Gang Xin

Apoptosis and Proinflammatory Cytokine Responses of Primary Mouse Microglia and Astrocytes Induced by Human H1N1 and Avian H5N1 Influenza Viruses

Patients with an influenza virus infection can be complicated by acute encephalopathy and encephalitis. To investigate the immune reactions involved in the neurocomplication, mouse microglia and astrocytes were isolated, infected with human H1N1 and avian H5N1 influenza viruses, and examined for their immune responses. We observed homogeneously distributed viral receptors, sialic acid (SA)-α2,3-Galactose (Gal) and SA-α2,6-Gal, on microglia and astrocytes. Both viruses were replicative and productive in microglia and astrocytes. Virus-induced apoptosis and cytopathy in infected cells were observed at 24 h post-infection (p.i.). Expression of IL-1β, IL-6 and TNF-α mRNA examined at 6 h and 24 h p.i. was up-regulated, and their expression levels were considerably higher in H5N1 infection. The amounts of secreted proinflammatory IL-1β, IL-6 and TNF-α at 6 h and 24 h p.i. were also induced, with greater induction by H5N1 infection. This study is the first demonstration that both human H1N1 and avian H5N1 influenza viruses can infect mouse microglia and astrocytes and induce apoptosis, cytopathy, and proinflammatory cytokine production in them in vitro. Our results suggest that the direct cellular damage and the consequences of immunopathological injury in the CNS contribute to the influenza viral pathogenesis.

Serum BAFF is elevated in patients with IgA nephropathy and associated with clinical and histopathological features.

BACKGROUND B-cell-activating factor belonging to the tumor necrosis factor family (BAFF) has been found to have the function of activating B cells and participating in the class switching of B cells; however, its clinical application needs further study. In the present study, the serum BAFF levels of patients with IgA nephropathy (IgAN) with different histopathological phenotypes were measured. METHODS Levels of serum BAFF in 153 patients with IgAN, 55 healthy controls and 20 disease controls were recorded using commercially available ELISA kits. Their correlations with clinical and histopathological features of patients with IgAN were further evaluated. RESULTS Levels of serum BAFF in patients with IgAN were significantly higher than in controls. Serum BAFF levels were significantly higher in patients with mesangial hypercellularity and segmental glomerulosclerosis than in those without. Serum BAFF levels were associated with the severity of tubular atrophy/interstitial fibrosis. Serum BAFF levels were significantly positively correlated with estimated glomerular filtration rate and serum creatinine. Patients with elevated serum BAFF levels showed significantly greater severity in clinical and histopathological stages. CONCLUSION Levels of serum BAFF were elevated in patients with IgAN and were associated with clinical and pathological features of the disease. Serum BAFF levels could be a noninvasive biomarker for monitoring disease severity of IgAN.

Effects of NS1 variants of H5N1 influenza virus on interferon induction, TNFα response and p53 activity

Non-structural protein 1 (NS1) is an important virulence factor of the highly pathogenic H5N1 avian influenza virus. A five-amino-acid (5 aa) deletion at position 80–84 and an aspartic acid to glutamic acid substitution at position 92 (D92E) are two major NS1 mutations that are highly correlated with enhanced virulence. To investigate the effect of these mutations in H5N1 virulence, three H5N1-NS1 variants were constructed: NS51 (lacking 5 aa at position 80–84), NS51(I) (carrying a 5-aa insertion at position 80–84) and NS51(IM) (carrying both the 5-aa insertion and the D92E mutation). We examined the effects of these mutations on interferon (IFN) induction, tumor-necrosis factor (TNF)α response, p53 activity and apoptosis. We found that the D92E mutation eliminated NS1s repressive effect on IFN induction, while the 5-aa deletion resulted in enhanced resistance to TNFα responses. We also observed that all three variants exhibited a similar suppressive effect on p53 transcriptional activity, although none of them significantly influenced apoptosis of host cells. Our findings shed new light on the role of NS1 in the pathogenicity of H5N1 virus.

Serum B-cell Activating Factor in Myecloperoxiase-antineutrophil Cytoplasmic Antibodies-associated Vasculitis

Background:In this study, the serum B-cell activating factor belonging to tumor necrosis factor family (BAFF) levels in patients with myeloperoxidase (MPO)-antineutrophil cytoplasmic antibodies (ANCA)–associated vasculitis (AAV) were measured, and their clinical significance was further analyzed. Methods:One hundred twenty-one patients with MPO-AAV were enrolled in this study. Eighty-three patients had active vasculitis and 38 were in remission. Fifty-five healthy individuals were used as healthy controls. The levels of serum BAFF were assessed using commercial available enzyme-linked immunosorbent assay kits. The correlations between serum BAFF and Birmingham Vasculitis Activity Score, erythrocyte sedimentation rate and MPO-ANCA were further evaluated. Results:The levels of serum BAFF of patients with MPO-AAV in both active (6.06 ± 5.02 ng/mL) and remission phases (3.60 ± 3.83 ng/mL) were significantly higher than those in healthy controls (0.87 ± 0.31 ng/mL) (P < 0.001, respectively). The serum BAFF levels in patients with active vasculitis were significantly higher than those in remission (P < 0.001). Serum BAFF levels were significantly correlated with Birmingham Vasculitis Activity Score (r = 0.320, P < 0.001) and erythrocyte sedimentation rate value (r = 0.311, P < 0.01) in all patients, but no correlation was found between the levels of serum BAFF and MPO-ANCA. Using receiver-operating characteristics statistics, the cutoff values of serum BAFF level for indicating the presence of MPO-AAV and active vasculitis were 1.58 and 4.20 ng/mL, respectively. Conclusions:The levels of serum BAFF were elevated in patients with MPO-AAV and associated with disease activity, but they were not related with the levels of MPO-ANCA.

Serum BAFF and APRIL might be associated with disease activity and kidney damage in patients with anti‐glomerular basement membrane disease

B cell activating factor belonging to the tumour necrosis factor family (BAFF) and a proliferation inducing ligand (APRIL) are two tumour necrosis factor (TNF)‐like cytokines that were found to be elevated in many autoimmune diseases. Anti‐glomerular basement membrane (GBM) disease is a typical severe autoimmune disease characterized by raised serum anti‐GBM antibodies. In this study we aimed to detect the serum levels of BAFF and APRIL in patients with anti‐GBM disease, and their clinical significance was further analyzed.

Induction of cytopathic effect and cytokines in coxsackievirus B3-infected murine astrocytes

BackgroundCoxsackievirus commonly infects children and occasionally causes severe meningitis and/or encephalitis in the newborn. The underlying mechanism(s) behind the central nervous system pathology is poorly defined.MethodsIt is hypothesized that astrocytes may be involved in inflammatory response induced by CVB3 infection. Here we discuss this hypothesis in the context of CVB3 infection and associated inflammatory response in primary mouse astrocytes.ResultsThe results showed that coxsackievirus receptor (CAR) was distributed homogeneously on the astrocytes, and that CVB3 could infect and replicate in astrocytes, with release of infectious virus particles. CVB3 induced cytopathic effect and production of proinflammatory cytokines IL-1β, TNF-α, IL-6, and chemokine CXCL10 from astrocytes.ConclusionThese data suggest that direct astrocyte damage and cytokines induction could be a mechanism of virus-induced meningitis and/or encephalitis.

An attenuated coxsackievirus b3 vector: a potential tool for viral tracking study and gene delivery.

Cardiomyocytes are quite resistant to gene transfer using standard techniques. We developed an expression vector carrying an attenuated but infectious and replicative coxsackievirus B3 (CVB3) genome, and unique ClaI-StuI cloning sites for an exogenous gene, whose product can be released from the nascent viral polyprotein by 2Apro cleavage. This vector was tested as an expression vehicle for green fluorescent protein (GFP). The vector transiently expressed GFP in cell cultures for at least ten passages and delivered functional GFP to the infected cardiomyocytes for at least 6 days. Moreover, the recombinant viruses showed virulence attenuation in vitro and in vivo. The findings suggest that the recombinant CVB3 vector could be a useful tool for viral tracking study and delivering exogenous proteins to cardiomyocytes.

LPS-Primed Release of HMGB-1 from Cortical Astrocytes is Modulated Through PI3K/AKT Pathway

Abstract Studies have shown that LPS-preconditioned tolerant state could protect against brain injury to subsequent challenges. We hypothesized astrocytes were directly involved in the readjustment to confer neuroprotective effects with LPS pretreatment. High-mobility group box 1(HMGB-1) from LPS-preconditioned astrocytes, presumably serving as a positive regulator, might contribute to the favorable preconditioned effects. Furthermore, a potential cellular pathway (PI3K/AKT pathway), has been proposed for the active regulation of LPS-primed reactive astrocytes to secrete HMGB-1. In the present study, we used a low concentration of LPS to directly prime the astrocytes in vitro, and the subsequent astrocytic reactions, including cytokine secretion, the expression of transcription factors, and the release of HMGB-1 were examined after the blockade of the PI3K pathway. The data showed that LPS preconditioning could reduce some capacity of astrocytes to subsequent challenge in vitro. PI3K/AKT pathway was partially involved in the modulation of the release HMGB-1 from reactive astrocytes. These findings offer direct evidence supporting the flexible roles of astrocytes in mediating LPS-primed neuroprotection, and highlight additional targets for future attempts to modify the protective effects of astrocytes through LPS preconditioning.

Predator exposure-induced cerebral interleukins are modulated heterogeneously by behavioral asymmetry.

Predator exposure is a naturalistic stressor that is likely to elicit a stressful response pattern similar to those experienced in the real world. As a consequence of stress, HPA hormonal activity and the alteration of mediators such as cytokines may result. Behavioral asymmetry, as assessed by paw preference, exerted effects on immune responses and peripheral cytokine production, observed after exposure to the physical stimuli. Thus, we hypothesized that behavioral asymmetry can modulate mouse brain interleukins and HPA activity after exposure to an internally generated psychological stress source. To determine the impact of behavioral asymmetry, mice were divided into left- and right-pawed groups by paw preference. Then, the mice received either a single 60-min or a daily 60-min predator exposure (cat exposure) for 14 consecutive days. After receiving predator exposure, trunk blood was collected and brain tissues, including the cerebral cortex, hippocampus and hypothalamus, were separated. Plasma corticosterone (CS) was detected by EIA, and IL-1β and IL-6 levels in the cortex, hippocampus and hypothalamus, were quantified by ELISA. The results revealed that predator stress, in particular chronic stress, could enhance plasma CS concentration and could alter IL-1β and IL-6 concentrations in the cortex, hippocampus and hypothalamus. Simultaneously, predator stress-induced CS and brain interleukin levels were modulated by behavioral asymmetry. The left-pawed mice showed a decreased variation in CS, less than right-pawed mice, and both left-pawed and right-pawed mice displayed heterogeneous direction and intensity of changes for IL-1β and IL-6 in the cortex, hippocampus and hypothalamus after predator exposure. From these results, it can be concluded that the alteration of cytokines depends on the characteristics of the stressor. Furthermore, the asymmetric cytokine responses within the brain to a natural, psychological stressor may be involved in the immunomodulation of behavioral asymmetry. These findings likely reflect the flexibility in reactivity patterns of a population in response to various insults.

Lipopolysaccharide enhances asymmetrical production of cytokines and nitric oxide by left and right cerebral cortical microglial cells in BALB/C mice

Lipopolysaccharide (LPS)‐induced inflammatory factors production by the cerebral cortical glial cells in two sides of the murine brain are different. To determine if microglial cells, a subset of glial cells, are involved in asymmetric production, interleukin‐6 (IL‐6), interleukin‐1β (IL‐1β) and nitric oxide (NO) responses to LPS by microglial cells in the right and left cerebral cortices were examined. Primary microglial cells were isolated from BALB/C neonatal mice, treated with LPS (10 µg ml−1) for 24 h and examined for IL‐6, IL‐1β and NO production. At untreated state, the levels of IL‐6, IL‐1β and NO showed no statistical difference between left and right. However, after LPS treatment, the levels of IL‐6, IL‐1β and NO for the right microglial cells was statistically significant higher than the left (P < 0·05). Our results denote that enhanced production of IL‐6, IL‐1β and NO after LPS treatment in microglia is directly proportional to their basal‐state levels, and right cortical microglia produce higher levels of IL‐6, IL‐1β and NO than left cortical microglia. Copyright