Virus-induced Immune Responses Research Articles

Upregulation of IFN-γ and IL-12 is associated with a milder form of hantavirus hemorrhagic fever with renal syndrome.

Hantavirus hemorrhagic fever with renal syndrome (HFRS) is a zoonotic disease characterized by acute onset, fever, malaise, and back pain. As the disease progresses, hemorrhagic disturbances and kidney dysfunctions predominate. The examination of tissue collected postmortem supports the premise that virus replication is not responsible for this pathology; therefore, it is widely believed that virus-induced immune responses lead to the clinical manifestations associated with HFRS. The overproduction of inflammatory cytokines is commonly reported in subjects with HFRS and has given rise to the hypothesis that a so-called "cytokine storm" may play a pivotal role in the pathogenesis of this disease. Currently, supportive care remains the only effective treatment for HFRS. Our data show that serum levels of interferon (IFN)-γ, interleukin (IL)-10, CCL2, and IL-12 are upregulated in HFRS cases when compared to healthy controls and the level of upregulation is dependent on the phase and severity of the disease. Furthermore, we observed an association between the mild form of the disease and elevated serum levels of IFN-γ and IL-12. Collectively, these observations suggest that the administration of exogenous IFN-γ and IL-12 may provide antiviral benefits for the treatment of HFRS and, thus, warrants further investigations.

MicroRNA‐155 controls Toll‐like receptor 3‐ and hepatitis C virus‐induced immune responses in the liver

The hepatitis C virus (HCV) establishes persistent infections despite strong activation of the innate immune system through TLR3 and other sensors. Therefore, we analysed regulatory mechanisms of TLR3-induced immune responses in nonparenchymal liver cells (NPCs). Effects of Interleukin-10 (IL-10), transforming growth factor beta (TGF-β) and immunoregulatory miR-155 on poly I:C-activated murine (C57BL/6) Kupffer cells (KC) and sinusoidal endothelial cells (LSEC) were assessed in vitro. NPCs were assayed for inflammatory and antiviral cytokines and T-cell (Balb/c)-activating factors. Gene expression of miR-155, IL-10, TGF-β and interferon sensitive genes (ISGs) in biopsies of patients with HCV was determined by qrt-PCR. TLR3-induced antiviral activity in murine NPCs was potently suppressed by IL-10 and TGF-β which correlated with decreased TLR3 expression and inhibition of NF-κB and IRF-3 activation. T-cell activation, induced by TLR3-activated NPCs, was also suppressed by IL-10 and TGF-β, which was associated with a down-regulation of CD80 and CD86. Pretreatment with IL-10 or TGF-β suppressed TLR3-induced miR-155 expression, which itself positively regulated poly I:C-mediated immune responses, thus counteracting IL-10 or TGF-β-induced immunosuppression. In addition, hepatic expression of miR-155 was elevated in chronically infected patients with HCV, was associated with an IL-28B SNP (rs12979860) and was inversely correlated with HCV serum load and ISG expression levels. As miR-155 is a key regulator of anti-inflammatory mechanisms that control innate and adaptive hepatic immune responses during HCV infection, miR-155 based therapies may represent a novel mechanism to control HCV in the future.

Response to Rhinovirus Infection by Human Airway Epithelial Cells and Peripheral Blood Mononuclear Cells in an In Vitro Two-Chamber Tissue Culture System

Human rhinovirus (HRV) infections are associated with the common cold, occasionally with more serious lower respiratory tract illnesses, and frequently with asthma exacerbations. The clinical features of HRV infection and its association with asthma exacerbation suggest that some HRV disease results from virus-induced host immune responses to infection. To study the HRV-infection-induced host responses and the contribution of these responses to disease, we have developed an in vitro model of HRV infection of human airway epithelial cells (Calu-3 cells) and subsequent exposure of human peripheral blood mononuclear cells (PBMCs) to these infected cells in a two-chamber trans-well tissue culture system. Using this model, we studied HRV 14 (species B) and HRV 16 (species A) induced cytokine and chemokine responses with PBMCs from four healthy adults. Infection of Calu-3 cells with either virus induced HRV-associated increases in FGF-Basic, IL-15, IL-6, IL-28A, ENA-78 and IP-10. The addition of PBMCs to HRV 14-infected cells gave significant increases in MIP-1β, IL-28A, MCP-2, and IFN-α as compared with mock-infected cells. Interestingly, ENA-78 levels were reduced in HRV 14 infected cells that were exposed to PBMCs. Addition of PBMCs to HRV 16-infected cells did not induce MIP-1β, IL-28A and IFN-α efficiently nor did it decrease ENA-78 levels. Our results demonstrate a clear difference between HRV 14 and HRV 16 and the source of PBMCs, in up or down regulation of several cytokines including those that are linked to airway inflammation. Such differences might be one of the reasons for variation in disease associated with different HRV species including variation in their link to asthma exacerbations as suggested by other studies. Further study of immune responses associated with different HRVs and PBMCs from different patient groups, and the mechanisms leading to these differences, should help characterize pathogenesis of HRV disease and generate novel approaches to its treatment.

Genome-wide RNAi Screen Reveals a New Role of a WNT/CTNNB1 Signaling Pathway as Negative Regulator of Virus-induced Innate Immune Responses

To identify new regulators of antiviral innate immunity, we completed the first genome-wide gene silencing screen assessing the transcriptional response at the interferon-β (IFNB1) promoter following Sendai virus (SeV) infection. We now report a novel link between WNT signaling pathway and the modulation of retinoic acid-inducible gene I (RIG-I)-like receptor (RLR)-dependent innate immune responses. Here we show that secretion of WNT2B and WNT9B and stabilization of β-catenin (CTNNB1) upon virus infection negatively regulate expression of representative inducible genes IFNB1, IFIT1 and TNF in a CTNNB1-dependent effector mechanism. The antiviral response is drastically reduced by glycogen synthase kinase 3 (GSK3) inhibitors but restored in CTNNB1 knockdown cells. The findings confirm a novel regulation of antiviral innate immunity by a canonical-like WNT/CTNNB1 signaling pathway. The study identifies novel avenues for broad-spectrum antiviral targets and preventing immune-mediated diseases upon viral infection.

Acute Renal Failure in a Child With Thrombocytopenic Purpura Caused by Acute Epstein-Barr Virus Infection After Treatment With Anti–D Immunoglobulin

Immune thrombocytopenia (ITP) in children is usually a benign, self-limiting disorder. An acute Epstein-Barr virus (EBV) infection usually causes atypical lymphocytosis and mild decrease in platelets. Severe thrombocytopenia is an extremely rare complication. Anti-D immunoglobulin has been used for treatment of ITP in Rh(D)-positive nonsplenectomized patients. Severe hemolysis and acute renal failure are extremely rare complications that may be aggravated by the presence of an acute EBV infection. It is believed that anti-D immunoglobulin triggers an unusual virus-induced immune response causing hemolysis. We present a 4-year-old girl with ITP caused by an acute EBV infection that developed acute kidney injury following treatment with anti-D immunoglobulin. The patient recovered completely from thrombocytopenia and renal dysfunction. Intravascular hemolysis and acute kidney injury are consistent with anti-D immunoglobulin mechanism of action. Pediatric patients treated with anti-D immunoglobulin for ITP should be closely monitored for signs and symptoms of hemolysis that may be aggravated by the presence of EBV infection leading to impaired renal function.

Systematic Analysis of the Mechanisms of Virus-Triggered Type I IFN Signaling Pathways through Mathematical Modeling

Based on biological experimental data, we developed a mathematical model of the virus-triggered signaling pathways that lead to induction of type I IFNs and systematically analyzed the mechanisms of the cellular antiviral innate immune responses, including the negative feedback regulation of ISG56 and the positive feedback regulation of IFNs. We found that the time between 5 and 48 hours after viral infection is vital for the control and/or elimination of the virus from the host cells and demonstrated that the ISG56-induced inhibition of MITA activation is stronger than the ISG56-induced inhibition of TBK1 activation. The global parameter sensitivity analysis suggests that the positive feedback regulation of IFNs is very important in the innate antiviral system. Furthermore, the robustness of the innate immune signaling network was demonstrated using a new robustness index. These results can help us understand the mechanisms of the virus-induced innate immune response at a system level and provide instruction for further biological experiments.

UBXN1 Interferes with Rig-I-like Receptor-Mediated Antiviral Immune Response by Targeting MAVS

RNA viruses are sensed by RIG-I-like receptors (RLRs), which signal through a mitochondria-associated adaptor molecule, MAVS, resulting in systemic antiviral immune responses. Although RLR signaling is essential for limiting RNA virus replication, it must be stringently controlled to prevent damage from inflammation. We demonstrate here that among all tested UBX-domain-containing protein family members, UBXN1 exhibits the strongest inhibitory effect on RNA-virus-induced type I interferon response. UBXN1 potently inhibits RLR- and MAVS-induced, but not TLR3-, TLR4-, or DNA-virus-induced innate immune responses. Depletion of UBXN1 enhances virus-induced innate immune responses, including those resulting from RNA viruses such as vesicular stomatitis, Sendai, West Nile, and dengue virus infection, repressing viral replication. Following viral infection, UBXN1 is induced, binds to MAVS, interferes with intracellular MAVS oligomerization, and disrupts the MAVS/TRAF3/TRAF6 signalosome. These findings underscore a critical role of UBXN1 in the modulation of a major antiviral signaling pathway.

Modeling and Dynamical Analysis of Virus-Triggered Innate Immune Signaling Pathways

The investigation of the dynamics and regulation of virus-triggered innate immune signaling pathways at a system level will enable comprehensive analysis of the complex interactions that maintain the delicate balance between resistance to infection and viral disease. In this study, we developed a delayed mathematical model to describe the virus-induced interferon (IFN) signaling process by considering several key players in the innate immune response. Using dynamic analysis and numerical simulation, we evaluated the following predictions regarding the antiviral responses: (1) When the replication ratio of virus is less than 1, the infectious virus will be eliminated by the immune system’s defenses regardless of how the time delays are changed. (2) The IFN positive feedback regulation enhances the stability of the innate immune response and causes the immune system to present the bistability phenomenon. (3) The appropriate duration of viral replication and IFN feedback processes stabilizes the innate immune response. The predictions from the model were confirmed by monitoring the virus titer and IFN expression in infected cells. The results suggest that the balance between viral replication and IFN-induced feedback regulation coordinates the dynamical behavior of virus-triggered signaling and antiviral responses. This work will help clarify the mechanisms of the virus-induced innate immune response at a system level and provide instruction for further biological experiments.

Enterovirus-induced gene expression profile is critical for human pancreatic islet destruction

Virally induced inflammatory responses, beta cell destruction and release of beta cell autoantigens may lead to autoimmune reactions culminating in type 1 diabetes. Therefore, viral capability to induce beta cell death and the nature of virus-induced immune responses are among key determinants of diabetogenic viruses. We hypothesised that enterovirus infection induces a specific gene expression pattern that results in islet destruction and that such a host response pattern is not shared among all enterovirus infections but varies between virus strains. The changes in global gene expression and secreted cytokine profiles induced by lytic or benign enterovirus infections were studied in primary human pancreatic islet using DNA microarrays and viral strains either isolated at the clinical onset of type 1 diabetes or capable of causing a diabetes-like condition in mice. The expression of pro-inflammatory cytokine genes (IL-1-α, IL-1-β and TNF-α) that also mediate cytokine-induced beta cell dysfunction correlated with the lytic potential of a virus. Temporally increasing gene expression levels of double-stranded RNA recognition receptors, antiviral molecules, cytokines and chemokines were detected for all studied virus strains. Lytic coxsackievirus B5 (CBV-5)-DS infection also downregulated genes involved in glycolysis and insulin secretion. The results suggest a distinct, virus-strain-specific, gene expression pattern leading to pancreatic islet destruction and pro-inflammatory effects after enterovirus infection. However, neither viral replication nor cytotoxic cytokine production alone are sufficient to induce necrotic cell death. More likely the combined effect of these and possibly cellular energy depletion lie behind the enterovirus-induced necrosis of islets.

Accelerated and Improved Quantification of Lymphocytic Choriomeningitis Virus (LCMV) Titers by Flow Cytometry

Lymphocytic choriomeningitis virus (LCMV), a natural murine pathogen, is a member of the Arenavirus family, may cause atypical meningitis in humans, and has been utilized extensively as a model pathogen for the study of virus-induced disease and immune responses. Historically, viral titers have been quantified by a standard plaque assay, but for non-cytopathic viruses including LCMV this requires lengthy incubation, so results cannot be obtained rapidly. Additionally, due to specific technical constraints of the plaque assay including the visual detection format, it has an element of subjectivity along with limited sensitivity. In this study, we describe the development of a FACS-based assay that utilizes detection of LCMV nucleoprotein (NP) expression in infected cells to determine viral titers, and that exhibits several advantages over the standard plaque assay. We show that the LCMV-NP FACS assay is an objective and reproducible detection method that requires smaller sample volumes, exhibits a ∼20-fold increase in sensitivity to and produces results three times faster than the plaque assay. Importantly, when applied to models of acute and chronic LCMV infection, the LCMV-NP FACS assay revealed the presence of infectious virus in samples that were determined to be negative by plaque assay. Therefore, this technique represents an accelerated, enhanced and objective alternative method for detection of infectious LCMV that is amenable to adaptation for other viral infections as well as high throughput diagnostic platforms.

Host response to mechanical ventilation for viral respiratory tract infection

Respiratory syncytial virus (RSV) bronchiolitis causes severe respiratory tract infection in infants, frequently necessitating mechanical ventilatory support. However, life-saving, mechanical ventilation aggravates lung inflammation. We set up a model to dissect the host molecular response to mechanical ventilation in RSV infection. Furthermore, the response to induced hypercapnic acidosis, reported to dampen the inflammatory response to mechanical ventilation in non-infectious models, was assessed. BALB/c mice were inoculated with RSV or mock-suspension and ventilated for 5 h on day 5 post inoculation. Mechanical ventilation of infected mice resulted in enhanced cellular influx and increased concentrations of pro-inflammatory cytokines in the bronchoalveolar space. Microarray analysis showed that enhanced inflammation was associated with a molecular signature of a stress response to mechanical ventilation with little effect on the virus-induced innate immune response. Hypercapnic acidosis during mechanical ventilation of infected mice did not change host transcript profiles. We conclude that mechanical ventilation during RSV infection adds a robust but distinct molecular stress response to virus-induced innate immunity activation, emphasising the importance of lung-protective mechanical ventilation strategies. Induced hypercapnic acidosis has no major effect on host transcription profiles during mechanical ventilation for RSV infection, suggesting that this is a safe approach to minimise ventilator-induced lung injury.

Myxoma Virus Induces Type I Interferon Production in Murine Plasmacytoid Dendritic Cells via a TLR9/MyD88-, IRF5/IRF7-, and IFNAR-Dependent Pathway

Poxviruses are large DNA viruses that replicate in the cytoplasm of infected cells. Myxoma virus is a rabbit poxvirus that belongs to the Leporipoxvirus genus. It causes a lethal disease called myxomatosis in European rabbits but cannot sustain any detectable infection in nonlagomorphs. Vaccinia virus is a prototypal orthopoxvirus that was used as a vaccine to eradicate smallpox. Myxoma virus is nonpathogenic in mice, whereas systemic infection with vaccinia virus can be lethal even in immunocompetent mice. Plasmacytoid dendritic cells (pDCs) are potent type I interferon (IFN)-producing cells that play important roles in antiviral innate immunity. How poxviruses are sensed by pDCs to induce type I IFN production is not well understood. Here we report that infection of primary murine pDCs with myxoma virus, but not with vaccinia virus, induces IFN-α, IFN-β, tumor necrosis factor (TNF), and interleukin-12p70 (IL-12p70) production. Using pDCs derived from genetic knockout mice, we show that the myxoma virus-induced innate immune response requires the endosomal DNA sensor TLR9 and its adaptor MyD88, transcription factors IRF5 and IRF7, and the type I IFN positive-feedback loop mediated by IFNAR1. It is independent of the cytoplasmic RNA sensing pathway mediated by the mitochondrial adaptor molecule MAVS, the TLR3 adaptor TRIF, or the transcription factor IRF3. Using pharmacological inhibitors, we demonstrate that myxoma virus-induced type I IFN and IL-12p70 production in murine pDCs is also dependent on phosphatidylinositol 3-kinase (PI3K) and Akt. Furthermore, our results reveal that the N-terminal Z-DNA/RNA binding domain of vaccinia virulence factor E3, which is missing in the orthologous M029 protein expressed by myxoma virus, plays an inhibitory role in poxvirus sensing and innate cytokine production by murine pDCs.

The hemagglutinin protein of influenza A/Vietnam/1203/2004 (H5N1) contributes to hyperinduction of proinflammatory cytokines in human epithelial cells

Live attenuated influenza A/Vietnam/1203/04 (H5N1) (VN04 cold adapted [ ca]) and A/Hong Kong/213/03 (H5N1) (HK03 ca) vaccine viruses were compared with the A/New Caledonia/20/99 (H1N1) (NC99 ca) seasonal vaccine virus for induction of host gene expression in infected human epithelial cells. Levels of proinflammatory cytokines and interferon-related genes were significantly upregulated in VN04 ca virus-infected A549 cells compared to cells infected with the HK03 ca and NC99 ca viruses as examined by microarray analysis and confirmed by quantitative RT–PCR and ELISA assays. Further mapping studies demonstrated that the hemagglutinin (HA) protein of the VN04 ca virus contributed to the hyperinduction of cytokines. The inactivated viruses could also induce the production of the cytokines and chemokines, albeit at a much lower level than live viruses. Compared to HK03 ca virus, VN04 ca virus differs by 9 amino acids including an additional glycosylation site at residue 158N of the HA protein and a shortened stalk in the neuraminidase (NA) protein. Increased cytokine production by HK03 ca virus was only observed when HK03 ca virus acquired an additional glycosylation in the HA protein and when its NA protein was replaced by that of VN04. Thus, our data indicate that the HA protein and its interaction with the NA protein play a role in triggering cytokine responses. The full implications of cytokine induction in vaccine virus-induced immune responses remain to be explored.

Targeted Delivery of Small Interfering RNA to Human Dendritic Cells To Suppress Dengue Virus Infection and Associated Proinflammatory Cytokine Production

Dengue is a common arthropod-borne flaviviral infection in the tropics, for which there is no vaccine or specific antiviral drug. The infection is often associated with serious complications such as dengue hemorrhagic fever (DHF) or dengue shock syndrome (DSS), in which both viral and host factors have been implicated. RNA interference (RNAi) is a potent antiviral strategy and a potential therapeutic option for dengue if a feasible strategy can be developed for delivery of small interfering RNA (siRNA) to dendritic cells (DCs) and macrophages, the major in vivo targets of the virus and also the source of proinflammatory cytokines. Here we show that a dendritic cell-targeting 12-mer peptide (DC3) fused to nona-D-arginine (9dR) residues (DC3-9dR) delivers siRNA and knocks down endogenous gene expression in heterogenous DC subsets, (monocyte-derived DCs [MDDCs], CD34(+) hematopoietic stem cell [HSC])-derived Langerhans DCs, and peripheral blood DCs). Moreover, DC3-9dR-mediated delivery of siRNA targeting a highly conserved sequence in the dengue virus envelope gene (siFvE(D)) effectively suppressed dengue virus replication in MDDCs and macrophages. In addition, DC-specific delivery of siRNA targeting the acute-phase cytokine tumor necrosis factor alpha (TNF-alpha), which plays a major role in dengue pathogenesis, either alone or in combination with an antiviral siRNA, significantly reduced virus-induced production of the cytokine in MDDCs. Finally to validate the strategy in vivo, we tested the ability of the peptide to target human DCs in the NOD/SCID/IL-2Rgamma(-/-) mouse model engrafted with human CD34(+) hematopoietic stem cells (HuHSC mice). Treatment of mice by intravenous (i.v.) injection of DC3-9dR-complexed siRNA targeting TNF-alpha effectively suppressed poly(I:C)-induced TNF-alpha production by DCs. Thus, DC3-9dR can deliver siRNA to DCs both in vitro and in vivo, and this delivery approach holds promise as a therapeutic strategy to simultaneously suppress virus replication and curb virus-induced detrimental host immune responses in dengue infection.

Essential Impact of NF-κB Signaling on the H5N1 Influenza A Virus-Induced Transcriptome

Systemic infections of humans and birds with highly pathogenic avian influenza A viruses of the H5N1 subtype are characterized by inner bleedings and a massive overproduction of cytokines known as cytokine storm. Growing evidence supports the role of endothelial cells in these processes. The aim of this study was to elucidate determinants of this strong response in endothelial cells with a focus on the transcription factor NF-kappaB. This factor is known as a major regulator of inflammatory response; however, its role in influenza virus replication and virus-induced immune responses is controversially discussed. By global mRNA profiling of infected cells in the presence or absence of a dominant negative mutant of IkappaB kinase 2 that specifically blocks the pathway, we could show that almost all H5N1 virus-induced genes depend on functional NF-kappaB signaling. In particular, activation of NF-kappaB is a bottleneck for the expression of IFN-beta and thus influences the expression of IFN-dependent genes indirectly in the primary innate immune response against H5N1 influenza virus. Control experiments with a low pathogenic influenza strain revealed a much weaker and less NF-kappaB-dependent host cell response.

Adenovirus-mediated down-regulation of X-linked inhibitor of apoptosis protein inhibits colon cancer

Our previous studies and those of others have indicated that X-linked inhibitor of apoptosis protein (XIAP) holds promise as a target gene in colon cancer gene therapy. In this study, we constructed an adenoviral vector to deliver small hairpin RNA (shRNA) against XIAP (XIAP-shRNA) into colon cancer cells and tested its therapeutic efficacy in vitro and in vivo. We first confirmed an overexpression of XIAP in colon cancer cells and human cancer tissues. We then designed XIAP-small interfering RNA (siRNA) and confirmed the knockdown effect of these siRNAs in colon cancer cells. The sequences of the effective siRNAs were converted into shRNA and then packed into replication-deficient adenoviral vectors using BLOCK-iT Adenoviral RNAi Expression System to generate Adv-XIAP-shRNA. Infection of HT29 and HCT116 cells with Adv-XIAP-shRNA led to enhanced caspase-3 activity, which was associated with increased apoptosis and reduced cell proliferation. The therapeutic effect of Adv-XIAP-shRNA was then tested in xenograft tumors in nude mice. We showed that treatment of the xenograft tumors derived from HCT116 cells with Adv-XIAP-shRNA resulted in a retardation of tumor growth, which was associated with enhanced apoptosis, increased caspase-3 activity, and reduced expression of proliferating cell nuclear antigen in the tumor tissues. Treatment of xenograft tumors with Adv-XIAP-shRNA did not affect the expressions of inflammatory cytokines in tumor-bearing mice. Thus, Adv-XIAP-shRNA-mediated down-regulation of XIAP exerts a therapeutic effect in colon cancer by promoting apoptosis and inhibiting proliferation of colon cancer cells, and the antitumor effect of Adv-XIAP-shRNA was unlikely to be related to virus-induced immune response.

Therapeutic Monoclonal Antibody Treatment Targeting Respiratory Syncytial Virus (RSV) G Protein Mediates Viral Clearance and Reduces the Pathogenesis of RSV Infection in BALB/c Mice

Because the G protein of respiratory syncytial virus (RSV) has a CX3C chemokine motif that has been associated with the ability of RSV G protein to modulate the virus-induced host immune response, we examined whether therapeutic treatment with an anti-RSV G monoclonal antibody (mAb), 131-2G, that blocks the CX3C-associated activity of RSV G protein might decrease the pulmonary inflammation associated with infection in BALB/c mice. The results show that treatment with mAb 131-2G on day 3 after RSV infection reduces both inflammation and RSV titer in the lungs. Later administration of anti-RSV G mAb (day 5 after RSV infection) effectively reduced the viral titer but had a minimal effect on pulmonary inflammation. This study suggests that an anti-RSV G mAb might be an effective antiviral, either alone or in combination with anti-RSV F protein neutralizing antibodies, for decreasing the virus-induced host response to infection and improve treatment outcome.

S05: Immune mechanisms in viral bronchiolitis-a model for virus induced asthma exacerbations?

Bronchiolitis in infancy is a severe inflammatory disease of the distal airways, caused by respiratory viruses, most often respiratory syncytial virus (RSV). Respiratory viruses are also the most important triggers for asthma exacerbations. The immune pathology of either condition is not fully understood. The immune response to RSV can be subdivided into: (1) the epithelial phase in the first hours after infection during which epithelial cells and some macrophages release pro-inflammatory cytokines and chemokines, (2) the innate immune response which is characterised by an early influx of natural killer cells and concomitant recruitment of granulocytes, macrophages and dendritic cells (DC), (3) the adaptive immune response by antigen-specific effector T-cells and activated B-cell following activation by DC. T-cells are involved in cytotoxicity, clearing the virus, and in both pro-inflammatory and regulatory pathways. Activated B-cells produce antigen specific antibodies which contribute to viral clearance, in particular in secondary infections. Given the apparent role of T-cells in viral bronchiolitis and viral asthma exacerbations, we studied the impact of RSV infection in vivo on phenotype and function of pulmonary DC, the professional antigen presenting cells that are thought to activate T-cells during infection. Further we investigated the role of airway epithelial cells (AEC) in regulating DC and T-cell activation. Using a mouse model, we observed increases in numbers of lung DC after the initial phase of RSV-infection. These seem to be triggered by an early release of GM-CSF from lung epithelial cells leading to an expansion of local DC-precursors in the lung which mature and become efficient T-cell activators. Early in infection DC preferentially induce Th1- and pro-inflammatory Th17-cells. The latter may contribute to excessive pulmonary inflammation in bronchiolitis and also during subsequent allergen sensitisation possibly facilitating the development of asthma symptoms which have been observed following RSV-bronchiolitis. The subset of plasmacytoid DC which are potent producers of interferon-alpha has a direct antiviral role during RSV-infection limiting maximum replication of RSV and contributing to its clearance. In addition they may have direct regulatory effects, reducing RSV-induced inflammation and lung function changes. Healthy AEC seem to prevent activation of mucosal T cells and DC. However this inhibition is lost after RSV infection of AEC. We conclude that in the early phase of RSV-infection innate, epithelial cell dependent mechanisms dominate the antiviral response, releasing DC and T cells from inhibition, inducing expansion of lung DC and allowing activation of these immune cells. Subsequent innate and adaptive immune responses, including activation of DC and subsequently T-cell subsets contribute to the development and maintenance of pulmonary inflammation and associated lung function changes. Further, these virus-induced immune responses may also contribute to aberrant immune pulmonary responses after the resolution of RSV-bronchiolitis.

Cigarette smoke selectively enhances viral PAMP- and virus-induced pulmonary innate immune and remodeling responses in mice.

Viral infections have more severe consequences in patients who have been exposed to cigarette smoke (CS) than in those not exposed to CS. For example, in chronic obstructive pulmonary disease (COPD), viruses cause more severe disease exacerbation, heightened inflammation, and accelerated loss of lung function compared with other causes of disease exacerbation. Symptomatology and mortality in influenza-infected smokers is also enhanced. To test the hypothesis that these outcomes are caused by CS-induced alterations in innate immunity, we defined the effects of CS on pathogen-associated molecular pattern-induced (PAMP-induced) pulmonary inflammation and remodeling in mice. CS was found to enhance parenchymal and airway inflammation and apoptosis induced by the viral PAMP poly(I:C). CS and poly(I:C) also induced accelerated emphysema and airway fibrosis. The effects of a combination of CS and poly(I:C) were associated with early induction of type I IFN and IL-18, later induction of IL-12/IL-23 p40 and IFN-gamma, and the activation of double-stranded RNA-dependent protein kinase (PKR) and eukaryotic initiation factor-2alpha (eIF2alpha). Further analysis using mice lacking specific proteins indicated a role for TLR3-dependent and -independent pathways as well as a pathway or pathways that are dependent on mitochondrial antiviral signaling protein (MAVS), IL-18Ralpha, IFN-gamma, and PKR. Importantly, CS enhanced the effects of influenza but not other agonists of innate immunity in a similar fashion. These studies demonstrate that CS selectively augments the airway and alveolar inflammatory and remodeling responses induced in the murine lung by viral PAMPs and viruses.

Regulatory T cells hasten infant AIDS

HIV infection tends to develop into AIDS more quickly in infants than in adults. A study by Hartigan-O'Conner et al. (page 2679) now suggests that more potent infant regulatory T (T reg) cells are to blame. Figure 1 Infants (black lines) have more powerful T reg cells that suppress SIV-fighting CD4+ T cells. T reg cells cool down virus-induced immune responses by suppressing activated T cells. It has been suggested that too much suppression allows HIV to get the upper hand. Others, however, have suggested that activated T cell suppression by T reg cells prevents inflammatory T cell cytokines from damaging tissues and thus enhancing disease. Hartigan-O'Conner and colleagues now show that healthy infant monkeys have more—and more potent—T reg cells than do adult monkeys. When monkeys were infected with SIV—the primate version of HIV—most of the infants rapidly developed AIDS. Their T reg cells suppressed anti-SIV T cell functions, whereas the adult T reg cells did not. Humans also have more T reg cells during infancy. The authors suspect that, like primate T reg cells, the potency of human T reg cells might also decrease with age. Determining why T reg cell numbers decrease with age and why aging T reg cells lose their steam is the next step.