Innate Antiviral Genes Research Articles

Bacterial homologs of innate eukaryotic antiviral defenses with anti-phage activity highlight shared evolutionary roots of viral defenses

Prokaryotes have evolved a multitude of defense systems to protect against phage predation. Some of these resemble eukaryotic genes involved in antiviral responses. Here, we set out to systematically project the current knowledge of eukaryotic-like antiviral defense systems onto prokaryotic genomes, using Pseudomonas aeruginosa as a model organism. Searching for phage defense systems related to innate antiviral genes from vertebrates and plants, we uncovered over 450 candidates. We validated six of these phage defense systems, including factors preventing viral attachment, R-loop-acting enzymes, the inflammasome, ubiquitin pathway, and pathogen recognition signaling. Collectively, these defense systems support the concept of deep evolutionary links and shared antiviral mechanisms between prokaryotes and eukaryotes.

Poly(I:C), chitosan oligosaccharide, and chitosan enhance inactivated GCRV vaccine potency in grass carp Ctenopharyngodon idella

Vaccination is the most promising strategy to combat infectious diseases. Vaccine adjuvants are considered as key components in modern vaccinology since they provide the necessary help in enhancing the immune responses. This study evaluated the protective effects of poly (I:C), chitosan oligosaccharides (COS) and chitosan as new adjuvants in enhancing immune response and inhibiting GCRV replication using inactivated GCRV vaccines. Complete Freund's adjuvant (CFA) was used as a control adjuvant, and the survival rate of each group of grass carp was assessed. The CiIgM protein levels in sera were obviously boosted and sustained by the three novel adjuvant supplies, examining by enzyme-linked immunosorbent assay (ELISA) and western blotting (WB) methods. Meanwhile, mRNA expression of CiIgM in spleen and head kidney were elevated, checking by real time quantitative RT-PCR (qRT-PCR) assays. The Th1, not Th2 type immune responses, pro-inflammation cytokines and innate antiviral genes were also reinforced in spleen and head kidney tissues at early stages. The GCRV replication was significantly reduced post GCRV injection. The tissue sectioning showed a significant reduction in tissue damage in muscles, intestines, and hepatopancreatic tissues under the protection of the three adjuvants. The results indicated that poly(I:C), COS and chitosan can serve as novel and efficient adjuvants in inactivated vaccine in grass carp.

Association between Patient-Reported Social Determinant of Health Outcomes and a Social Genomics Profile in Allogeneic Hematopoietic Cell Transplantation: A Center for International Blood and Marrow Transplant Research (CIBMTR) Analysis

Introduction Patient-reported outcomes (PROs) capture the subjective experience of a patient's social and environmental context, providing a firsthand account of this and other social determinants of health (SDOH). An individual's socioenvironmental context is linked to a wide array of health outcomes, including for patients undergoing hematopoietic cell transplantation (HCT) [1, 2]. Social genomics, the study of socioenvironmental effects on human gene expression, offers a framework for understanding these complex biobehavioral relationships [3]. The conserved transcriptional response to adversity (CTRA) is a genomic biomarker characterized by increased expression of pro-inflammatory genes and decreased innate anti-viral gene expression [4]. The CTRA profile has been associated with psychosocial adversity and is predictive of clinical outcomes in HCT, and thus may be a means to map the biologic sequelae of social hardship [2, 4]. The objective of this study was to examine the association of pre-transplant CTRA profile with psychosocial PROs in allogeneic HCT recipients. Methods A cross-sectional analysis of data from a subset of participants enrolled on a prior CIBMTR study testing the feasibility of centralized collection of PROs in allogeneic HCT [5] was completed. For the present analysis, eligible patients from this larger cohort were those aged >18 years with baseline (pre-HCT) PRO measures and available pre-transplant banked biospecimens. PROs included the 36-Item Short Form Health Survey (SF-36) and the Functional Assessment of Cancer Therapy-Bone Marrow Transplant (FACT-BMT). Descriptive analyses of patient-, disease-, and transplant-related characteristics were prepared. CTRA was derived from banked whole blood specimens using RNA sequencing. Transcript-per-million values of the 47 indicator genes were log2 transformed and analyzed using mixed effects linear regression models testing relative average gene expression levels to standardized PRO score controlling for age, sex, race, malignant versus non-malignant disease, and performance status. Results A total of n=140 patients representing 8 transplant centers in the United States were included. Patient characteristics are depicted in Table 1. The median age was 55 years (range 18-75y), 44% were female, and 90% identified as White. A majority (96%) of patients had a hematologic malignancy diagnosis, with acute myelogenous leukemia (AML) being most common (35%). There were statistically significant associations between lower CTRA gene expression and higher FACT-BMT total scores (β-coefficient -0.041 difference in log2 CTRA RNA abundance/SD PRO score, p-value=0.003) social well-being (β-coefficient -0.058 difference in log2 CTRA RNA abundance/SD PRO score, p-value=0.0001), physical well-being, (β-coefficient -0.031 difference in log2 CTRA RNA abundance/SD PRO score, p-value=0.031) and FACT-G scores (β-coefficient -0.042 difference in log2 CTRA RNA abundance/SD PRO score, p-value=0.003) as depicted in Figure 1. This corresponds to a 10.7%, 14.8%, 8.2%, and 11.1% difference, respectively, in CTRA gene expression per 4-SD range of variation in each PRO. Conclusions Patient reports of socioenvironmental experience were associated with the CTRA immunoregulatory pattern in adult allogeneic HCT recipients, with social well-being emerging as the most significant domain. This finding is consistent with prior literature and supports the importance of social well-being and community context in the setting of allogeneic HCT. Heightened CTRA expression could provide a potential molecular mechanism for social gradients in HCT outcomes, although this was not experimentally tested in this study. Further research and intervention that targets this pathway could support optimal multidimensional outcomes in this high-risk population. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Enhanced Immune Activation Following Acute Social Stress Among Adolescents With Early-Life Adversity

BackgroundEarly-life adversity (ELA) has been linked to higher depression risk across the life span and chronic inflammatory conditions that contribute to earlier mortality. In this study, we characterized innate immune responses to acute social stress in a community sample of adolescents (mean age = 13.9 ± 1.6 years; 46.4% female) as a potential pathway linking ELA and depression pathogenesis. MethodsParents reported their child’s exposure to 9 ELAs, and adolescents participated in the Trier Social Stress Test for Children, with blood collected immediately before and then at 60 and 90 minutes thereafter. Overall, 65 adolescents had complete data for analysis of stress-induced changes in gene expression and 84 adolescents had complete data for circulating inflammatory markers. ResultsRelative to adolescents exposed to no ELA (11.9%) or low ELA (ELA = 1−3; 67.9%), those exposed to high ELA (ELA = 4+; 20.2%) showed larger stress-associated increases in expression of both proinflammatory and innate antiviral gene transcripts in circulating blood. Consistent with a potential mediating role of sympathetic nervous system activity, promoter-based bioinformatics analyses implicated CREB transcription factor activity in structuring observed gene expression differences. These effects were accompanied by a smaller initial but protracted increase in circulating interleukin 6 in adolescents with high ELA. ConclusionsResults are consistent with the hypothesis that ELA may enhance cellular and gene regulatory reactivity to stress, which may, in turn, increase vulnerability to depression and other inflammation-related disease processes.

Flourishing in Healthcare Trainees: Psychological Well-Being and the Conserved Transcriptional Response to Adversity.

While much attention has been paid to healthcare provider and trainee burnout, less is known about provider well-being (i.e., flourishing) or about the effects of well-being on immune function. This study examined the demographic and psycho-social correlates of well-being among healthcare trainees (resident physicians and physician assistant (PA) trainees) and evaluated the association of well-being with the “conserved transcriptional response to adversity” (CTRA) characterized by up-regulated expression of pro-inflammatory genes and down-regulated expression of innate antiviral genes. Participants (n = 58) completed self-reported assessments of sleep disturbance, loneliness, depressive symptoms, anxiety, stress, and well-being (flourishing). Blood sample RNA profiles were analyzed by RNA sequencing to assess the CTRA. Slightly over half (n = 32; 55.2%) of healthcare trainees were categorized as flourishing. Flourishing was less prevalent among primary caregivers, and more prevalent among trainees who exercised more frequently and those with fewest days sick. Loneliness (AOR = 0.75; 95% CI = 0.61, 0.91; p = 0.003) and stress (AOR = 0.65; 95% CI = 0.45, 0.94; p = 0.02) were associated with decreased odds of flourishing when controlling for other variables. Flourishing was associated with down-regulated CTRA gene expression, whereas loneliness was associated with up-regulated CTRA gene expression (both p < 0.05). Assessing these relationships in a larger, multi-site study is of critical importance to inform policy, curricula, and interventions to bolster sustainable trainee well-being.

Myeloid cell interferon responses correlate with clearance of SARS-CoV-2

Emergence of mutant SARS-CoV-2 strains associated with an increased risk of COVID-19-related death necessitates better understanding of the early viral dynamics, host responses and immunopathology. Single cell RNAseq (scRNAseq) allows for the study of individual cells, uncovering heterogeneous and variable responses to environment, infection and inflammation. While studies have reported immune profiling using scRNAseq in terminal human COVID-19 patients, performing longitudinal immune cell dynamics in humans is challenging. Macaques are a suitable model of SARS-CoV-2 infection. Our longitudinal scRNAseq of bronchoalveolar lavage (BAL) cell suspensions from young rhesus macaques infected with SARS-CoV-2 (n = 6) demonstrates dynamic changes in transcriptional landscape 3 days post- SARS-CoV-2-infection (3dpi; peak viremia), relative to 14-17dpi (recovery phase) and pre-infection (baseline) showing accumulation of distinct populations of both macrophages and T-lymphocytes expressing strong interferon-driven inflammatory gene signature at 3dpi. Type I interferon response is induced in the plasmacytoid dendritic cells with appearance of a distinct HLADR+CD68+CD163+SIGLEC1+ macrophage population exhibiting higher angiotensin-converting enzyme 2 (ACE2) expression. These macrophages are significantly enriched in the lungs of macaques at 3dpi and harbor SARS-CoV-2 while expressing a strong interferon-driven innate anti-viral gene signature. The accumulation of these responses correlated with decline in viremia and recovery.

Immunostimulant Bathing Influences the Expression of Immune- and Metabolic-Related Genes in Atlantic Salmon Alevins.

Simple SummaryActivation of immune cells is bioenergetically expensive, requiring precise control of cellular metabolism. This applies also to innate immune cells. The current study shows that the immunostimulants, Astragalus, Hyaluronic acid, Imiquimod, and Poly I:C can modulate the expression of genes involved in the innate antiviral immune, as well as genes associated with metabolism, in the early life stages of Atlantic salmon.Disease resistance of fish larvae may be improved by bath treatment in water containing immunostimulants. Pattern recognition receptors, such as TLR3, TLR7, and MDA5, work as an “early warning” to induce intracellular signaling and facilitate an antiviral response. A single bath of newly hatched larvae, with Astragalus, upregulated the expression of IFNα, IFNc, ISG15, MDA5, PKR, STAT1, TLR3, and TLR7 immune genes, on day 4 post treatment. Similar patterns were observed for Hyaluronic acid and Poly I:C. Increased expression was observed for ISG15, MDA5, MX, STAT1, TLR3, TLR7, and RSAD2, on day 9 for Imiquimod. Metabolic gene expression was stimulated on day 1 after immunostimulant bath in ULK1, MYC, SLC2A1, HIF1A, MTOR, and SIX1, in Astragalus, Hyaluronic acid, and Imiquimod. Expression of NOS2 in Poly I:C was an average fourfold above that of control at the same timepoint. Throughout the remaining sampling days (2, 4, 9, 16, 32, and 45 days post immunostimulant bath), NOS2 and IL1B were consistently overexpressed. In conclusion, the immunostimulants induced antiviral gene responses, indicating that a single bath at an early life stage could enable a more robust antiviral defense in fish. Additionally, it was demonstrated, based on gene expression data, that cell metabolism was perturbed, where several metabolic genes were co-regulated with innate antiviral genes.

Baseline pro-inflammatory gene expression in whole blood is related to adverse long-term outcomes after transcatheter aortic valve replacement: a case control study

BackgroundAge-associated inflammation and immune system dysfunction have been implicated as mechanisms that increase risk for adverse long-term procedural outcomes in older adults. The purpose of this study was to investigate relationships between baseline inflammatory and innate antiviral gene expression and outcomes after transcatheter aortic valve replacement (TAVR) in older adults with severe aortic stenosis.MethodsWe performed a retrospective case–control study comparing pre-procedural pro-inflammatory and Type 1 interferon (IFN) gene expression in 48 controls with favorable outcomes (alive 1 year after TAVR with improved quality of life [QoL]) versus 48 individuals with unfavorable outcomes (dead by 1 year or alive at 1 year but with reduced QoL). Gene expression was evaluated in whole blood via (1) pre-defined composite scores of 19 inflammation-associated genes and 34 Type I IFN response genes, and (2) pro-inflammatory and antiviral transcription factor activity inferred from promotor based bioinformatics analyses of genes showing > 25% difference in average expression levels across groups. All analyses were adjusted for age, gender, body mass index, diabetes, immunosuppression, cardiovascular disease (CVD), and frailty.ResultsRelative to controls, those with unfavorable outcomes demonstrated higher expression of the pro-inflammatory gene composite prior to TAVR (p < 0.01) and bioinformatic indicators of elevated Nuclear Factor kB (p < 0.001) and Activator Protein 1 (p < 0.001) transcription factor activity, but no significant differences in Type I IFN-related gene expression.ConclusionsThese results demonstrate that a pro-inflammatory state prior to TAVR, independent of CVD severity and frailty status, is associated with worse long-term procedural outcomes.

Overexpression of Toll-like receptor 4 enhances LPS-induced inflammatory response and inhibits Salmonella Typhimurium growth in ovine macrophages

The Toll-like receptor 4 (TLR4) plays a crucial role in innate inflammatory responses, as it recognizes gram-negative bacteria (or their products) and contributes greatly to host defense against invading pathogens. Though TLR4 overexpressing transgenic sheep, resistant to certain diseases related with gram-negative bacteria, had been bred in our previous research, the effects of overexpression of TLR4 on innate immune response remained unclear. In this study, TLR4 overexpressing ovine macrophages were obtained from peripheral blood, and it was found that the overexpression of TLR4 initially promoted the production of proinflammatory cytokines TNFα and IL-6 by activating TLR4-mediated IRAK4-dependent NF-κB and MAPK (JNK and ERK1/2) signaling following LPS stimulation. However, this effect was later impaired due to increased internalization of TLR4 into endosomal compartment of the macrophages. Then the overexpression of TLR4 triggered TBK1-dependent interferon-regulatory factor-3 (IRF-3) expression, which in turn led to the induction of IFN-β and IFN-inducible genes (i.e.IP10, IRG1 and GARG16). Understandably, an increased IFN-β level facilitated phosphorylation of STAT1 to induce expression of innate antiviral genes Mx1 and ISG15, suggesting that TLR4 overexpressing macrophages were equipped better against viral infection. Correspondingly, the bacterial burden in these macrophages, after infection with live S. Typhimurium, was decreased significantly. In summary, the results indicated that overexpression of TLR4 could enhance innate inflammatory responses, initiate the innate antiviral immunity, and control effectively S. Typhimurium growth in ovine macrophages.

Conditional reprogramming of pediatric airway epithelial cells: A new human model to investigate early-life respiratory disorders.

Airway epithelial cells (AEC) are quite difficult to access in newborns and infants. It is critically important to develop robust life-extended models to conduct translational studies in this age group. We propose the use of a recently described cell culture technology (conditionally reprogrammed cells-CRC) to generate continuous primary cell cultures from nasal and bronchial AEC of young children. We collected nasal and/or bronchial AEC from a total of 23 subjects of different ages including newborns/infants/toddlers (0-2years; N=9), school-age children (4-11years; N=6), and a group of adolescent/adult donors (N=8). For CRC generation, we used conditioned medium from mitotically inactivated 3T3 fibroblasts and Rho-associated kinase (ROCK) inhibitor (Y-27632). Antiviral immune responses were studied using 25 key antiviral genes and protein production of type III epithelial interferon (IFN λ1) after double-stranded (ds) RNA exposure. CRC derived from primary AEC of neonates/infants and young children exhibited: (i) augmented proliferative capacity and life extension, (ii) preserved airway epithelial phenotype after multiple passages, (iii) robust immune responses characterized by the expression of innate antiviral genes and parallel nasal/bronchial production of IFN λ1 after exposure to dsRNA, and (iv) induction of airway epithelial inflammatory and remodeling responses to dsRNA (eg, CXCL8 and MMP9). Conditional reprogramming of AEC from young children is a feasible and powerful translational approach to investigate early-life airway epithelial immune responses in humans.

IFN-γ Influences Epithelial Antiviral Responses via Histone Methylation of the RIG-I Promoter.

The asthmatic lung is prone to respiratory viral infections that exacerbate the symptoms of the underlying disease. Recent work has suggested that a deficient T-helper cell type 1 response in early life may lead to these aberrant antiviral responses. To study the development of long-term dysregulation of innate responses, which is a hallmark of asthma, we investigated whether the inflammatory environment of the airway epithelium can modulate antiviral gene expression via epigenetic mechanisms. We primed AALEB cells, a human bronchial epithelial cell line, with IFN-γ and IL-13, and subsequently infected the cells with respiratory syncytial virus (RSV). We then analyzed the expression of innate antiviral genes and their epigenetic markers. Priming epithelial cells with IFN-γ reduced the RSV viral load. Microarray analysis identified that IFN-γ priming enhanced retinoic acid-inducible gene (RIG)-I mRNA expression, and this expression correlated with epigenetic changes at the RIG-I promoter that influenced its transcription. Using chromatin immunoprecipitation, we observed a reduction of trimethylated histone 3 lysine 9 at the RIG-I promoter. Addition of inhibitor BIX-01294 to this model indicated an involvement of lysine methyltransferase G9a in RIG-I epigenetic regulation. These data suggest that prior exposure to IFN-γ may leave an epigenetic mark on the chromatin that enhances airway cells' ability to resist infection, possibly via epigenetic upregulation of RIG-I. These observations provide further evidence for a crucial role of IFN-γ in the development of mature antiviral responses within a model of respiratory infection. Further clinical validation is required to determine whether this effect in early life leads to changes in antiviral responses associated with asthma.

Innate Immune Gene Transcript Level Associated with the Infection of Macrophages with Ectromelia Virus in Two Different Mouse Strains

Poxviruses have evolved numerous mechanisms to avoid the immune response of the infected host, and many of these mechanisms have not been fully described. Here, we studied the transcriptional response of innate immune genes in BALB/c and C57BL/6 peritoneal macrophages following infection with the Moscow strain of ectromelia virus (ECTV-Mos) with the aim of delineating innate immune genes that contribute to the difference between susceptibility and resistance to lethal infection. We show a generalized downregulation of many genes in four categories (toll-like receptor signaling, NOD-like receptor signaling, RIG-I-like receptor signaling, and type I interferon signaling) of antiviral innate immune receptors, downstream signaling pathways, and responsive components. Two important observations were made. First, 14 innate antiviral genes were differentially expressed with fold change upregulation of two and above occurring in C57BL/6 mice, known to be resistant to ECTV-Mos infection, whereas the same genes were downregulated in BALB/c mice with fold change of two and below. Second, the cathepsin group of genes was downregulated in both strains of mice but with profound fold changes of 17, 38, and 62 downregulation for CtsL, CtsB, and CtsS, respectively, in C57BL/6 mice. We show that a poxvirus profoundly downregulates both the mRNA and protein expression of these three cathepsins and this change appears to support virus replication. Based on these data we propose that the variations in gene expression observed may contribute to the difference in resistance/susceptibility between BALB/c and C57BL/6 mice to lethal infection by ECTV-Mos.

Prolonged activation of innate antiviral gene signature after childbirth is determined by IFNL3 genotype

Maternal innate and adaptive immune responses are modulated during pregnancy to concurrently defend against infection and tolerate the semiallogeneic fetus. The restoration of these systems after childbirth is poorly understood. We reasoned that enhanced innate immune activation may extend beyond gestation while adaptive immunity recovers. To test this hypothesis, the transcriptional profiles of total peripheral blood mononuclear cells following delivery in healthy women were compared with those of nonpregnant control subjects. Interestingly, interferon-stimulated genes (ISGs) encoding proteins such as IFIT1, IFIT2, and IFIT3, as well as signaling proteins such as STAT1, STAT2, and MAVS, were enriched postpartum. Antiviral genes were primarily expressed in CD14(+) cells and could be stratified according to genetic variation at the interferon-λ3 gene (IFNL3, also named IL28B) SNP rs12979860. Antiviral gene expression was sustained beyond 6 mo following delivery in mothers with a CT or TT genotype, but resembled baseline nonpregnant control levels following delivery in mothers with a CC genotype. CT and TT IFNL3 genotypes have been associated with persistent elevated ISG expression in individuals chronically infected with hepatitis C virus. Together, these data suggest that postpartum, the normalization of the physiological rheostat controlling IFN signaling depends on IFNL3 genotype.

Impaired innate, humoral, and cellular immunity despite a take in smallpox vaccine recipients

Smallpox vaccine is highly effective, inducing protective immunity to smallpox and diseases caused by related orthopoxviruses. Smallpox vaccine efficacy was historically defined by the appearance of a lesion or “take” at the vaccine site, which leaves behind a characteristic scar. Both the take and scar are readily recognizable and were used during the eradication effort to indicate successful vaccination and to categorize individuals as “protected.” However, the development of a typical vaccine take may not equate to the successful development of a robust, protective immune response. In this report, we examined two large (>1000) cohorts of recipients of either Dryvax® or ACAM2000 using a testing and replication study design and identified subgroups of individuals who had documented vaccine takes, but who failed to develop robust neutralizing antibody titers. Examination of these individuals revealed that they had suboptimal cellular immune responses as well. Further testing indicated these low responders had a diminished innate antiviral gene expression pattern (IFNA1, CXCL10, CXCL11, OASL) upon in vitro stimulation with vaccinia virus, perhaps indicative of a dysregulated innate response. Our results suggest that poor activation of innate antiviral pathways may result in suboptimal immune responses to the smallpox vaccine. These genes and pathways may serve as suitable targets for adjuvants in new attenuated smallpox vaccines and/or effective antiviral therapy targets against poxvirus infections.

Genome wide host gene expression analysis in chicken lungs infected with avian influenza viruses

Background: The molecular pathogenesis of avian influenza viruses vary greatly with individual bird species and virus strain. The molecular pathogenesis of the highly pathogenic avian influenza virus (HPAIV) or the low pathogenic avian influenza virus (LPAIV) in avian species remains poorly understood. Methods & Materials: Thus, global immune response of chickens infected with HPAI H5N1 (A/duck/India/02CA10/2011) and LPAI H9N2 (A/duck/India/249800/2010) viruses was studied using microarray to identify crucial host genetic components responsive to these infection. Results: HPAI H5N1 virus induced excessive mRNA expression of type I IFNs (IFNA and IFNG), cytokines (IL1B, IL18, IL22, IL13, and IL12B), chemokines (CCL4, CCL19, CCL10, and CX3CL1) and IFN stimulated genes (OASL, MX1, RSAD2, IFITM5, IFIT5, GBP 1, and EIF2AK) in lung tissues. This dysregulation of host innate antiviral genes may be the critical determinant of the severity and the outcome of the influenza infection in chickens. In contrast, the expression levels of most of these genes remained unchanged in the lungs of LPAI H9N2 virus infected chickens. Conclusion: This study indicated the relationship between host antiviral genes and their roles in pathogenesis of HPAIV infection in chickens.

Glucocorticosteroids enhance replication of respiratory viruses: effect of adjuvant interferon.

Glucocorticosteroids (GCS) are used on a daily basis to reduce airway inflammation in asthma and chronic obstructive pulmonary disease (COPD). This treatment is usually escalated during acute disease exacerbations, events often associated with virus infections. We examined the impact of GCS on anti-viral defences and virus replication and assessed supplementary interferon (IFN) treatment. Here, we report that treatment of primary human airway cells in vitro with GCS prior to rhinovirus (RV) or influenza A virus (IAV) infection significantly reduces the expression of innate anti-viral genes and increases viral replication. Mice given intranasal treatment with GCS prior to IAV infection developed more severe disease associated with amplified virus replication and elevated inflammation in the airways. Adjuvant IFN treatment markedly reduced GCS-amplified infections in human airway cells and in mouse lung. This study demonstrates that GCS cause an extrinsic compromise in anti-viral defences, enhancing respiratory virus infections and provides a rationale for adjuvant IFN treatment.

S129 Inflammatory Cytokines Influence Respiratory Epithelial Anti-viral Immune Responses Via Inducible Epigenetic Control Of Rig1 Expression: A Model Of Early Life Origins Of Asthma?

The development of asthma is linked to early life environmental exposures and the occurrence of severe viral infections. Rapid maturation of adaptive immunity from a tolerant (Th2) to an anti-infective (Th1) state occurs in the neonatal period. We hypothesised that the airway inflammatory milieu, driven by the maturing immune response to environmental exposures may have important effects on the development of anti-viral innate immunity at the level of the epithelium. We studied whether the inflammatory environment of the airway epithelium modulates gene expression via epigenetic regulation of anti-viral genes as a model of the development of a long term abnormality which is a hallmark of asthma. We optimised an in-vitro model using AALEB, human immortalised bronchial epithelial-derived cells which were pre-treated for 24 h with cytokines that mimic Th1 environment (IFNy, 10 ng/ml) and Th2 (IL-13, 10 ng/ml) before being infected with RSV A, MOI=2 for 48 h. Quantitative real-time PCR with Taqman primers was used to assess expression of innate genes. Cells were collected after 48 h and stored in Trizol. Chromatin Immuno Precipitation (ChIP) with antibodies against histone modifications was used to assess epigenetic controls. In order to confirm epigenetic regulation of innate genes we used a panel of HAT, HDAC and histone demethylase inhibitors. We initially studied the impact of cytokines on a range of innate anti-viral genes. RIG1 was differentially expressed and reductions in expression associated with higher viral titres. IFNy priming induced increases in RIG1 mRNA at 48 h that correlated at the promoter with enrichment of H3K9ac and RNApolII (active-promoters) and reduction of H3K9me3 (repressive-promoters). We observed a statistically significant increase of RIG1 expression by IFNγ when co-incubated with SAHA (HDAC I and II inhibitors) and JIB-04 (Pan-Jumanji histone demethylase inhibitor). No effects of Th2 priming were seen at the level of antiviral responses. This in-vitro study suggests the inflammatory environment of naive epithelial cells can induce epigenetic modulation of innate immune responses at the level of histone methylation and acetylation and hence potentially lead to long term impacts on anti-viral immunity. The presence of a Th1 milieu appears key to the development of effective anti-viral responses.

Enhanced Innate Antiviral Gene Expression, IFN-α, and Cytolytic Responses Are Predictive of Mucosal Immune Recovery during Simian Immunodeficiency Virus Infection

The mucosa that lines the respiratory and gastrointestinal (GI) tracts is an important portal of entry for pathogens and provides the first line of innate immune defense against infections. Although an abundance of memory CD4(+) T cells at mucosal sites render them highly susceptible to HIV infection, the gut and not the lung experiences severe and sustained CD4(+) T cell depletion and tissue disruption. We hypothesized that distinct immune responses in the lung and gut during the primary and chronic stages of viral infection contribute to these differences. Using the SIV model of AIDS, we performed a comparative analysis of the molecular and cellular characteristics of host responses in the gut and lung. Our findings showed that both mucosal compartments harbor similar percentages of memory CD4(+) T cells and displayed comparable cytokine (IL-2, IFN-γ, and TNF-α) responses to mitogenic stimulations prior to infection. However, despite similar viral replication and CD4(+) T cell depletion during primary SIV infection, CD4(+) T cell restoration kinetics in the lung and gut diverged during acute viral infection. The CD4(+) T cells rebounded or were preserved in the lung mucosa during chronic viral infection, which correlated with heightened induction of type I IFN signaling molecules and innate viral restriction factors. In contrast, the lack of CD4(+) T cell restoration in the gut was associated with dampened immune responses and diminished expression of viral restriction factors. Thus, unique immune mechanisms contribute to the differential response and protection of pulmonary versus GI mucosa and can be leveraged to enhance mucosal recovery.

Influenza A virus infection in zebrafish recapitulates mammalian infection and sensitivity to anti-influenza drug treatment.

Seasonal influenza virus infections cause annual epidemics and sporadic pandemics. These present a global health concern, resulting in substantial morbidity, mortality and economic burdens. Prevention and treatment of influenza illness is difficult due to the high mutation rate of the virus, the emergence of new virus strains and increasing antiviral resistance. Animal models of influenza infection are crucial to our gaining a better understanding of the pathogenesis of and host response to influenza infection, and for screening antiviral compounds. However, the current animal models used for influenza research are not amenable to visualization of host-pathogen interactions or high-throughput drug screening. The zebrafish is widely recognized as a valuable model system for infectious disease research and therapeutic drug testing. Here, we describe a zebrafish model for human influenza A virus (IAV) infection and show that zebrafish embryos are susceptible to challenge with both influenza A strains APR8 and X-31 (Aichi). Influenza-infected zebrafish show an increase in viral burden and mortality over time. The expression of innate antiviral genes, the gross pathology and the histopathology in infected zebrafish recapitulate clinical symptoms of influenza infections in humans. This is the first time that zebrafish embryos have been infected with a fluorescent IAV in order to visualize infection in a live vertebrate host, revealing a pattern of vascular endothelial infection. Treatment of infected zebrafish with a known anti-influenza compound, Zanamivir, reduced mortality and the expression of a fluorescent viral gene product, demonstrating the validity of this model to screen for potential antiviral drugs. The zebrafish model system has provided invaluable insights into host-pathogen interactions for a range of infectious diseases. Here, we demonstrate a novel use of this species for IAV research. This model has great potential to advance our understanding of influenza infection and the associated host innate immune response.

146. Does gene regulation make it complicated? Differences in gene expression responses to Complicated Grief and Non-Complicated Grief

Increased morbidity and mortality in recently bereaved spouses (e.g. “widowhood effect”) may be due to alterations in immune function. However, little is known about the impact of bereavement on gene regulation of immune cells. Complicated Grief (CG), a syndrome characterized by persistent separation distress with feelings of overwhelming yearning and preoccupation with the deceased is distinct to Non-complicated Grief (Non-CG) in various psychoneuroimmunologic aspects. In this study we examined whether the type of grief (CG vs. Non-CG) differentially affects leukocyte gene expression. Sixty-three older adults were recruited for genome-wide transcriptional profiling and bioinformatic analyses; 36 had lost their spouse/partner, with 12 of them meeting criteria for CG. The remaining 27 were nonbereaved married controls. Bereavement (both CG and Non-CG, compared to nonbereaved controls) was linked to an increased expression of genes involved in general immunologic activation and a decreased expression of genes involved in B lymphocyte responses. In contrast, CG and non-CG showed significantly different gene expression of Type I interferon-related transcripts: compared to nonbereaved controls there was substantial upregulation in Non-CG subjects and substantial downregulation in CG subjects. Bereavement significantly modulates immune function gene expression. Furthermore, innate antiviral genes are differentially expressed in response to the magnitude of bereavement-related distress (i.e., CG vs. non-CG). These molecular insights might help to understand the health effects of bereavement.