One hundred thirty-eight new Epstein-Barr virus (EBV) genome sequences have been determined. One hundred twenty-five of these and 116 from previous reports were combined to produce a ...multiple-sequence alignment of 241 EBV genomes, which we have used to analyze variation within the viral genome. The type 1/type 2 classification of EBV remains the major form of variation and is defined mostly by EBNA2 and EBNA3, but the type 2 single-nucleotide polymorphisms (SNPs) at the EBNA3 locus extend into the adjacent gp350 and gp42 genes, whose products mediate infection of B cells by EBV. A small insertion within the BART microRNA region of the genome was present in 21 EBV strains. EBV from saliva of U.S. patients with chronic active EBV infection aligned with the wild-type EBV genome with no evidence of WZhet rearrangements. The V3 polymorphism in the Zp promoter for BZLF1 was found to be frequent in nasopharyngeal carcinoma cases from both Hong Kong and Indonesia. Codon usage was found to differ between latent and lytic cycle EBV genes, and the main forms of variation of the EBNA1 protein have been identified.
Epstein-Barr virus causes most cases of infectious mononucleosis and posttransplant lymphoproliferative disease. It contributes to several types of cancer, including Hodgkin's lymphoma, Burkitt's lymphoma, diffuse large B cell lymphoma, nasopharyngeal carcinoma, and gastric carcinoma. EBV genome variation is important because some of the diseases associated with EBV have very different incidences in different populations and geographic regions, and differences in the EBV genome might contribute to these diseases. Some specific EBV genome alterations that appear to be significant in EBV-associated cancers are already known, and current efforts to make an EBV vaccine and antiviral drugs should also take account of sequence differences in the proteins used as targets.
Control of Streptococcus pneumoniae colonisation at human mucosal surfaces is critical to reducing the burden of pneumonia and invasive pneumococcal disease, interrupting transmission, and achieving ...herd protection. Here, we use an experimental human pneumococcal carriage model (EHPC) to show that S. pneumoniae colonisation is associated with epithelial surface adherence, micro-colony formation and invasion, without overt disease. Interactions between different strains and the epithelium shaped the host transcriptomic response in vitro. Using epithelial modules from a human epithelial cell model that recapitulates our in vivo findings, comprising of innate signalling and regulatory pathways, inflammatory mediators, cellular metabolism and stress response genes, we find that inflammation in the EHPC model is most prominent around the time of bacterial clearance. Our results indicate that, rather than being confined to the epithelial surface and the overlying mucus layer, the pneumococcus undergoes micro-invasion of the epithelium that enhances inflammatory and innate immune responses associated with clearance.
Formulation of lipid core nanocapsules Venturini, Cristina G.; Jäger, Eliézer; Oliveira, Catiuscia P. ...
Colloids and surfaces. A, Physicochemical and engineering aspects,
02/2011, Letnik:
375, Številka:
1
Journal Article
Recenzirano
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▶ A study of the quality of nanoparticulated formulations was carried out to obtain lipid-core nanocapsules (LNC). ▶ Those new nanocarriers can present higher loading capacity than ...other due to a dispersion of lipids in their core. ▶ The analytical approach showed a rational design to develop high performance nanocarrier platform. ▶ The LNC are new nanocarriers with potential application in nanomedicine.
Polymeric nanoparticle aqueous suspensions have been proposed as drug carriers to improve the efficacy of medicines. Considering those nanocarriers, nanocapsules are vesicular structures containing an oil core surrounded by a polymeric wall. Recently, we proposed the supramolecular model for a new kind of nanocapsule prepared with triacylglycerol, sorbitan monostearate (SM), polyester and polysorbate 80. Varying the proportions of the raw materials in the organic phase, different kinds of colloids could be obtained. So, our objective was to formulate exclusively lipid-core nanocapsules (LNC) in aqueous suspensions. In this way, the analytical approach to verify the quality of the different formulations was based on light scattering measurements (dynamic light scattering, multiple light scattering and laser diffractometry) and density gradient. The increase in the SM concentration showed a slight tendency of both sedimentation and creaming, while the increase in the oil concentration resulted in creaming. For the latter, size distribution as function of time indicated the presence of nanoemulsion simultaneously with LNC. Finally, density gradient showed an exclusive band for formulations prepared using 1:4.1:2.6 (w/w) of SM, medium chain triacylglycerol and polyester, respectively.
Abstract
Objectives
Cytokines released by infiltrating T cells may promote mechanisms leading to fibrosis in scleroderma. The aim of this study was to investigate the role of the Th2 cytokine IL-31, ...and its receptor IL-31RA, in scleroderma skin and lung fibrosis.
Methods
IL-31 was measured by ELISA of plasma, and by immunochemistry of fibrotic skin and lung tissue of scleroderma patients. The receptor, IL-31RA, was assayed by qPCR of tissue resident cells. Next-generation sequencing was used to profile the responses of normal skin fibroblasts to IL-31. In wild-type Balb/c mice, IL-31 was administered by subcutaneous mini pump, with or without additional TGFβ, and the fibrotic reaction measured by histology and ELISA of plasma.
Results
IL-31 was present at high levels in plasma and fibrotic skin and lung lesions in a subset of scleroderma patients, and the receptor overexpressed by downstream cells relevant to the disease process, including skin and lung fibroblasts, through loss of epigenetic regulation by miR326. In skin fibroblasts, IL-31 induced next generation sequencing profiles associated with cellular growth and proliferation, anaerobic metabolism and mineralization, and negatively associated with angiogenesis and vascular repair, as well as promoting phenotype changes including migration and collagen protein release via pSTAT3, resembling the activation state in the disease. In mice, IL-31 induced skin and lung fibrosis. No synergy was seen with TGFβ, which supressed IL-31RA.
Conclusion
IL-31/IL-31RA is confirmed as a candidate pro-fibrotic pathway, which may contribute to skin and lung fibrosis in a subset of scleroderma patients.
Many regions of the Epstein-Barr virus (EBV) genome, repeated and unique sequences, contribute to the geographical variation observed between strains. Here we use a large alignment of curated EBV ...genome sequences to identify major sites of variation in the genome of type 1 EBV strains; the CAO deletion in latent membrane protein 1 (LMP1) is the most frequent major indel present in the unique regions of EBV strains from various parts of the world. Principal component analysis was used to identify patterns of sequence variation and nucleotide positions in the sequences that can distinguish EBV from some different geographical regions. Viral genome sequence variation also affects interpretation of genetic content; known genes, origins of replication and gene expression control regions explain most of the viral genome but there are still a few sections of unknown function. One of these EBV genome regions contains a large inverted repeat sequence (invR) within the IR-1 major internal repeat array. We deleted this invR sequence and showed that this abolished the ability of the virus to transform human B cells into lymphoblastoid cell lines. This article is part of the theme issue 'Silent cancer agents: multi-disciplinary modelling of human DNA oncoviruses'.
Objective
In systemic sclerosis (SSc), a persistent tissue repair process leads to progressive fibrosis of the skin and internal organs. The role of mesenchymal stem cells (MSCs), which ...characteristically initiate and regulate tissue repair, has not been fully evaluated. We undertook this study to investigate whether dividing metakaryotic MSCs are present in SSc skin and to examine whether exposure to the disease microenvironment activates MSCs and leads to transdifferentiation.
Methods
Skin biopsy material from patients with recent‐onset diffuse SSc was examined by collagenase spread of 1‐mm–thick surface‐parallel sections, in order to identify dividing metakaryotic stem cells in each tissue plane. Adipose‐derived MSCs from healthy controls were treated with dermal blister fluid (BF) from patients with diffuse SSc and profiled by next‐generation sequencing, or they were evaluated for phenotypic changes relevant to SSc. Differential responses of dermal fibroblasts were studied in parallel.
Results
MSC‐like cells undergoing active metakaryotic division were identified in SSc sections (but not control sections) most prominently in the deep dermis and adjacent to damaged microvessels, in both clinically involved and uninvolved skin. Furthermore, exposure to SSc BF caused selective MSC activation, inducing a myofibroblast signature, while reducing signatures of vascular repair and adipogenesis and enhancing migration and contractility. Microenvironmental factors implicated in inducing transdifferentiation included the profibrotic transforming growth factor β, the presence of lactate, and mechanosensing, while the microenvironment Th2 cytokine, interleukin‐31, enhanced osteogenic commitment (calcinosis).
Conclusion
Dividing MSC‐like cells are present in the SSc disease microenvironment where multiple factors, likely acting in concert, promote transdifferentiation and lead to a complex and resistant disease state.
In this study, two types of cutaneous-directed nanoparticles are proposed for the co-encapsulation of imiquimod (a drug approved for the treatment of basal cell carcinoma) and copaiba oil (oil that ...exhibits anti-proliferative properties). Nanostructured copaiba capsules (NCCImq) were prepared using the interfacial deposition method, and nanostructured Brazilian lipids (NBLImq) were prepared by high-pressure homogenization. The formulations exhibited average diameter, zeta potential, pH and drug content of approximately 200nm, −12mV, 6 and 1mgmL−1, respectively. In addition, the formulations exhibited homogeneity regarding particle size, high encapsulation efficiency and stability. Both nanocarriers controlled imiquimod release, and NBLImq exhibited slower drug release (p < 0.05), likely due to increased interaction of the drug with the solid lipid (cupuaçu seed butter). The in vitro evaluation of the imiquimod-loaded nanocarriers was performed using healthy skin cells (keratinocytes, HaCaT); no alteration was observed, suggesting the biocompatibility of the nanocarriers. In addition, in vitro skin permeation/penetration using pig skin was performed, and NCCImq led to increased drug retention in the skin layers and reduced amounts of drug found in the receiver solution. Thus, NCCImq is considered the most promising nanoformulation for the treatment of skin carcinoma.
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Excessive neutrophil extravasation can drive immunopathology, exemplified in pyogenic meningitis caused by
infection. Insufficient knowledge of the mechanisms that amplify neutrophil extravasation ...has limited innovation in therapeutic targeting of neutrophil mediated pathology. Attention has focussed on neutrophil interactions with endothelia, but data from mouse models also point to a role for the underlying pericyte layer, as well as perivascular macrophages, the only other cell type found within the perivascular space in the cerebral microvasculature. We tested the hypothesis that human brain vascular pericytes (HBVP) contribute to neutrophil extravasation in a transwell model of the cerebral post-capillary venule. We show that pericytes augment endothelial barrier formation. In response to inflammatory cues, they significantly enhance neutrophil transmigration across the endothelial barrier, without increasing the permeability to small molecules. In our model, neither pericytes nor endothelia responded directly to bacterial stimulation. Instead, we show that paracrine signalling by multiple cytokines from monocyte derived macrophages drives transcriptional upregulation of multiple neutrophil chemokines by pericytes. Pericyte mediated amplification of neutrophil transmigration was independent of transcriptional responses by endothelia, but could be mediated by direct chemokine translocation across the endothelial barrier. Our data support a model in which microbial sensing by perivascular macrophages generates an inflammatory cascade where pericytes serve to amplify production of neutrophil chemokines that are translocated across the endothelial barrier to act directly on circulating neutrophils. In view of the striking redundancy in inflammatory cytokines that stimulate pericytes and in the neutrophil chemokines they produce, we propose that the mechanism of chemokine translocation may offer the most effective therapeutic target to reduce neutrophil mediated pathology in pyogenic meningitis.
Summary
Chronic active Epstein–Barr virus (CAEBV) disease is a rare condition characterised by persistent EBV infection in previously healthy individuals. Defective EBV genomes were found in East ...Asian patients with CAEBV. In the present study, we sequenced 14 blood EBV samples from three UK patients with CAEBV, comparing the results with saliva CAEBV samples and other conditions. We observed EBV deletions in blood, some of which may disrupt viral replication, but not saliva in CAEBV. Deletions were lost overtime after successful treatment. These findings are compatible with CAEBV being associated with the evolution and persistence of EBV+ haematological clones that are lost on successful treatment.
Aqueous solutions of lipid-core nanocapsules are interesting drug delivery systems for passive drug targeting. In this study, we hypothesized that the drug distribution mechanisms in lipid-core ...nanocapsule formulations could be categorized into six different types. To experimentally determine the type of drug distribution in these formulations, we proposed the use of an algorithm as an innovative strategy. The approach is shown to be a valuable tool to optimize and select formulations intended for drug delivery. The best physico-chemical parameter in terms of predicting the type of distribution was the log
D
value. In conclusion, the use of the algorithm developed in this study represents a simple and rapid approach through which it was possible to experimentally determine the drug distribution in colloidal formulations for eight drug models.
The algorithm described in this work is a simple and rapid approach to experimentally determine the drug distribution in colloidal formulations.
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