Histopathological and ultrastructural aspects of mice lungs experimentally infected with dengue virus serotype 2

Histological and ultrastructural alterations in lung tissue of BALB/c mice infected with dengue virus serotype 2 (non-neuroadapted), by intraperitoneal and intravenous routes were analyzed. Lung tissues were processed following the standard techniques for photonic and electron transmission microscopies. Histopathological and ultrastructural studies showed interstitial pneumonia, characterized by the presence of mononuclear cells. In the mouse model, the dengue virus serotype 2 seems to led to a transient inflammatory process without extensive damage to the interalveolar septa, but caused focal alterations of the blood-exchange barrier. Endothelial cells of blood capillaries exhibited phyllopodia suggesting activation by presence of dengue virus. Morphometrical analysis of mast cells showed an expressive increase of the number of these cells in peribronchiolar spaces and adjacent areas to the interalveolar septa. Alveolar macrophages showed particles dengue virus-like inside rough endoplasmic reticulum and Golgi complex, suggesting viral replication. The tissue alterations observed in our experimental model were similar to the observed in human cases of dengue fever and dengue hemorrhagic fever. Our results show that BALB/c mice are permissive host for dengue virus serotype 2 replication and therefore provides an useful model to study of morphological aspects of dengue virus infection.

Analysis of synaptic-like microvesicle exocytosis of B-cells using a live imaging technique.

Pancreatic β-cells play central roles in blood glucose homeostasis. Beside insulin, these cells release neurotransmitters and other signaling molecules stored in synaptic-like microvesicles (SLMVs). We monitored SLMV exocytosis by transfecting a synaptophysin-pHluorin construct and by visualizing the cells by Total Internal Reflection Fluorescence (TIRF) microscopy. SLMV fusion was elicited by 20 mM glucose and by depolarizing K(+) concentrations with kinetics comparable to insulin secretion. SLMV exocytosis was prevented by Tetanus and Botulinum-C neurotoxins indicating that the fusion machinery of these organelles includes VAMP-2/-3 and Syntaxin-1, respectively. Sequential visualization of SLMVs by TIRF and epifluorescence microscopy showed that after fusion the vesicle components are rapidly internalized and the organelles re-acidified. Analysis of single fusion episodes revealed the existence of two categories of events. While under basal conditions transient fusion events prevailed, long-lasting episodes were more frequent upon secretagogue exposure. Our observations unveiled similarities between the mechanism of exocytosis of insulin granules and SLMVs. Thus, diabetic conditions characterized by defective insulin secretion are most probably associated also with inappropriate release of molecules stored in SLMVs. The assessment of the contribution of SLMV exocytosis to the manifestation of the disease will be facilitated by the use of the imaging approach described in this study.

Reconstituted human polyclonal plasma-derived secretory-like IgM and IgA maintain the barrier function of epithelial cells infected with an enteropathogen.

Intravenous administration of polyclonal and monoclonal antibodies has proven to be a clinically valid approach in the treatment, or at least relief, of many acute and chronic pathologies, such as infection, immunodeficiency, and a broad range of autoimmune conditions. Plasma-derived IgG or recombinant IgG are most frequently used for intravenous or subcutaneous administration, whereas a few IgM-based products are available as well. We have established recently that secretory-like IgA and IgM can be produced upon association of plasma-derived polymeric IgA and IgM with a recombinant secretory component. As a next step toward potential future mucosal administration, we sought to unravel the mechanisms by which these secretory Igs protect epithelial cells located at the interface between the environment and the inside of the body. By using polarized epithelial Caco-2 cell monolayers and Shigella flexneri as a model enteropathogen, we found that polyspecific plasma-derived SIgA and SIgM fulfill many protective functions, including dose-dependent recognition of the antigen via formation of aggregated immune complexes, reduction of bacterial infectivity, maintenance of epithelial cell integrity, and inhibition of proinflammatory cytokine/chemokine production by epithelial cells. In this in vitro model devoid of other cellular or molecular interfering partners, IgM and secretory IgM showed stronger bacterial neutralization than secretory IgA. Together, these data suggest that mucosally delivered antibody preparations may be most effective when combining both secretory-like IgA and IgM, which, together, play a crucial role in preserving several levels of epithelial cell integrity.

What should a virtual University Library environment look like?

Over the past year, the Open University of Catalonia library has been designing its new website with this question in mind. Our main concern has been how to integrate the library in the student day to day study routine to not to be only a satellite tool. We present the design of the website that, in a virtual library like ours, it is not only a website but the whole library itself. The central point of the web is my library, a space that associates the library resources with the student's curriculum and their course subjects. There the students can save the resources as favourites, comment or share them. They have also access to all the services the library offers them. The resources are imported from multiple tools such as Millennium, SFX, Metalib and Dspace to the Drupal CMS. Then the resources' metadata can be enriched with other contextual information from other sources, for example the course subjects. And finally they can be exported in standard, open data formats making them available for linked data applications.

Karyotypic variation and geographic distribution of Anopheles campestris-like (Diptera: Culicidae) in Thailand

Seventy-one isolines of Anopheles campestris-like were established from wild-caught females collected from human-biting and animal-biting traps at 12 locations in Thailand. All isolines had an average branch summation of seta 2-VI pupal skins ranging from 20.3-30.0 branches, which is in the range of An. campestris (17-58 branches). They showed three different karyotypes based on the amount of extra heterochromatin in the sex chromosomes, namely Forms B (X2, Y2), E (X1, X2, X3, Y5) and a new karyotypic Form F (X2, X3, Y6). Form B has been found only in Chaing Mai and Kamphaeng Phet populations, while Forms E and F are widely distributed throughout the species range. Genetic crosses between the 12 isolines, which were arbitrarily selected as representatives of An. campestris-like Forms B, E and F, revealed genetic compatibility that provided viable progeny through F2 generations, suggesting a conspecific nature of these karyotypic forms. These results are supported by the very low intraspecies variation (genetic distance < 0.005) of ITS2, COI and COII from genomic DNA of the three karyotypic forms.

The Thioredoxin TRX-1 Modulates the Function of the Insulin-Like Neuropeptide DAF-28 during Dauer Formation in Caenorhabditis elegans

Thioredoxins comprise a conserved family of redox regulators involved in many biological processes, including stress resistance and aging. We report that the C. elegans thioredoxin TRX-1 acts in ASJ head sensory neurons as a novel modulator of the insulin-like neuropeptide DAF-28 during dauer formation. We show that increased formation of stress-resistant, long-lived dauer larvae in mutants for the gene encoding the insulin-like neuropeptide DAF-28 requires TRX-1 acting in ASJ neurons, upstream of the insulin-like receptor DAF-2. Genetic rescue experiments demonstrate that redox-independent functions of TRX-1 specifically in ASJ neurons are needed for the dauer formation constitutive (Daf-c) phenotype of daf-28 mutants. GFP reporters of trx-1 and daf-28 show opposing expression patterns in dauers (i.e. trx-1 is up-regulated and daf-28 is down-regulated), an effect that is not observed in growing L2/L3 larvae. In addition, functional TRX-1 is required for the down-regulation of a GFP reporter of daf-28 during dauer formation, a process that is likely subject to DAF-28-mediated feedback regulation. Our findings demonstrate that TRX-1 modulates DAF-28 signaling by contributing to the down-regulation of daf-28 expression during dauer formation. We propose that TRX-1 acts as a fluctuating neuronal signaling modulator within ASJ neurons to monitor the adjustment of neuropeptide expression, including insulin-like proteins, during dauer formation in response to adverse environmental conditions.

Candidate gene analysis of ocular toxoplasmosis in Brazil: evidence for a role for toll-like receptor 9 (TLR9)

Toxoplasma gondii infection is an important mediator of ocular disease in Brazil more frequently than reported from elsewhere. Infection and pathology are characterized by a strong proinflammatory response which in mice is triggered by interaction of the parasite with the toll-like receptor (TLR)/MyD88 pathway. A powerful way to identify the role of TLRs in humans is to determine whether polymorphisms at these loci influence susceptibility to T. gondii-mediated pathologies. Here we report on a small family-based study (60 families; 68 affected offspring) undertaken in Brazil which was powered for large effect sizes using single nucleotide polymorphisms with minor alleles frequencies > 0.3. Of markers in TLR2, TLR5 and TLR9 that met these criteria, we found an association Family Based Association Tests [(FBAT) Z score = 4.232; p = 1.5 x 10-5; p corrected = 1.2 x 10-4] between the C allele (frequency = 0.424; odds ratio = 7; 95% confidence interval 1.6-30.8) of rs352140 at TLR9 and toxoplasmic retinochoroiditis in Brazil. This supports the hypothesis that direct interaction between T. gondii and TLR9 may trigger proinflammatory responses that lead to severe pathologies such as the ocular disease that is associated with this infection in Brazil.

An isoform of microtubule-associated protein 2 (MAP2) containing four repeats of the tubulin-binding motif.

Microtubule-associated protein 2 (MAP2) exists in both high- and low-molecular mass isoforms, each of which has a tubulin-binding domain consisting of 3 imperfect tandem repeats of 31 amino acids containing a more highly conserved 18 amino acid 'core' sequence. We describe here a novel form of low molecular mass MAP2 (MAP2c) that contains an additional 4th repeat of this tubulin-binding motif. Like the 3 previously known repeat sequences, this 4th copy is highly conserved between MAP2 and the two other known members of the same gene family, tau and MAP4. In each of these three genes the additional 4th repeat is inserted between the 1st and 2nd repeats of the 3-repeat form of the molecule. Experiments with brain cell cultures, in which the relative proportions of neurons and glia had been manipulated by drug treatment, showed that 4-repeat MAP2c is associated with glial cells whereas 3-repeat MAP2c is expressed in neurons. Whereas 3-repeat MAP2c is expressed early in development and then declines, the level of 4-repeat MAP2c increases later in development, corresponding to the relatively late differentiation of glial cells compared to neurons. When transfected into non-neuronal cells, the 4-repeat version of MAP2c behaved indistinguishably from the 3-repeat form in stabilising and rearranging cellular microtubules. The presence of an additional 4th repeat of the tubulin-binding motif in all three members of the MAP2 gene family suggests that this variant arose prior to their differentiation from an ancestral gene.

Polymorphisms IL10-819 and TLR-2 are potentially associated with sepsis in Brazilian patients

Genetic variation in immune response is probably involved in the progression of sepsis and mortality in septic patients. However, findings in the literature are sometimes conflicting or their significance is uncertain. Thus, we investigated the possible association between 12 polymorphisms located in the interleukin-6 (IL6), IL10, TLR-2, Toll-like receptor-4 (TLR-4), tumor necrosis factor-α and tumor necrosis factor-β (lymphotoxin α - LTA) genes and sepsis. Critically ill patients classified with sepsis, severe sepsis and septic shock and 207 healthy volunteers were analyzed and genotyped. Seven of the nine polymorphisms showed similar distributions in allele frequencies between patients and controls. Interestingly, our data suggest that the IL10-819 and TLR-2 polymorphisms may be potential predictors of sepsis.

Differential prefrontal-like deficit in children after cerebellar astrocytoma and medulloblastoma tumor

BACKGROUND This study was realized thanks to the collaboration of children and adolescents who had been resected from cerebellar tumors. The medulloblastoma group (CE+, n = 7) in addition to surgery received radiation and chemotherapy. The astrocytoma group (CE, n = 13) did not receive additional treatments. Each clinical group was compared in their executive functioning with a paired control group (n = 12). The performances of the clinical groups with respect to controls were compared considering the tumor's localization (vermis or hemisphere) and the affectation (or not) of the dentate nucleus. Executive variables were correlated with the age at surgery, the time between surgery-evaluation and the resected volume. METHODS The executive functioning was assessed by means of WCST, Complex Rey Figure, Controlled Oral Word Association Test (letter and animal categories), Digits span (WISC-R verbal scale) and Stroop test. These tests are very sensitive to dorsolateral PFC and/or to medial frontal cortex functions. The scores for the non-verbal Raven IQ were also obtained. Direct scores were corrected by age and transformed in standard scores using normative data. The neuropsychological evaluation was made at 3.25 (SD = 2.74) years from surgery in CE group and at 6.47 (SD = 2.77) in CE+ group. RESULTS The Medulloblastoma group showed severe executive deficit (

Pulmonary Sarcoid-like Granulomatosis after Multiple Vaccinations of a Long-term Surviving Patient with Metastatic Melanoma.

Autoimmune side effects are frequent in patients with cancer treated with immune checkpoint-targeting antibodies, but are rare with cancer vaccines. Here, we present a case report on a patient with metastatic melanoma who developed pulmonary sarcoid-like granulomatosis following repetitive vaccinations with peptides and CpG. Despite multiple metastases, including one lesion in the brain, the patient is alive and well more than 13 years after the diagnosis of metastatic disease. The strongly activated tumor-specific CD8(+) T cells showed robust long-term memory and effector functions. It is possible that long-term survival and adverse autoimmune events may become more common for vaccines inducing robust anticancer immune responses as were present in this patient. Cancer Immunol Res; 2(12); 1148-53. ©2014 AACR.

Altered expression of beta-galactosidase-1-like protein 3 (Glb1l3) in the retinal pigment epithelium (RPE)-specific 65-kDa protein knockout mouse model of Leber's congenital amaurosis

Purpose: In this study, we investigated the expression of the gene encoding beta-galactosidase (Glb)-1-like protein 3 (Glb1l3), a member of the glycosyl hydrolase 35 family, during retinal degeneration in the retinal pigment epithelium (RPE)-specific 65-kDa protein knockout (Rpe65(-/-)) mouse model of Leber congenital amaurosis (LCA). Additionally, we assessed the expression of the other members of this protein family, including beta-galactosidase-1 (Glb1), beta-galactosidase-1-like (Glb1l), and beta-galactosidase-1-like protein 2 (Glb1l2).Methods: The structural features of Glb1l3 were assessed using bioinformatic tools. mRNA expression of Glb-related genes was investigated by oligonucleotide microarray, real-time PCR, and reverse transcription (RT) -PCR. The localized expression of Glb1l3 was assessed by combined in situ hybridization and immunohistochemistry.Results: Glb1l3 was the only Glb-related member strongly downregulated in Rpe65(-/-) retinas before the onset and during progression of the disease. Glb1l3 mRNA was only expressed in the retinal layers and the RPE/choroid. The other Glb-related genes were ubiquitously expressed in different ocular tissues, including the cornea and lens. In the healthy retina, expression of Glb1l3 was strongly induced during postnatal retinal development; age-related increased expression persisted during adulthood and aging.Conclusions: These data highlight early-onset downregulation of Glb1l3 in Rpe65-related disease. They further indicate that impaired expression of Glb1l3 is mostly due to the absence of the chromophore 11-cis retinal, suggesting that Rpe65 deficiency may have many metabolic consequences in the underlying neuroretina.

Tumor necrosis-like weak inducer of apoptosis as a proinflammatory cytokine in human adipocyte cells: up-regulation in severe obesity is mediated by inflammation but not hypoxia.

CONTEXT Adipose tissue hypoxia and endoplasmic reticulum (ER) stress may link the presence of chronic inflammation and macrophage infiltration in severely obese subjects. We previously reported the up-regulation of TNF-like weak inducer of apoptosis (TWEAK)/fibroblast growth factor-inducible 14 (Fn14) axis in adipose tissue of severely obese type 2 diabetic subjects. OBJECTIVES The objective of the study was to examine TWEAK and Fn14 adipose tissue expression in obesity, severe obesity, and type 2 diabetes in relation to hypoxia and ER stress. DESIGN In the obesity study, 19 lean, 28 overweight, and 15 obese nondiabetic subjects were studied. In the severe obesity study, 23 severely obese and 35 control subjects were studied. In the type 2 diabetes study, 11 type 2 diabetic and 36 control subjects were studied. The expression levels of the following genes were analyzed in paired samples of sc and visceral adipose tissue: Fn14, TWEAK, VISFATIN, HYOU1, FIAF, HIF-1a, VEGF, GLUT-1, GRP78, and XBP-1. The effect of hypoxia, inflammation, and ER stress on the expression of TWEAK and Fn14 was examined in human adipocyte and macrophage cell lines. RESULTS Up-regulation of TWEAK/Fn14 and hypoxia and ER stress surrogate gene expression was observed in sc and visceral adipose tissue only in our severely obese cohort. Hypoxia modulates TWEAK or Fn14 expression in neither adipocytes nor macrophages. On the contrary, inflammation up-regulated TWEAK in macrophages and Fn14 expression in adipocytes. Moreover, TWEAK had a proinflammatory effect in adipocytes mediated by the nuclear factor-kappaB and ERK but not JNK signaling pathways. CONCLUSIONS Our data suggest that TWEAK acts as a pro-inflammatory cytokine in the adipose tissue and that inflammation, but not hypoxia, may be behind its up-regulation in severe obesity.

Comparison of the efficacy of long-lasting insecticidal nets PermaNet® 2.0 and Olyset® against Anopheles albimanus under laboratory conditions

Insecticide-treated nets provide a reduction in human-vector contact through physical barrier, mortality and/or repellent effects that protect both users and non-users, thereby protecting the wider community from vector-borne diseases like malaria. Long-lasting insecticide-treated nets (LLINs) are the best alternative. This study evaluated the bioefficacy of LLINs PermaNet® 2.0 and Olyset® under laboratory conditions with Anopheles albimanus. The laboratory strain was evaluated for insecticide susceptibility with selected insecticides used for malarial control. Regeneration time and wash resistance were evaluated with the standard bioassay cone technique following WHO guidelines. Heat assistance was used for Olyset® nets; the nets were exposed to four different temperatures to speed the regeneration process. The regeneration study of PermaNet® 2.0 showed that efficacy was fully recovered by 24 h after one and three washes and wash resistance persisted for 15 washes. Regeneration of Olyset® nets was not observed for nets washed three times, even with the different temperature exposures for up to seven days. Thus, for Olyset® the wash resistance evaluation could not proceed. Differences in response between the two LLINs may be associated with differences in manufacturing procedures and species response to the evaluated LLINs. PermaNet® 2.0 showed higher and continuous efficacy against An. albimanus.