Orthologous gene-expression profiling in multi-species models: search for candidate genes

Microarray-driven gene-expression profiles are generally produced and analyzed for a single specific experimental model. We have assessed an analytical approach that simultaneously evaluates multi-species experimental models within a particular biological condition using orthologous genes as linkers for the various Affymetrix microarray platforms on multi-species models of ventilator-associated lung injury. The results suggest that this approach may be a useful tool in the evaluation of biological processes of interest and selection of process-related candidate genes.     

Physiological Heterothallism and Sexuality in Euascomycetes: A Partial History

Copyright 1998 Academic Press.     

Functionality of Dengue Virus Specific Memory T Cell Responses in Individuals Who Were Hospitalized or Who Had Mild or Subclinical Dengue Infection

Background

Although antibody responses to dengue virus (DENV) in naturally infected individuals have been extensively studied, the functionality of DENV specific memory T cell responses in relation to clinical disease severity is incompletely understood.

Methodology/Principal findings

Using ex vivo IFNγ ELISpot assays, and by determining cytokines produced in ELISpot supernatants, we investigated the functionality of DENV-specific memory T cell responses in a large cohort of individuals from Sri Lanka (n=338), who were naturally infected and were either hospitalized due to dengue or had mild or sub clinical dengue infection. We found that T cells of individuals with both past mild or sub clinical dengue infection and who were hospitalized produced multiple cytokines when stimulated with DENV-NS3 peptides. However, while DENV-NS3 specific T cells of those with mild/sub clinical dengue infection were more likely to produce only granzyme B (p=0.02), those who were hospitalized were more likely to produce both TNFα and IFNγ (p=0.03) or TNFα alone.We have also investigated the usefulness of a novel T cell based assay, which can be used to determine the past infecting DENV serotype. 92.4% of DENV seropositive individuals responded to at least one DENV serotype of this assay and none of the seronegatives responded. Individuals who were seronegative, but had received the Japanese encephalitis vaccine too made no responses, suggesting that the peptides used in this assay did not cross react with the Japanese encephalitis virus.

Conclusions/significance

The types of cytokines produced by DENV-specific memory T cells appear to influence the outcome of clinical disease severity. The novel T cell based assay, is likely to be useful in determining the past infecting DENV serotype in immune-epidemiological studies and also in dengue vaccine trials.     

HLA class I and class II associations in dengue viral infections in a Sri Lankan population

Background

HLA class I and class II alleles have been shown to be associated with the development of dengue hemorrhagic fever (DHF)/dengue shock syndrome (DSS) in different populations. However, the majority of studies have been based on limited numbers of patients. In this study we aimed to investigate the HLA-class I and class II alleles that are positively and negatively associated with the development of DSS in a cohort of patients with DHF and also the alleles associated with development of DHF during primary dengue infections in a Sri Lankan population.

Methodology/Principal Findings

The allele frequencies of HLA class I and class II alleles were compared in 110 patients with DHF and 119 individuals from the population who had never reported a symptomatic dengue infection at the time of recruitment. We found that HLA-A*31 (corrected P = 0.01) and DRB1*08 (corrected P = 0.009) were associated with susceptibility to DSS when infected with the dengue virus, during secondary dengue infection. The frequency of DRB1*08 allele was 28.7 times higher than in the normal population in patients with DSS. HLA-A*31 allele was increased 16.6 fold in DHF who developed shock when compared to those who did not develop shock. A*24 (corrected P = 0.03) and DRB1*12 (corrected P = 0.041) were strongly associated with the development of DHF during primary dengue infection.

Conclusions/Significance

These data suggest that certain HLA alleles confer susceptibility/protection to severe dengue infections. As T cell epitope recognition depend on the HLA type of an individual, it would be now important to investigate how epitope specific T cells associate with primary and secondary dengue infections and in severe dengue infections.     

Expression of Toll-like receptor 2 is up-regulated in monocytes from patients with chronic obstructive pulmonary disease

Background

Chronic obstructive pulmonary disease (COPD) is characterised by pulmonary and systemic inflammation which flare-up during episodes of acute exacerbation (AECOPD). Given the role of Toll-like receptors (TLRs) in the induction of inflammatory responses we investigated the involvement of TLRs in COPD pathogenesis.

Methods

The expression of TLR-2, TLR-4 and CD14 in monocytes was analyzed by flow cytometry. To study the functional responses of these receptors, monocytes were stimulated with peptidoglycan or lipopolysaccharide and the amounts of TNFα and IL-6 secreted were determined by ELISA.

Results

We found that the expression of TLR-2 was up-regulated in peripheral blood monocytes from COPD patients, either clinically stable or during AECOPD, as compared to never smokers or smokers with normal lung function. Upon stimulation with TLR-2 ligand monocytes from COPD patients secreted increased amounts of cytokines than similarly stimulated monocytes from never smokers and smokers. In contrast, the expressions of TLR-4 and CD14 were not significantly different between groups, and the response to lipopolysaccharide (a TLR-4 ligand) stimulation was not significantly different either. At discharge from hospital TLR-2 expression was down-regulated in peripheral blood monocytes from AECOPD patients. This could be due to the treatment with systemic steroids because, in vitro, steroids down-regulated TLR-2 expression in a dose-dependent manner. Finally, we demonstrated that IL-6, whose plasma levels are elevated in patients, up-regulated in vitro TLR-2 expression in monocytes from never smokers.

Conclusion

Our results reveal abnormalities in TLRs expression in COPD patients and highlight its potential relationship with systemic inflammation in these patients.

     

Big game hunting in New Zealand: per capita effort,harvest and expenditure in 2011–2012

Recreational big game hunters make a significant contribution to conservation through kills of deer, pigs, chamois and tahr. New opportunities for managing recreational hunting through the proposed Game Animal Council underscore the need to understand the implications of potential changes in recreational hunting participation and harvests. Based on a survey of hunters' recall over a year, hunters averaged 15.63 (SEM = 0.58) big game hunts per year, spending 30.53 (SEM = 0.85) days hunting and killing 8.92 (SEM = 0.69) big game animals. Hunters commonly targeted several species on a single hunt, with highly skewed distributions for hunter effort and kills. Mean monthly expenditure on big game hunting items was $296.78 (SEM = $8.95). Results demonstrate that big game hunting is a significant activity in New Zealand, but this varies considerably among hunters with a small number responsible for the vast majority of kills. These are important considerations for future big game hunting management.     

Expression of AMPA receptor subunits at synapses in laminae I–III of the rodent spinal dorsal horn

Neuropathic pain may arise following peripheral nerve injury though the molecular mechanisms associated with this are unclear. We used proteomic profiling to examine changes in protein expression associated with the formation of hyper-excitable neuromas derived from rodent saphenous nerves. A two-dimensional difference gel electrophoresis (2D-DIGE) profiling strategy was employed to examine protein expression changes between developing neuromas and normal nerves in whole tissue lysates. We found around 200 proteins which displayed a >1.75-fold change in expression between neuroma and normal nerve and identified 55 of these proteins using mass spectrometry. We also used immunoblotting to examine the expression of low-abundance ion channels Nav1.3, Nav1.8 and calcium channel α2δ-1 subunit in this model, since they have previously been implicated in neuronal hyperexcitability associated with neuropathic pain. Finally, S³⁵methionine in vitro labelling of neuroma and control samples was used to demonstrate local protein synthesis of neuron-specific genes. A number of cytoskeletal proteins, enzymes and proteins associated with oxidative stress were up-regulated in neuromas, whilst overall levels of voltage-gated ion channel proteins were unaffected. We conclude that altered mRNA levels reported in the somata of damaged DRG neurons do not necessarily reflect levels of altered proteins in hyper-excitable damaged nerve endings. An altered repertoire of protein expression, local protein synthesis and topological re-arrangements of ion channels may all play important roles in neuroma hyper-excitability.     

Trichogynes and Fertilization in Uni- and Bimating Type Colonies of Neurospora tetrasperma

Microscopic examination of living, protoperithecium-bearing colonies of a, A, and a + A that have been challenged by macroconidia from the same three colony types has shown that active trichogynes, i.e., those that grow to and fuse with a conidium, are to be found only in the first two types. Thus, in the a colony the trichogynes respond to conidia from the A and a + A colonies while in the A colony they respond to conidia from a and a + A colonies. In contrast to this ability of conidia from a + A colonies to function as fertilizing elements, the trichogynes of these colonies, if indeed they are formed at all, do not so respond. This nonresponse in a + A colonies may be due to the perithecia that are developing at the time of the challenge. Evidence for this conclusion comes from unimating type colonies in which the two halves of each colony were challenged at different times, 48 h apart. Trichogynes and perithecia developed in the first half; neither developed in the second. This inhibition of trichogyne development and response in the presence of developing perithecia may be only one manifestation of a more general inhibitory action by these structures.     

Cellular and Cytokine Correlates of Severe Dengue Infection

Background

The occurrence of dengue haemorrhagic fever (DHF) is thought to result from a complex interplay between the virus, host genetics and host immune factors. Existing published data are not consistent, in part related to relatively small sample sizes. We set out to determine possible associations between dengue virus (DEN-V) NS3 specific T cells and cytokine and chemokine levels and the pathogenesis of severe disease in a large cohort of individuals with DHF.

Methodology/Principal Findings

By using ex vivo IFNγ ELISpot assays we determined DENV-NS3 specific responses in patients with varying severity of DHF. Other cytokines produced by DENV-NS3 specific T cells were determined by using multiple bead array analysis (MBAA). We also determined the serum cytokine levels using MBAA, lymphocyte subsets and Annexin V expression of lymphocytes in patients with varying severity of DHF. Of the 112 DHF patients studied, 29 developed shock. Serum IL-10 and IP-10 levels positively and significantly correlated with T cell apoptosis while IL-10 levels inversely correlated with T cell numbers. In contrast, TGFß showed a very significant (P<0.0001) and positive correlation (Spearman’s R = 0.65) with the platelet counts, consistent with platelet release. We found that whilst patients with severe dengue had lower total T cell numbers, the DV-NS3 specific T cells persisted and produced high levels of IFNγ but not TNFα, IL-3, IL-13, IL-2, IL-10 or IL-17.

Conclusions/Significance

Our data suggest that serum IL-10, TNFα and TGFβ differentially associate with dengue disease severity.     

Calcium‐permeable AMPA receptors and silent synapses in cocaine‐conditioned place preference

Exposure to cocaine generates silent synapses in the nucleus accumbens (NAc), whose eventual unsilencing/maturation by recruitment of calcium‐permeable AMPA‐type glutamate receptors (CP‐AMPARs) after drug withdrawal results in profound remodeling of NAc neuro‐circuits. Silent synapse‐based NAc remodeling was shown to be critical for several drug‐induced behaviors, but its role in acquisition and retention of the association between drug rewarding effects and drug‐associated contexts has remained unclear. Here, we find that the postsynaptic proteins PSD‐93, PSD‐95, and SAP102 differentially regulate excitatory synapse properties in the NAc. Mice deficient for either of these scaffold proteins exhibit distinct maturation patterns of silent synapses and thus provided instructive animal models to examine the role of NAc silent synapse maturation in cocaine‐conditioned place preference (CPP). Wild‐type and knockout mice alike all acquired cocaine‐CPP and exhibited increased levels of silent synapses after drug‐context conditioning. However, the mice differed in CPP retention and CP‐AMPAR incorporation. Collectively, our results indicate that CP‐AMPAR‐mediated maturation of silent synapses in the NAc is a signature of drug–context association, but this maturation is not required for establishing or retaining cocaine‐CPP.