation techniques and legislative issues.

Journal Article↗
PubMed2000

Long-term persistence of Coxiella burnetii in the host after primary Q fever.

After a primary infection Coxiella burnetii may persist covertly in animals and recrudesce at parturition to be shed in the products of conception and the milk. Similar latent persistence and recrudescence occurs in man: namely, infection of placenta, heart valve or mural endocardium, bone or liver. The numbers of organisms, their viability and cellular form, and the underlying organ sites of latent infection for the coxiella are obscure. During investigations of 29 patients with a chronic sequel to acute Q fever, the post-Q fever fatigue syndrome (QFS) [1-3], sensitive conventional and TaqMan-based PCR revealed low levels of C. burnetii DNA in blood mononuclear cells (5/29; 17%), thin needle liver biopsies (2/14; 14%) and, notably, in bone marrow aspirates (13/20; 65%). Irrespective of the ultimate significance of coxiella persistence for QFS, the detection of C. burnetii genomic DNA in bone marrow several years after a primary infection unveils a new pathological dimension for Q fever.

Bone Marrow↗
PubMed2000

A role for ectophosphatase in xenobiotic resistance.

Xenobiotic resistance in animals, plants, yeast, and bacteria is known to involve ATP binding cassette transporters that efflux invading toxins. We present data from yeast and a higher plant indicating that xenobiotic resistance also involves extracellular ATP degradation. Transgenic upregulation of ecto-ATPase alone confers resistance to organisms that have had no previous exposure to toxins. Similarly, cells that are deficient in extracellular ATPase activity are more sensitive to xenobiotics. On the basis of these and other supporting data, we hypothesize that the hydrolysis of extracellular ATP by phosphatases and ATPases may be necessary for the resistance conferred by P-glycoprotein.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2000

Mutants of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase resistant to nonnucleoside reverse transcriptase inhibitors demonstrate altered rates of RNase H cleavage that correlate with HIV-1 replication fitness in cell culture.

Three mutants of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (V106A, V179D, and Y181C), which occur in clinical isolates and confer resistance to nonnucleoside reverse transcriptase inhibitors (NNRTIs), were analyzed for RNA- and DNA-dependent DNA polymerization and RNase H cleavage. All mutants demonstrated processivities of polymerization that were indistinguishable from wild-type enzyme under conditions in which deoxynucleoside triphosphates were not limiting. The V106A reverse transcriptase demonstrated a three- to fourfold slowing of both DNA 3'-end-directed and RNA 5'-end-directed RNase H cleavage relative to both wild-type and V179D enzymes, similar to what was observed for P236L in a previously published study (P. Gerondelis et al., J. Virol. 73:5803-5813, 1999). In contrast, the Y181C reverse transcriptase demonstrated a selective acceleration of the secondary RNase H cleavage step during both modes of RNase H cleavage. The relative replication fitness of these mutants in H9 cells was assessed in parallel infections as well as in growth competition experiments. Of the NNRTI-resistant mutants, V179D was more fit than Y181C, and both of these mutants were more fit than V106A, which demonstrated the greatest reduction in RNase H cleavage. These findings, in combination with results from previous work, suggest that abnormalities in RNase H cleavage are a common characteristic of HIV-1 mutants resistant to NNRTIs and that combined reductions in the rates of DNA 3'-end- and RNA 5'-end-directed cleavages are associated with significant reductions in the replication fitness of HIV-1.

Cells, Cultured↗
PubMed2000

Expression of MMPs and TIMPs in human pterygia and cultured pterygium epithelial cells.

PURPOSE: Pterygia are a common, benign, fibrovascular, and infiltrative process of the corneal-conjunctival junction of unknown pathogenesis. Matrix metalloproteinases (MMPs) are a family of proteolytic enzymes active against all components of the extracellular matrix, whose activity is specifically neutralized by tissue inhibitors of MMPs (TIMPs). In the current study the hypothesis was that MMPs and TIMPs may actively participate in the formation and progression of pterygia. METHODS: In this study, 25 pterygium specimens and 15 normal conjunctival biopsies obtained from subjects undergoing surgery for glaucoma and cataract, were processed for immunohistochemistry or in situ hybridization. Pterygium epithelial cells (PECs) were cultured under serum-free conditions and exposed to proinflammatory cytokines to determine both the mRNA and protein expression profiles of MMPs and TIMPs. RESULTS: Collagenase-1 and gelatinase A were expressed in all pterygia examined, specifically localized to the epithelium (directly adjacent to collagen type III), with gelatinase B expression exclusively associated with neutrophils. No collagenase-1 or gelatinase A was detected in normal conjunctiva. TIMP-1 and -3 were localized to epithelial cells with additional TIMP-3 immunoreactivity detected in the extracellular matrix, endothelial cells and leukocytes of all diseased tissue. TIMP-3 protein was evident in 4 of 15 normal conjunctiva. Induction of collagenase-1, gelatinase A, and TIMP-1 mRNA and protein was demonstrated in epithelial cells treated with tumor necrosis factor-alpha and interleukin-1alpha, whereas TIMP-3 expression was unaltered. CONCLUSIONS: This is the first study to document the cellular expression of MMPs and TIMPs in pterygia and cultured human PECs. MMPs and TIMPs may contribute to the inflammation, tissue remodeling, and angiogenesis that characterize pterygia. Understanding the role these proteins play may lead to novel therapies intended to reduce the progressive nature of pterygia.

Blotting, Western↗
PubMed1999

Abundant expression of chemokines in malignant and infective human lymphadenopathies.

Lymph nodes can be the primary target of infection or malignant transformation and may exhibit characteristic patterns of leukocyte infiltration analogous to those seen in inflammation of other tissues. Leukocyte migration to lymph nodes in vivo is a highly regulated, multi-step process that depends upon adhesion molecules and as yet, uncharacterized chemotactic signals. Chemokines are a key part of the orchestrated code of signals that directs leukocyte subsets to sites of inflammation or immune response. The potential role of these chemoattractants in selective trafficking of leukocyte subsets into lymph nodes was assessed by determining the expression of chemokines on a range of pathological and normal human lymph nodes and by evaluating the cellular composition of each lymph node. In situ hybridization using chemokine riboprobes and immunohistochemistry using specific antibodies were performed in order to correlate the mRNA and protein expression of the chemokines. The cellular source(s) of each chemokine was assessed by immunohistochemical staining of adjacent sections using antibodies directed against distinctive cellular markers. Substantial, but varied, expression of macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, RANTES, macrophage chemotactic protein (MCP)-1, eotaxin, and interleukin 8 (IL-8) were detected in the pathological lymph nodes by diverse cell types. Control lymph nodes showed expression only of RANTES, mainly by high endothelial venules. In all lymph nodes, except the nodes infiltrated with breast cancer, chemokine mRNA expression was highly concordant with the corresponding protein. In contrast with in vitro studies that have suggested discrete target cell specificity of chemokines, this study showed that with the possible exception of the neutrophil chemoattractant, IL-8, no chemokine appeared to be uniquely associated with the accumulation of a specific leukocyte subset. These data implicate chemokines in the recruitment of leukocytes to lymph nodes affected by diverse disease states.

Antigens, CD↗
PubMed1999

Prevalence of true vein graft aneurysms: implications for aneurysm pathogenesis.

BACKGROUND: Circumstantial evidence suggests that arterial aneurysms have a different cause than atherosclerosis and may form part of a generalized dilating diathesis. The aim of this study was to compare the rates of spontaneous aneurysm formation in vein grafts performed either for popliteal aneurysms or for occlusive disease. The hypothesis was that if arterial aneurysms form a part of a systemic process, then the rates of vein graft aneurysms should be higher for patients with popliteal aneurysms than for patients with lower limb ischemia caused by atherosclerosis. METHODS: Infrainguinal vein grafting procedures performed from 1990 to 1995 were entered into a prospective audit and graft surveillance program. Aneurysmal change was defined as a focal increase in the graft diameter of 1.5 cm or greater, excluding false aneurysms and dilatations after graft angioplasty. RESULTS: During the study period, 221 grafting procedures were performed in 200 patients with occlusive disease and 24 grafting procedures were performed in 21 patients with popliteal aneurysms. Graft surveillance revealed spontaneous aneurysm formation in 10 of the 24 bypass grafts (42%) for popliteal aneurysms but in only 4 of the 221 grafting procedures (2%) that were performed for chronic lower limb ischemia. CONCLUSION: This study provides further evidence that aneurysmal disease is a systemic process, and this finding has clinical implications for the treatment of popliteal aneurysms.

Adult↗