IL-4-secreting CD4+ T cells are crucial to the development of CD8+ T-cell responses against malaria liver stages.

CD4+ T cells are crucial to the development of CD8+ T cell responses against hepatocytes infected with malaria parasites. In the absence of CD4+ T cells, CD8+ T cells initiate a seemingly normal differentiation and proliferation during the first few days after immunization. However, this response fails to develop further and is reduced by more than 90%, compared to that observed in the presence of CD4+ T cells. We report here that interleukin-4 (IL-4) secreted by CD4+ T cells is essential to the full development of this CD8+ T cell response. This is the first demonstration that IL-4 is a mediator of CD4/CD8 cross-talk leading to the development of immunity against an infectious pathogen

Heparan sulfate proteoglycans: structure, protein interactions and cell signaling

Heparan sulfate proteoglycans are ubiquitously found at the cell surface and extracellular matrix in all the animal species. This review will focus on the structural characteristics of the heparan sulfate proteoglycans related to protein interactions leading to cell signaling. The heparan sulfate chains due to their vast structural diversity are able to bind and interact with a wide variety of proteins, such as growth factors, chemokines, morphogens, extracellular matrix components, enzymes, among others. There is a specificity directing the interactions of heparan sulfates and target proteins, regarding both the fine structure of the polysaccharide chain as well precise protein motifs. Heparan sulfates play a role in cellular signaling either as receptor or co-receptor for different ligands, and the activation of downstream pathways is related to phosphorylation of different cytosolic proteins either directly or involving cytoskeleton interactions leading to gene regulation. The role of the heparan sulfate proteoglycans in cellular signaling and endocytic uptake pathways is also discussed.Proteoglicanos de heparam sulfato são encontrados tanto superfície celular quanto na matriz extracelular em todas as espécies animais. Esta revisão tem enfoque nas características estruturais dos proteoglicanos de heparam sulfato e nas interações destes proteoglicanos com proteínas que levam à sinalização celular. As cadeias de heparam sulfato, devido a sua variedade estrutural, são capazes de se ligar e interagir com ampla gama de proteínas, como fatores de crescimento, quimiocinas, morfógenos, componentes da matriz extracelular, enzimas, entreoutros. Existe uma especificidade estrutural que direciona as interações dos heparam sulfatos e proteínas alvo. Esta especificidade está relacionada com a estrutura da cadeia do polissacarídeo e os motivos conservados da cadeia polipeptídica das proteínas envolvidas nesta interação. Os heparam sulfatos possuem papel na sinalização celular como receptores ou coreceptores para diferentes ligantes. Esta ligação dispara vias de sinalização celular levam à fosforilação de diversas proteínas citosólicas ou com ou sem interações diretas com o citoesqueleto, culminando na regulação gênica. O papel dos proteoglicanos de heparam sulfato na sinalização celular e vias de captação endocítica também são discutidas nesta revisão.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Universidade Federal de São Paulo (UNIFESP) Departamento de BioquímicaUniversidade Federal de São Paulo (UNIFESP) Departamento de OftalmologiaUNIFESP, Depto. de BioquímicaUNIFESP, Depto. de OftalmologiaSciEL

Amphipathic DNA polymers exhibit antiviral activity against systemic Murine Cytomegalovirus infection

<p>Abstract</p> <p>Background</p> <p>Phosphorothioated oligonucleotides (PS-ONs) have a sequence-independent, broad spectrum antiviral activity as amphipathic polymers (APs) and exhibit potent in vitro antiviral activity against a broad spectrum of herpesviruses: HSV-1, HSV-2, HCMV, VZV, EBV, and HHV-6A/B, and in vivo activity in a murine microbiocide model of genital HSV-2 infection. The activity of these agents against animal cytomegalovirus (CMV) infections in vitro and in vivo was therefore investigated.</p> <p>Results</p> <p>In vitro, a 40 mer degenerate AP (REP 9) inhibited both murine CMV (MCMV) and guinea pig CMV (GPCMV) with an IC₅₀of 0.045 μM and 0.16 μM, respectively, and a 40 mer poly C AP (REP 9C) inhibited MCMV with an IC₅₀of 0.05 μM. Addition of REP 9 to plaque assays during the first two hours of infection inhibited 78% of plaque formation whereas addition of REP 9 after 10 hours of infection did not significantly reduce the number of plaques, indicating that REP 9 antiviral activity against MCMV occurs at early times after infection. In a murine model of CMV infection, systemic treatment for 5 days significantly reduced virus replication in the spleens and livers of infected mice compared to saline-treated control mice. REP 9 and REP 9C were administered intraperitoneally for 5 consecutive days at 10 mg/kg, starting 2 days prior to MCMV infection. Splenomegaly was observed in infected mice treated with REP 9 but not in control mice or in REP 9 treated, uninfected mice, consistent with mild CpG-like activity. When REP 9C (which lacks CpG motifs) was compared to REP 9, it exhibited comparable antiviral activity as REP 9 but was not associated with splenomegaly. This suggests that the direct antiviral activity of APs is the predominant therapeutic mechanism <it>in vivo</it>. Moreover, REP 9C, which is acid stable, was effective when administered orally in combination with known permeation enhancers.</p> <p>Conclusion</p> <p>These studies indicate that APs exhibit potent, well tolerated antiviral activity against CMV infection in vivo and represent a new class of broad spectrum anti-herpetic agents.</p

CMV Infection Attenuates the Disease Course in a Murine Model of Multiple Sclerosis

Recent evidence in multiple sclerosis (MS) suggests that active CMV infection may result in more benign clinical disease. The goal of this pilot study was to determine whether underlying murine CMV (MCMV) infection affects the course of the Theiler's murine encephalitis virus (TMEV) induced murine model of MS. A group of eight TMEV-infected mice were co-infected with MCMV at 2 weeks prior to TMEV infection while a second group of TMEV-infected mice received MCMV two weeks post TMEV. We also used 2 control groups, where at the above time points MCMV was replaced with PBS. Outcome measures included (1) monthly monitoring of disability via rotarod for 8 months; (2) in vivo MRI for brain atrophy studies and (3) FACS analysis of brain infiltrating lymphocytes at 8 months post TMEV infection. Co-infection with MCMV influenced the disease course in mice infected prior to TMEV infection. In this group, rotarod detectable motor performance was significantly improved starting 3 months post-infection and beyond (p≤0.024). In addition, their brain atrophy was close to 30% reduced at 8 months, but this was only present as a trend due to low power (p = 0.19). A significant reduction in the proportion of brain infiltrating CD3+ cells was detected in this group (p = 0.026), while the proportion of CD45+ Mac1+ cells significantly increased (p = 0.003). There was also a strong trend for a reduced proportion of CD4+ cells (p = 0.17) while CD8 and B220+ cell proportion did not change. These findings support an immunomodulatory effect of MCMV infection in this MS model. Future studies in this co-infection model will provide insight into mechanisms which modulate the development of demyelination and may be utilized for the development of novel therapeutic strategies

Membrane Properties and the Balance between Excitation and Inhibition Control Gamma-Frequency Oscillations Arising from Feedback Inhibition

Computational studies as well as in vivo and in vitro results have shown that many cortical neurons fire in a highly irregular manner and at low average firing rates. These patterns seem to persist even when highly rhythmic signals are recorded by local field potential electrodes or other methods that quantify the summed behavior of a local population. Models of the 30–80 Hz gamma rhythm in which network oscillations arise through ‘stochastic synchrony’ capture the variability observed in the spike output of single cells while preserving network-level organization. We extend upon these results by constructing model networks constrained by experimental measurements and using them to probe the effect of biophysical parameters on network-level activity. We find in simulations that gamma-frequency oscillations are enabled by a high level of incoherent synaptic conductance input, similar to the barrage of noisy synaptic input that cortical neurons have been shown to receive in vivo. This incoherent synaptic input increases the emergent network frequency by shortening the time scale of the membrane in excitatory neurons and by reducing the temporal separation between excitation and inhibition due to decreased spike latency in inhibitory neurons. These mechanisms are demonstrated in simulations and in vitro current-clamp and dynamic-clamp experiments. Simulation results further indicate that the membrane potential noise amplitude has a large impact on network frequency and that the balance between excitatory and inhibitory currents controls network stability and sensitivity to external inputs

A Novel Cre Recombinase Imaging System for Tracking Lymphotropic Virus Infection In Vivo

BACKGROUND:Detection, isolation, and identification of individual virus infected cells during long term infection are critical to advance our understanding of mechanisms of pathogenesis for latent/persistent viruses. However, current approaches to study these viruses in vivo have been hampered by low sensitivity and effects of cell-type on expression of viral encoded reporter genes. We have designed a novel Cre recombinase (Cre)-based murine system to overcome these problems, and thereby enable tracking and isolation of individual in vivo infected cells. METHODOLOGY/PRINCIPAL FINDINGS:Murine gammaherpesvirus 68 (MHV-68) was used as a prototypic persistent model virus. A Cre expressing recombinant virus was constructed and characterised. The virus is attenuated both in lytic virus replication, producing ten-fold lower lung virus titres than wild type virus, and in the establishment of latency. However, despite this limitation, when the sEGFP7 mouse line containing a Cre-activated enhanced green fluorescent protein (EGFP) was infected with the Cre expressing virus, sites of latent and persistent virus infection could be identified within B cells and macrophages of the lymphoid system on the basis of EGFP expression. Importantly, the use of the sEGFP7 mouse line which expresses high levels of EGFP allowed individual virus positive cells to be purified by FACSorting. Virus gene expression could be detected in these cells. Low numbers of EGFP positive cells could also be detected in the bone marrow. CONCLUSIONS/SIGNIFICANCE:The use of this novel Cre-based virus/mouse system allowed identification of individual latently infected cells in vivo and may be useful for the study and long-term monitoring of other latent/persistent virus infections

Sparse Gamma Rhythms Arising through Clustering in Adapting Neuronal Networks

Gamma rhythms (30–100 Hz) are an extensively studied synchronous brain state responsible for a number of sensory, memory, and motor processes. Experimental evidence suggests that fast-spiking interneurons are responsible for carrying the high frequency components of the rhythm, while regular-spiking pyramidal neurons fire sparsely. We propose that a combination of spike frequency adaptation and global inhibition may be responsible for this behavior. Excitatory neurons form several clusters that fire every few cycles of the fast oscillation. This is first shown in a detailed biophysical network model and then analyzed thoroughly in an idealized model. We exploit the fact that the timescale of adaptation is much slower than that of the other variables. Singular perturbation theory is used to derive an approximate periodic solution for a single spiking unit. This is then used to predict the relationship between the number of clusters arising spontaneously in the network as it relates to the adaptation time constant. We compare this to a complementary analysis that employs a weak coupling assumption to predict the first Fourier mode to destabilize from the incoherent state of an associated phase model as the external noise is reduced. Both approaches predict the same scaling of cluster number with respect to the adaptation time constant, which is corroborated in numerical simulations of the full system. Thus, we develop several testable predictions regarding the formation and characteristics of gamma rhythms with sparsely firing excitatory neurons

Ion Channel Density Regulates Switches between Regular and Fast Spiking in Soma but Not in Axons

The threshold firing frequency of a neuron is a characterizing feature of its dynamical behaviour, in turn determining its role in the oscillatory activity of the brain. Two main types of dynamics have been identified in brain neurons. Type 1 dynamics (regular spiking) shows a continuous relationship between frequency and stimulation current (f-Istim) and, thus, an arbitrarily low frequency at threshold current; Type 2 (fast spiking) shows a discontinuous f-Istim relationship and a minimum threshold frequency. In a previous study of a hippocampal neuron model, we demonstrated that its dynamics could be of both Type 1 and Type 2, depending on ion channel density. In the present study we analyse the effect of varying channel density on threshold firing frequency on two well-studied axon membranes, namely the frog myelinated axon and the squid giant axon. Moreover, we analyse the hippocampal neuron model in more detail. The models are all based on voltage-clamp studies, thus comprising experimentally measurable parameters. The choice of analysing effects of channel density modifications is due to their physiological and pharmacological relevance. We show, using bifurcation analysis, that both axon models display exclusively Type 2 dynamics, independently of ion channel density. Nevertheless, both models have a region in the channel-density plane characterized by an N-shaped steady-state current-voltage relationship (a prerequisite for Type 1 dynamics and associated with this type of dynamics in the hippocampal model). In summary, our results suggest that the hippocampal soma and the two axon membranes represent two distinct kinds of membranes; membranes with a channel-density dependent switching between Type 1 and 2 dynamics, and membranes with a channel-density independent dynamics. The difference between the two membrane types suggests functional differences, compatible with a more flexible role of the soma membrane than that of the axon membrane

A novel role of dendritic gap junction and mechanisms underlying its interaction with thalamocortical conductance in fast spiking inhibitory neurons

<p>Abstract</p> <p>Background</p> <p>Little is known about the roles of dendritic gap junctions (GJs) of inhibitory interneurons in modulating temporal properties of sensory induced responses in sensory cortices. Electrophysiological dual patch-clamp recording and computational simulation methods were used in combination to examine a novel role of GJs in sensory mediated feed-forward inhibitory responses in barrel cortex layer IV and its underlying mechanisms.</p> <p>Results</p> <p>Under physiological conditions, excitatory post-junctional potentials (EPJPs) interact with thalamocortical (TC) inputs within an unprecedented few milliseconds (i.e. over 200 Hz) to enhance the firing probability and synchrony of coupled fast-spiking (FS) cells. Dendritic GJ coupling allows fourfold increase in synchrony and a significant enhancement in spike transmission efficacy in excitatory spiny stellate cells. The model revealed the following novel mechanisms: <b><it>1) </it></b>rapid capacitive current (I_cap) underlies the activation of voltage-gated sodium channels; <b><it>2) </it></b>there was less than 2 milliseconds in which the I_capunderlying TC input and EPJP was coupled effectively; <b><it>3) </it></b>cells with dendritic GJs had larger input conductance and smaller membrane response to weaker inputs; <b><it>4) </it></b>synchrony in inhibitory networks by GJ coupling leads to reduced sporadic lateral inhibition and increased TC transmission efficacy.</p> <p>Conclusion</p> <p>Dendritic GJs of neocortical inhibitory networks can have very powerful effects in modulating the strength and the temporal properties of sensory induced feed-forward inhibitory and excitatory responses at a very high frequency band (>200 Hz). Rapid capacitive currents are identified as main mechanisms underlying interaction between two transient synaptic conductances.</p

IgG Fc Receptors Provide an Alternative Infection Route for Murine Gamma-Herpesvirus-68

BACKGROUND: Herpesviruses can be neutralized in vitro but remain infectious in immune hosts. One difference between these settings is the availability of immunoglobulin Fc receptors. The question therefore arises whether a herpesvirus exposed to apparently neutralizing antibody can still infect Fc receptor(+) cells. PRINCIPAL FINDINGS: Immune sera blocked murine gamma-herpesvirus-68 (MHV-68) infection of fibroblasts, but failed to block and even enhanced its infection of macrophages and dendritic cells. Viral glycoprotein-specific monoclonal antibodies also enhanced infection. MHV-68 appeared to be predominantly latent in macrophages regardless of whether Fc receptors were engaged, but the infection was not abortive and new virus production soon overwhelmed infected cultures. Lytically infected macrophages down-regulated MHC class I-restricted antigen presentation, endocytosis and their response to LPS. CONCLUSIONS: IgG Fc receptors limit the neutralization of gamma-herpesviruses such as MHV-68