Long-term neurological sequela is common among herpes simplex encephalitis (HSE) survivors. Animal models for HSE are used to investigate mechanisms of acute disease, but little has been done to model chronic manifestations of HSE. The current study presents a detailed, systematic analysis of chronic neuropathology, including characterization of topography and sequential progression of degenerative lesions and inflammation. Subsequent to intranasal HSV-1 infection, inflammatory responses that were temporally and spatially distinct persisted in infected cortical and brain stem regions. Neutrophils were present exclusively within the olfactory bulb and brain stem regions during the acute phase of infection, while the chronic inflammation was marked by plasma cells, lymphocytes and activated microglia. The chronic lymphocytic infiltrate, cytokine production, and activated microglia were associated with the loss of cortical neuropile in the entorhinal cortex and hippocampus. Animals surviving the acute infection showed a spectrum of chronic lesions from decreased brain volume, neuronal loss, activated astrocytes, and glial scar formation to severe atrophy and cavitations of the cortex. These lesions were also associated with severe spatial memory deficits in surviving animals. Taken together, this model can be utilized to further investigate the mechanisms of neurological defects that follow in the wake of HSE.

Murine cytomegalovirus (MCMV) brain infection stimulates microglial cell-driven proinflammatory chemokine production which precedes the presence of brain-infiltrating systemic immune cells. Here, we show that in response to MCMV brain infection, antigen-specific CD8(+) T-cells migrated into the brain and persisted as long-lived memory cells. The role of these persistent T-cells in the brain is unclear because most of our understanding of antimicrobial T-cell responses comes from analyses of lymphoid tissue. Strikingly, memory T-cells isolated from the brain exhibited an effector phenotype and produced IFN-γ upon restimulation with viral peptide. Furthermore, we observed time-dependent and long-term activation of resident microglia, indicated by chronic MHC class II up-regulation and TNF-α production. The immune response in this immunologically restricted site persisted in the absence of active viral replication. Lymphocyte infiltrates were detected until 30 d p.i., with CD8(+) and CD4(+) T-cells present at a 3:1 ratio, respectively. We then investigated the role of IFN-γ in chronic microglial activation by using IFN-γ-knockout (GKO) mice. At 30 d p.i., GKO mice demonstrated a similar phenotypic brain infiltrate when compared to wild-type mice (Wt), however, MHC class II expression on microglia isolated from these GKO mice was significantly lower compared to Wt animals. When IFN-γ producing CD8(+) T-cells were reconstituted in GKO mice, MHC class II up-regulation on microglial cells was restored. Taken together, these results suggest that MCMV brain infection results in long-term persistence of antigen specific CD8(+) T-cells which produce IFN-γ and drive chronic microglial cell activation. This response was found to be dependent on IFN-γ production by viral Ag-specific T-cells during the chronic phase of disease.

BackgroundUsing a murine model of herpes simplex virus (HSV)-1 encephalitis, our laboratory has determined that induction of proinflammatory mediators in response to viral infection is largely mediated through a Toll-like receptor-2 (TLR2)-dependent mechanism. Published studies have shown that, like other inflammatory mediators, reactive oxygen species (ROS) are generated during viral brain infection. It is increasingly clear that ROS are responsible for facilitating secondary tissue damage during central nervous system infection and may contribute to neurotoxicity associated with herpes encephalitis.MethodsPurified microglial cell and mixed neural cell cultures were prepared from C57B/6 and TLR2-/- mice. Intracellular ROS production in cultured murine microglia was measured via 2', 7'-Dichlorofluorescin diacetate (DCFH-DA) oxidation. An assay for 8-isoprostane, a marker of lipid peroxidation, was utilized to measure free radical-associated cellular damage. Mixed neural cultures obtained from β-actin promoter-luciferase transgenic mice were used to detect neurotoxicity induced by HSV-infected microglia.ResultsStimulation with HSV-1 elevated intracellular ROS in wild-type microglial cell cultures, while TLR2-/- microglia displayed delayed and attenuated ROS production following viral infection. HSV-infected TLR2-/- microglia produced less neuronal oxidative damage to mixed neural cell cultures in comparison to HSV-infected wild-type microglia. Further, HSV-infected TLR2-/- microglia were found to be less cytotoxic to cultured neurons compared to HSV-infected wild-type microglia. These effects were associated with decreased activation of p38 MAPK and p42/p44 ERK in TLR2-/- mice.ConclusionsThese studies demonstrate the importance of microglial cell TLR2 in inducing oxidative stress and neuronal damage in response to viral infection.

Background Previous studies have shown that a western diet impairs, whereas physical exercise enhances hippocampus-dependent learning and memory. Both diet and exercise influence expression of hippocampal brain-derived neurotrophic factor (BDNF), which is associated with improved cognition. We hypothesized that exercise reverses diet-induced cognitive decline while increasing hippocampal BDNF. Methods To test the effects of exercise on hippocampal-dependent memory, we compared cognitive scores of Sprague-Dawley rats exercised by voluntary running wheel (RW) access or forced treadmill (TM) to sedentary (Sed) animals. Memory was tested by two-way active avoidance test (TWAA), in which animals are exposed to a brief shock in a specific chamber area. When an animal avoids, escapes or has reduced latency to do either, this is considered a measure of memory. In a second experiment, rats were fed either a high-fat diet or control diet for 16 weeks, then randomly assigned to running wheel access or sedentary condition, and TWAA memory was tested once a week for seven weeks of exercise intervention. Results Both groups of exercised animals had improved memory as indicated by reduced latency to avoid and escape shock, and increased avoid and escape episodes (p<0.05). Exposure to a high-fat diet resulted in poor performance during both the acquisition and retrieval phases of the memory test as compared to controls. Exercise reversed high-fat diet-induced memory impairment, and increased brain-derived neurotrophic factor (BDNF) in neurons of the hippocampal CA3 region. Conclusions These data suggest that exercise improves memory retrieval, particularly with respect to avoiding aversive stimuli, and may be beneficial in protecting against diet induced cognitive decline, likely via elevated BDNF in neurons of the CA3 region.

Wild-type mice control murine cytomegalovirus (MCMV) brain infection, but identical infection is lethal to animals deficient in interleukin (IL)-10. Here, we report that MCMV-infected IL-10 knockout (KO) mice displayed a marked increase in neutrophil infiltration into the infected, IL-10-deficient brain when compared to wild-type animals. Enhanced microglial cell activation, determined by MHC class II up-regulation, overexpression of CXCL2, and elevated P-selectin mRNA levels were observed. In vivo blocking of CXCL2 attenuated neutrophil infiltration and significantly improved the outcome of infection. Collectively, these data indicate that the absence of IL-10 results in pathologic neutrophil infiltration into MCMV-infected brains.