Drug Interactions with Glutaredoxin Orthologues

Glutaredoxin, an enzymatic protein, is an important component of cell viability and function. It catalyzes reactions involved in DNA synthesis and innate immunity [1,4]. Glutaredoxin is also essential in antibiotic resistance in pathogenic bacterial species. Pseudomonas aeruginosa in particular is responsible for infecting the lung tissue of its human hosts, resulting in the development of pneumonia and cystic fibrosis [3]. Because glutaredoxin is pertinent in cell proliferation of eukaryotic and bacterial cells alike, medicinal fragments that take advantage of the subtle differences in protein structure of the orthologous proteins can be synthesized and enhanced to bind bacterial glutaredoxins, without inhibiting the function of the human form. This can be accomplished by exploiting the mechanisms of fragment based drug discovery using NMR techniques. A library of small potential medicinal fragments are screened against each protein to determine which interact, or bind most efficiently to the bacterial orthologues with little to no interaction with eukaryotic cells. To confirm the ability of select fragments hits to kill bacterial cells without harming human cells, MTT and MIC assays are performed to determine what concentration of fragment is needed to obtain desired therapeutic results [6,7]. These assays were performed against selected lead fragments, particularly RK207 and RK395, which can be structurally enhanced to bind even more to the bacterial orthologues. This research can potentially lead to the development of drug targets against bacterial orthologues of glutaredoxin to treat life threatening diseases caused by pathogenic species

Estimation of hypocentral parameters of local earthquakes when crustal layers have constant P-velocities and dipping interfaces

The paper describes an algorithm for estimating the hypocentral coordinates and origin time of local earthquakes when the wave speed model to be employed is a layered one with dipping interfaces. A constrained least-squared error problem has been solved using the penalty function approach, in conjunction with the sequential unconstrained optimization technique of Fiacco and McCormick. Joint confidence intervals for the computed parameters are estimated using the approach of Bard for nonlinear problems. These results show that when a hypocentre lies outside the array of recording stations and head waves from a dipping interface are involved, then its inclination must be taken into account for dip angles exceeding 5°

Low Numbers of FOXP3 Positive Regulatory T Cells Are Present in all Developmental Stages of Human Atherosclerotic Lesions

BACKGROUND: T cell mediated inflammation contributes to atherogenesis and the onset of acute cardiovascular disease. Effector T cell functions are under a tight control of a specialized T cell subset, regulatory T cells (Treg). At present, nothing is known about the in situ presence of Treg in human atherosclerotic tissue. In the present study we investigated the frequency of naturally occurring Treg cells in all developmental stages of human atherosclerotic lesions including complicated thrombosed plaques. METHODOLOGY: Normal arteries, early lesions (American Heart Association classification types I, II, and III), fibrosclerotic plaques (types Vb and Vc) and 'high risk' plaques (types IV, Va and VI) were obtained at surgery and autopsy. Serial sections were immunostained for markers specific for regulatory T cells (FOXP3 and GITR) and the frequency of these cells was expressed as a percentage of the total numbers of CD3+ T cells. Results were compared with Treg counts in biopsies of normal and inflammatory skin lesions (psoriasis, spongiotic dermatitis and lichen planus). PRINCIPLE FINDINGS: In normal vessel fragments T cells were virtually absent. Treg were present in the intima during all stages of plaque development (0.5-5%). Also in the adventitia of atherosclerotic vessels Treg were encountered, in similar low amounts. High risk lesions contained significantly increased numbers of Treg compared to early lesions (mean: 3.9 and 1.2%, respectively). The frequency of FOXP3+ cells in high risk lesions was also higher compared to stable lesions (1.7%), but this difference was not significant. The mean numbers of intimal FOXP3 positive cells in atherosclerotic lesions (2.4%) was much lower than those in normal (24.3%) or inflammatory skin lesions (28%). CONCLUSION: Low frequencies of Treg in all developmental stages of human plaque formation could explain the smoldering chronic inflammatory process that takes place throughout the longstanding course of atherosclerosis

Negative Feedback Regulation of T Cells via Interleukin-2 and FOXP3 Reciprocity

As interleukin-2 (IL2) is central to the clonal expansion of antigen-selected T cells, we investigated the relationship between IL2 and the negative regulatory transcription factor FOXP3. We found IL2 to be responsible for T cell antigen receptor (TCR)-activated FOXP3 expression by both CD4+ and CD8+ human T cells, and as anticipated, FOXP3 expression restricted TCR-stimulated IL2 expression. However, no evidence could be found that FOXP3+ cells actively suppress IL2 expression by FOXP3- cells. These data are consistent with an IL2/FOXP3-dependent negative feedback loop that normally regulates the T cell immune response. It follows that a defect in this negative feedback loop as a result of a deficiency of either IL2 or FOXP3 will lead to a hyperproliferative autoimmune syndrome, without the necessity of invoking an active suppressive function for FOXP3+ T cells

CD4+ CD25+ FoxP3+ regulatory T cells suppress cytotoxicity of CD8+ effector T cells: implications for their capacity to limit inflammatory central nervous system damage at the parenchymal level

<p>Abstract</p> <p>Background</p> <p>CD4⁺CD25⁺forkhead box P3 (FoxP3)⁺regulatory T cells (T reg cells) are known to suppress adaptive immune responses, key control tolerance and autoimmunity.</p> <p>Methods</p> <p>We challenged the role of CD4⁺T reg cells in suppressing established CD8⁺T effector cell responses by using the OT-I/II system <it>in vitro </it>and an OT-I-mediated, oligodendrocyte directed <it>ex vivo </it>model (ODC-OVA model).</p> <p>Results</p> <p>CD4⁺T reg cells dampened cytotoxicity of an ongoing CD8⁺T effector cell attack <it>in vitro </it>and within intact central nervous system tissue <it>ex vivo</it>. However, their suppressive effect was limited by the strength of the antigen signal delivered to the CD8⁺T effector cells and the ratio of regulatory to effector T cells. CD8⁺T effector cell suppression required T cell receptor-mediated activation together with costimulation of CD4⁺T reg cells, but following activation, suppression did not require restimulation and was antigen non-specific.</p> <p>Conclusions</p> <p>Our results suggest that CD4⁺T reg cells are capable of suppressing CD8⁺T effector cell responses at the parenchymal site, that is, limiting parenchymal damage in autoimmune central nervous system inflammation.</p

Identification, frequency, activation and function of CD4+ CD25highFoxP3+ regulatory T cells in children with juvenile idiopathic arthritis

The aim of the study was to test the frequency of CD4+ CD25highFoxP3 regulatory T cells in JIA patients and to assess their activation status and functional activity. The study involved 12 children with JIA and 35 healthy control subjects. PBMC were stained with monoclonal antibodies (anti-CD25, anti-CD4, anti-CD127, anti-CD69, anti-CD71, and anti-FoxP3). The samples were evaluated using flow cytometer. CD4+ CD25− and CD4+ CD25+ cells were isolated by negative and positive selection with magnetic microbeads. CD4+ CD25+ and CD4+ CD25− cells were cultured separately and co-cultured (1:1) with or without PHA. The percentage of Tregs in JIA patients was significantly decreased in comparison with controls (median, 3.2 vs. 4.6; P = 0.042). Relative fluorescence intensities of FoxP3 were higher in JIA patients than in controls (median, 9.1 vs. 6.8). The percentage of activated Tregs (CD71+) was significantly higher in JIA patients in comparison with controls (median, 6.5 vs. 2.8; P = 0.00043). CD4+ CD25+ cells derived from JIA patients and controls were anergic upon PHA stimulation, while CD4+ CD25− cells showed intensive proliferative response. The proliferation rate of CD4+ CD25− cells stimulated by PHA was decreased in co-cultures. In JIA patients, the inhibition of proliferation of CD4+ CD25− cells by CD4+ CD25+ cells was 37.9%, whereas in controls it was significantly lower (55.7%, P = 0.046). JIA patients had statistically lower percentage of Tregs in peripheral blood compared to controls. CD4+ CD25+ cells sorted from peripheral blood of JIA patients had statistically lower ability to suppress CD4+ CD25− cell proliferation in comparison with cells obtained from controls

Novel magnetite nanoparticles coated with waste sourced bio- based substances as sustainable and renewable adsorbing materials

This study examines the possibility of using bio-based product isolated from urban solid wastes as a material for environmental technological applications. To this end, Fe3O4 nanoparticles coated with different amounts of soluble bio-based products (SBO) were synthesized as low-cost nanoadsorbent for the removal of pollutants in wastewater. Particles of 10 nm diameter with Fe3O4 core and SBO shell were obtained. The concentration of SBO employed in the synthesis had no effect on the size and structure of the NPs, but ruled the pHPZC and aggregation of the nanoparticles in water. The cationic dye crystal violet (CV) was used as a model pollutant to test the adsorption capacity of the nanoparticles. The results indicated that both the medium pH and NP dosage were significant parameters to enhance the removal of CV. The results contribute to the studies which show how wastes can become a source of revenue through the industrial exploitation of their chemical value.Fil: Magnacca, Giuliana. Università di Torino; ItaliaFil: Allera, Alex. Università di Torino; ItaliaFil: Montoneri, Enzo. Università di Torino; ItaliaFil: Celi, Luisella. Università di Torino; ItaliaFil: Benito, Damián Ezequiel. Universidad Nacional de La Plata. Laboratorio de Investigación y Desarrollo de Métodos Analíticos; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata; ArgentinaFil: Gagliardi, Leonardo Gabriel. Universidad Nacional de La Plata. Laboratorio de Investigación y Desarrollo de Métodos Analíticos; ArgentinaFil: Martire, Daniel Osvaldo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico la Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; ArgentinaFil: Gonzalez, Monica Cristina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico la Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; ArgentinaFil: Carlos, Luciano. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico la Plata. Instituto de Investigaciones Fisicoquímicas Teóricas y Aplicadas; Argentin

Characterization of regulatory T cells in urban newborns

© 2009 Ly et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution Licens

Methylation matters: binding of Ets-1 to the demethylated Foxp3 gene contributes to the stabilization of Foxp3 expression in regulatory T cells

The forkhead-box protein P3 (Foxp3) is a key transcription factor for the development and suppressive activity of regulatory T cells (Tregs), a T cell subset critically involved in the maintenance of self-tolerance and prevention of over-shooting immune responses. However, the transcriptional regulation of Foxp3 expression remains incompletely understood. We have previously shown that epigenetic modifications in the CpG-rich Treg-specific demethylated region (TSDR) in the Foxp3 locus are associated with stable Foxp3 expression. We now demonstrate that the methylation state of the CpG motifs within the TSDR controls its transcriptional activity rather than a Treg-specific transcription factor network. By systematically mutating every CpG motif within the TSDR, we could identify four CpG motifs, which are critically determining the transcriptional activity of the TSDR and which serve as binding sites for essential transcription factors, such as CREB/ATF and NF-κB, which have previously been shown to bind to this element. The transcription factor Ets-1 was here identified as an additional molecular player that specifically binds to the TSDR in a demethylation-dependent manner in vitro. Disruption of the Ets-1 binding sites within the TSDR drastically reduced its transcriptional enhancer activity. In addition, we found Ets-1 bound to the demethylated TSDR in ex vivo isolated Tregs, but not to the methylated TSDR in conventional CD4+ T cells. We therefore propose that Ets-1 is part of a larger protein complex, which binds to the TSDR only in its demethylated state, thereby restricting stable Foxp3 expression to the Treg lineage

Two Lysines in the Forkhead Domain of Foxp3 Are Key to T Regulatory Cell Function

Background: The forkhead box transcription factor, Foxp3, is master regulator of the development and function of CD4+CD25+ T regulatory (Treg) cells that limit autoimmunity and maintain immune homeostasis. The carboxyl-terminal forkhead (FKH) domain is required for the nuclear localization and DNA binding of Foxp3. We assessed how individual FKH lysines contribute to the functions of Foxp3 in Treg cells. Methodology/Principal Findings: We found that mutation of FKH lysines at position 382 (K17) and at position 393 (K18) impaired Foxp3 DNA binding and inhibited Treg suppressive function in vivo and in vitro. These lysine mutations did not affect the level of expression of Foxp3 but inhibited IL-2 promoter remodeling and had important and differing effects on Treg-associated gene expression. Conclusions/Significance: These data point to complex effects of post-translational modifications at individual lysines within the Foxp3 FKH domain that affect Treg function. Modulation of these events using small molecule inhibitors ma