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Biomedical subjects

E Beck

Publications and source records attributed to E Beck.

At least 37 records · Page 2Linked to original sources
PubMed2000

Tools for discovery of inhibitors of the 1-deoxy-D-xylulose 5-phosphate (DXP) synthase and DXP reductoisomerase: an approach with enzymes from the pathogenic bacterium Pseudomonas aeruginosa.

Two Pseudomonas aeruginosa genes encoding the enzymes 1-deoxy-D-xylulose 5-phosphate (DXP) synthase and DXP reductoisomerase, both involved in the mevalonate-independent biosynthesis of isoprenoids, have been expressed as recombinant enzymes in Escherichia coli. The purified P. aeruginosa DXP reductoisomerase was inhibited by submicromolar concentrations of the antibiotics fosmidomycin and FR-900098 in a well established method. A novel and convenient spectrophotometric assay was developed to determine activity and inhibition of P. aeruginosa DXP synthase. Fluoropyruvate is described as a first inhibitor of DXP synthase.

Aldose-Ketose Isomerases↗
PubMed2000

IL-18 potentiates the adjuvant properties of IL-12 in the induction of a strong Th1 type immune response against a recombinant antigen.

Due to the synergistic effects of IL-12 and IL-18, and to their importance in establishing a Th1 type immune response, we investigated the potential of a combined administration of both cytokines as an adjuvant for recombinant antigens. As a model system, we used a schistosome T cell antigen recently identified in our group. By co-adsorption of this antigen on alum in the presence of IL-12 and IL-18, we demonstrate that IL-18 enhances the effects of IL-12 in inducing an antigen-specific Th1 type CD4(+) T cell response as well as high titres of IgG1, IgG2a, and IgG2b antibodies.

Adjuvants, Immunologic↗
PubMed2000

Molecular characterization of a Litomosoides sigmodontis protein involved in the development of the microfilarial sheath during embryogenesis.

A cDNA clone Ls110 was isolated from a female Litomosoides sigmodontis expression library using an antiserum raised against the microfilarial sheath. The complete cDNA encodes a protein (Ls110) of 382 amino acids. Southern and PCR analyses revealed the presence of Ls110 in L. sigmodontis as a single copy gene. The transcription of the Ls110 gene was limited to female worms. In these worms the transcription was confined to the epithelial cells of the uterus. The protein Ls110 was detected not only in the epithelial layer of the uterus but also secreted in the lumen of the uterus. All the intra-uterine embryonic stages showed the protein bound to their egg shell/sheath, except the early multicellular embryonic stages and fully developed microfilariae. The transient occurrence of Ls110 on these structures of intra-uterine stages besides the presence of a cysteine-rich N-terminal region (SXC-like domain) suggest that the protein may play a role in the formation of the microfilarial sheath during embryogenesis.

Amino Acid Sequence↗
PubMed2000

Growth, ageing and death of a photoautotrophic plant cell culture.

Batch cultures of photoautotrophic cell suspensions of Chenopodium rubrum L., growing in an inorganic medium on CO2 under a daily balanced light-dark regime of 16: 8 h could be maintained for approximately 100 d without subcultivation. The long-lived cultures showed an initial cell division phase of 4 weeks, followed by a stationary phase of another 4 weeks, after which ageing and progressive cell death reduced the number of living cells and the cultures usually expired after another 3-4 weeks. These developmental phases of the cell culture were characterised with respect to photosynthetic performance, dark respiration, content of phytohormones and capacity of cell division. Cell division of the majority of the cells finished in the G1- or G0-phase of the cell cycle, caused by a pronounced decline in the endogenous levels of auxin and cytokinins. Supply of these growth factors to resting cells resulted in resumption of cytokinesis, at least by some of the cells. However, responsiveness to the phytohormones declined during the stationary phase, and subcultivation was no longer possible beyond day 60 when the phases of ageing and death commenced. Ageing was characterised by a further decline in the photosynthetic capacity of the cells, by a climacteric enhancement of dark respiration, but also by a slight increase in the level of IAA and cytokinins concomitant with a decrease in ethylene. Similarities and differences between the development of batch-cultured photoautotrophic cells of C. rubrum and that of a leaf are discussed with respect to using the cell culture as a model for a leaf.

Cell Death↗
PubMed2000

Improvement of a serodiagnostic strategy for foot-and-mouth disease virus surveillance in cattle under systematic vaccination: a combined system of an indirect ELISA-3ABC with an enzyme-linked immunoelectrotransfer blot assay.

Foot-and-mouth disease (FMD) recombinant non-capsideal viral antigens 3A, 3B, 2C, 3D and 3ABC were assessed individually in an indirect enzyme-linked immunosorbent assay (I-ELISA) for their ability to screen for persistent infection-specific antibodies in cattle, regardless of vaccination condition. Results of sequential serum samples from non-vaccinated animals with experimentally induced persistent infection, and their correlation with virus isolation, indicated that the polypeptides 3A, 3B and 3ABC showed the most adequate characteristics for further field studies. Reliable performance of the I-ELISA with the selected antigen 3ABC was indicated by the distinct patterns observed for the frequency distribution values of naive and true positive samples. For regularly vaccinated livestock, a clear negative profile was proved in samples from regions without recent history of FMD. In contrast, at 90 and 900 days post-outbreak, coexistence of a positive and a negative population was established. These findings indicated that, irrespective of vaccination, the test allowed a classification of the herd-disease status. A high degree of agreement was observed between I-ELISA-3ABC and EITB results for clearly reactive and non-reactive sera. For samples with reactivity values close to that of the cut-off, the EITB profiles upheld the definition of the infection condition. On this basis, screening by I-ELISA-3ABC, together with confirmation of suspect or positive samples by EITB is proposed as an adequate and accurate approach for large-scale epidemiological surveillance.

Animals↗
PubMed2000

Cell recognition by foot-and-mouth disease virus that lacks the RGD integrin-binding motif: flexibility in aphthovirus receptor usage.

Cell surface molecules that can act as virus receptors may exert an important selective pressure on RNA viral quasispecies. Large population passages of foot-and-mouth disease virus (FMDV) in cell culture select for mutant viruses that render dispensable a highly conserved Arg-Gly-Asp (RGD) motif responsible for integrin receptor recognition. Here, we provide evidence that viability of recombinant FMDVs including a Asp-143-->Gly change at the RGD motif was conditioned by a number of capsid substitutions selected upon FMDV evolution in cell culture. Multiply passaged FMDVs acquired the ability to infect human K-562 cells, which do not express integrin alpha(v)beta(3). In contrast to previously described cell culture-adapted FMDVs, the RGD-independent infection did not require binding to the surface glycosaminoglycan heparan sulfate (HS). Viruses which do not bind HS and lack the RGD integrin-binding motif replicate efficiently in BHK-21 cells. Interestingly, FMDV mutants selected from the quasispecies for the inability to bind heparin regained sensitivity to inhibition by a synthetic peptide that represents the G-H loop of VP1. Thus, a single amino acid replacement leading to loss of HS recognition can shift preferential receptor usage of FMDV from HS to integrin. These results indicate at least three different mechanisms for cell recognition by FMDV and suggest a potential for this virus to use multiple, alternative receptors for entry even into the same cell type.

Amino Acid Sequence↗
PubMed1999

Inhibitors of the nonmevalonate pathway of isoprenoid biosynthesis as antimalarial drugs.

A mevalonate-independent pathway of isoprenoid biosynthesis present in Plasmodium falciparum was shown to represent an effective target for chemotherapy of malaria. This pathway includes 1-deoxy-D-xylulose 5-phosphate (DOXP) as a key metabolite. The presence of two genes encoding the enzymes DOXP synthase and DOXP reductoisomerase suggests that isoprenoid biosynthesis in P. falciparum depends on the DOXP pathway. This pathway is probably located in the apicoplast. The recombinant P. falciparum DOXP reductoisomerase was inhibited by fosmidomycin and its derivative, FR-900098. Both drugs suppressed the in vitro growth of multidrug-resistant P. falciparum strains. After therapy with these drugs, mice infected with the rodent malaria parasite P. vinckei were cured.

Aldose-Ketose Isomerases↗
PubMed1999

General practice and the care of children with HIV infection: 6 month prospective interview study.

OBJECTIVES: To describe the use of primary care services by children infected with HIV and to explore the attitudes of their parents to the role of general practitioners in their children's care. DESIGN: A 6 month prospective study. Quantitative analysis of "contact diaries" kept by parents; qualitative analysis of face to face interviews with parents. PARTICIPANTS: Parents of children receiving care at a regional referral centre in London. RESULTS: Twenty four families (80% response rate) were recruited to the study. In 19 families the mother was black African. Half the children had been diagnosed with symptomatic HIV infection, half with AIDS. All the children were registered with a general practitioner who knew of the child's HIV infection. In five families there had initially been tensions in their relationship with their general practitioner but by the time of the study all but one family had established at least an "acceptable" relationship. Children with symptomatic HIV infection saw their general practitioner a mean of 7.5 times per patient year; for children with AIDS the figure was 5.8. Parents regarded the paediatric HIV team at the hospital as their primary source of medical care. Three factors constrained their use of general practice: their own anxieties about distinguishing "normal" symptoms from those related to HIV infection; their view that their general practitioner did not feel competent to treat HIV infected children; and their concerns about maintaining confidentiality in the surgery. CONCLUSIONS: Parents remain oriented towards the paediatric HIV team as their primary source of medical care and use general practice largely for routine prescriptions for their children. Any further development of the general practitioner's role will need to build on existing relationships with specialist providers and take account of parents' concerns.

Child↗
PubMed1999

Regulation of monocyte survival in vitro by deposited IgG: role of macrophage colony-stimulating factor.

IgG deposition at tissue sites characteristically leads to macrophage accumulation and organ injury. Although the mechanism by which deposited IgG induces tissue injury is not known, we have recently demonstrated that deposited IgG stimulates the release of IL-8 and monocyte chemoattractant protein-1 from normal human monocytes, which may drive inflammation. Since IgG also induces macrophage accumulation in these diseases, we hypothesized that deposited IgG protects monocytes from apoptosis. As an in vitro model of the effect of deposited IgG on monocyte survival, monocyte apoptosis was studied after FcgammaR cross-linking. Monocytes cultured on immobilized IgG, which induces FcgammaR cross-linking, were protected from apoptosis, whereas monocytes cultured with equivalent concentrations of F(ab')2 IgG or 50 times higher concentrations of soluble IgG, neither of which induces FcgammaR cross-linking, were not protected. Moreover, this protection was transferable, as supernatants from immobilized IgG-stimulated monocytes protected freshly isolated monocytes from apoptosis and contained functional M-CSF, a known monocyte survival factor. M-CSF mediated the monocyte survival induced by FcgammaR cross-linking, as neutralizing anti-human M-CSF Abs blocked the monocyte protection provided by either immobilized IgG or IgG-stimulated monocyte supernatants. These findings demonstrate a novel mechanism by which deposited IgG targets tissue macrophage accumulation through FcgammaR-mediated M-CSF release. This pathway may play an important role in promoting and potentiating IgG-mediated tissue injury.

Antigen-Antibody Complex↗
PubMed1999

A hookworm allergen which strongly resembles calreticulin.

Immmoglobulin E-rich plasma from patients from Papua New Guinea infected with Necator americanus has been used to probe an adult N. americanus cDNA library for the presence of hookworm allergens. Using this approach, one hookworm allergen has been identified as calreticulin, which was subsequently expressed in Escherichia coli. Little serological cross reactivity was seen between the recombinant calreticulins of this hookworm and its host. Prospective roles for hookworm calreticulin in the host-parasite relationship are discussed in depth.

Allergens↗
PubMed1999

Treatment with the antibiotic roxithromycin in patients with acute non-Q-wave coronary syndromes. The final report of the ROXIS Study.

AIMS: Mounting evidence suggests infection, specifically Chlamydia pneumoniae, plays a role in atherosclerosis. We tested whether antibiotic treatment with the macrolide roxithromycin improves clinical outcome in patients with acute non-Q-wave coronary syndromes. Preliminary reports revealed a reduction in events in the roxithromycin group at 30 days. We now report the long-term follow-up results. METHODS AND RESULTS: Sixty-four per cent of the initial 202 patients with unstable angina who were randomly assigned to receive either roxithromycin or placebo for 30 days completed the active treatment period. At day 30, the primary triple and double end-point rates were 9% and 4% in the placebo group compared to 2% and 0% in the roxithromycin group (unadjusted P = 0.032 and 0.058, respectively). The secondary triple and double end-point rates were again higher in the placebo group at day 90 (12.5% and 6.25% vs 4.37% and 0%, unadjusted P = 0.065 and 0.029, respectively), and at day 180 (14.6% and 7.29% vs 8.69% and 2.17%, unadjusted P = 0.259 and 0.17, respectively). Anti-C, pneumoniae IgG titres were unchanged in both groups while C-reactive protein levels decreased in both strategies, with a more significant decrease in the roxithromycin arm (P = 0.03). Elevated C-reactive protein levels predicted the need for revascularization. CONCLUSIONS: In this pilot trial, roxithromycin appears to extend the clinical benefit of preventing death and re-infarction for at least 6 months after initial treatment.

Aged↗
PubMed1999

Isolation of T-cell antigens by using a recombinant protein library and its application to the identification of novel vaccine candidates against schistosomiasis.

We present here a novel approach to identify T-cell antigens from any infectious agent by use of a library of purified recombinant proteins. Essential features of this strategy include (i) a highly efficient cDNA cloning system which negatively selects against nonrecombinant transformants by making use of the bacterial EcoK restriction system, (ii) affinity staining of cDNA clones expressing recombinant proteins, and (iii) a procedure of simultaneous purification of recombinant proteins from large numbers of isolated clones (representing the protein library) in a single step from pools consisting of up to 24 individual clones. The feasibility of the screening system was confirmed by constructing a protein library of the human parasite Schistosoma mansoni. The recombinant antigens of this library were used to stimulate CD4(+) T cells derived from the axillary lymph nodes of mice vaccinated with irradiated cercariae. In initial screening experiments, we detected parasite-specific proliferation and gamma interferon (IFN-gamma) secretion in response to several pools of cDNA clones. Further analysis of one particular pool revealed that only one of its constituents stimulated considerable IFN-gamma secretion by CD4(+) T cells and that the expressed antigen is identical to a small fragment of myosin heavy chain.

Animals↗
PubMed1999

Interaction of eukaryotic initiation factor eIF4B with the internal ribosome entry site of foot-and-mouth disease virus is independent of the polypyrimidine tract-binding protein.

Eukaryotic translation initiation factor 4B (eIF4B) binds directly to the internal ribosome entry site (IRES) of foot-and-mouth disease virus (FMDV). Mutations in all three subdomains of the IRES stem-loop 4 reduce binding of eIF4B and translation efficiency in parallel, indicating that eIF4B is functionally involved in FMDV translation initiation. In reticulocyte lysate devoid of polypyrimidine tract-binding protein (PTB), eIF4B still bound well to the wild-type IRES, even after removal of the major PTB-binding site. In conclusion, the interaction of eIF4B with the FMDV IRES is essential for IRES function but independent of PTB.

Aphthovirus↗
PubMed1999

Cohesion and organization patterns among family members coping with rheumatoid arthritis.

The effects of rheumatoid arthritis (RA) on family functioning are described from two perspectives, cohesiveness and organization. These concepts are believed to be essential for assessing levels of family stability following the onset of a life crisis. In this study we characterized the ways in which families organize their daily activities and achieve family cohesiveness. Fifteen families participated in the study. The data showed family members have positive perceptions about their family cohesiveness and organizational behaviors, which were characterized in personal interviews. Families also discussed active and passive coping styles in response to RA. Implications for social work practice and issues for future research are presented.

Adaptation, Psychological↗
PubMed1998

Protein heterogeneity of spinach pullulanase results from the coexistence of interconvertible isomeric forms of the monomeric enzyme.

Purified pullulanase (starch-debranching enzyme, R-enzyme, EC 3.2.1. 41) from spinach (Spinacia oleracea L.) chloroplasts separated into at least seven individual enzymically active proteins (isomers, numbered 1-7) on isoelectric focusing or column chromatofocusing. At their isoelectric points (between pH 4.7 and 5.2) these forms were rather stable. At slightly alkaline pH, each converted into the whole set of isomers. PAGE of the purified enzyme under denaturing or non-denaturing conditions resulted in one protein band. When substrate (amylopectin or pullulan) was included in the gel, the native enzyme as well as any of the individual isomers separated into two (sometimes three) bands ('substrate-induced forms', numbered I-III) with different specific activities, dissociation constants of the enzyme-substrate complexes and activation energies. Each substrate-induced form produced the whole set of seven isomers on isoelectric focusing. The specific activity of the total enzyme reflected the relative proportions of the substrate-induced forms. To some extent the relative proportions, as determined by crossed immunoelectrophoresis, could be shifted in favour of the more or the less active forms by reduction with dithiothreitol, and gentle oxidation respectively. Activation by dithiothreitol did not alter the mode of action of the enzyme but only increased the velocity of substrate degradation and extended its activity into the pH range of the chloroplast. As a consequence of isomer interconversion, microheterogeneity could serve to regulate pullulanase activity in a biochemical manner that shares some features with allosteric regulation.

Allosteric Regulation↗