IFN Expression Research Articles

HSP90 Complex From OLP Lesion Induces T-Cell Polarization via Activation of Dendritic Cells.

To investigate the effects of the heat shock protein 90 (HSP90) complex from oral lichen planus (OLP) lesion tissues on dendritic cell (DC) activation and the polarization of naïve T cells. Expression of HSP90 in OLP lesions and healthy control (HC) mucosa was evaluated by single-cell RNA sequence, IHC, qRT-PCR, and immunoblotting. HSP90 complex was extracted by immunoprecipitation from oral mucosa as the agonist of DCs. Expression of IFN-α and MHC was detected by flow cytometry. After cocultured with pre-stimulated DCs, polarization of naïve T cells was investigated by cytokine analysis. HSP90 was significantly higher in the lamina propria of OLP lesion and closely related to lymphocyte infiltration. HSP90 complex of OLP lesion activated DCs via TLR9 and increased their IFN-α secretion and MHC II expression. Pre-stimulated DCs increased the proportion of Th17 cells. HSP90 complex isolated from OLP lesion activated TLR9/IFN-α of DCs and further promoted the polarization of naïve T cells toward Th17 immunity.

Nucleos(t)ide analogues potentially activate T lymphocytes through inducing interferon expression in hepatic cells and patients with chronic hepatitis B

Chronic hepatitis B (CHB) leads to liver inflammation and dysfunction, resulting in liver fibrosis and cancer. Nucleos(t)ide analogues (NAs), inhibitors of hepatitis B virus (HBV), specifically suppress HBV replication. We proposed that immune modulation benefits seroconversion by HBsAg loss. However, activation of T lymphocytes also deteriorates hepatic inflammation. Therefore, we intended to investigate the T cell status and its relationship with hepatic functions in CHB patients treated with NAs. Serum markers, including liver function markers AST, ALT, and HBV-infected markers HBV DNA, HBsAg, HBeAg, and HBsAb were measured in the clinical routine. The T cell levels and markers, including CD69, CD107a, CXCR3, and PD-1 were investigated using flow cytometry. Meanwhile, IFNγ, IL-2, and CXCL10 as immune activation markers in the PBMCs were investigated using qPCR. To validate the effects of NAs on T cell status, qPCR and flow cytometry were used to investigate the gene expression in the HepG2 and PLC5 cells treated with NAs, and in the healthy PBMCs treated with the cell-cultured supernatant. We found that NAs significantly suppressed HBV DNA and reduced AST and ALT levels in the CHB patients. Meanwhile, AST and ALT were both positively correlated with activation marker CD107a in CD8+ T cells. In addition, we found that the CHB patients with seroconversion exhibited a higher CD4/CD8 ratio (p < 0.05) compared to non-seroconversion. We demonstrated that NAs potentially induced IFNs and PD-L1 expression in HepG2 and PLC5 cells. Moreover, the collected supernatant from NAs-treated HepG2 significantly activated PBMCs. This study revealed that the reduction of HBV by NAs may be the reason leading to less AST and ALT levels. We further demonstrated that NAs induced IFN expression in hepatic cells to potentially activate T lymphocytes, which was positively associated with AST and ALT levels in the CHB patients. The results may explain the phenomena in clinical that when the virus is reactivated by aborted use of NAs, it causes consequent T cells-mediated severe acute-on-chronic liver injury.

Normalized Interferon Signatures and Clinical Improvements by IFNAR1 Blocking Antibody (Anifrolumab) in Patients with Type I Interferonopathies

PurposeA causal role of type-I interferons (IFN-I) in autoinflammatory type-I interferonopathies such as SAVI (STING–associated vasculopathy with onset in infancy) and CANDLE (chronic atypical neutrophilic dermatosis with lipodystrophy and elevated temperatures) is suggested by elevated expression of IFN-I stimulated genes (ISGs). Hitherto, the lack of specific inhibitors of IFN-I signaling has prevented the verification of a causal role for IFN-I in these conditions. Commonly used inhibitors of the JAK/STAT pathway exert broad effects on multiple signaling pathways leading to more general immunosuppression beyond IFN-I signaling.MethodsHere we show in four patients with SAVI and one patient with CANDLE syndrome that blockade of the IFNAR1 receptor (Anifrolumab) exerts an additive effect over JAK-inhibitor alone. In two patients with SAVI, monotherapy with Anifrolumab is sufficient to retain a suppressed IFN-I signature and clinical improvement.ResultsAnifrolumab normalizes IFN-I signature genes and relieves symptoms beyond what is typically achieved by a JAK-inhibitor (Baricitinib) alone in patients with type-I interferonopathies. In two patients Anifrolumab was used successfully as monotherapy. Addition of Anifrolumab enabled steroid tapering and cessation with reduced overall immunosuppression and lower risks of opportunistic infections and improved metabolic states and growth which is highly beneficial in these young patients.ConclusionThese results verify a causal role of IFN-I signaling in type-I Interferonopathies SAVI and CANDLE and suggests Anifrolumab as an important new treatment option in autoinflammatory diseases with elevated IFN-I induced gene expression.Genia Kretzschmar, Laura Piñero Páez, and Ziyang Tan are shared-first authors.Sara Alehashemi, AnnaCarin Horne, and Petter Brodin are co-senior author.

DAMPs Drive Fibroinflammatory Changes in the Glaucomatous ONH.

The optic nerve head (ONH) is well known to be the initial site of glaucomatous damage; however, the molecular mechanisms initiating this pathology are not fully understood. To further understand the initiating factors in glaucomatous damage we utilized a novel mouse model of glaucoma, B6.EDA+/+ mice, which constitutively express fibronectin containing the extra domain A (FN+EDA). FN+EDA is a known damage-associated molecular pattern (DAMP) that activates Toll-like receptor 4 and elicits a fibro-inflammatory response. Eyes from B6.EDA+/+ and C57BL/6J mice were evaluated for retinal ganglion cell (RGC) death, retinal nerve fiber layer (RNFL) thickness, and optic nerve (ON) damage at 12 months and 22 months of age. ONH sections were isolated using laser capture microdissection for subsequent RNA-sequencing and Gene Set Enrichment Analysis (GSEA). GSEA results were confirmed using immunohistochemical (IHC) staining. B6.EDA+/+ mice exhibit significantly higher intraocular pressure, loss of RGCs, thinning of the RNFL, and progressive levels of ON damage at 12 months and 22 months of age compared to C57BL/6J controls. Protein expression of DAMPs FN+EDA and biglycan was significantly increased in B6.EDA+/+ mice compared to C57BL/6J controls. GSEA analysis identified significantly up- and downregulated gene groupings at both 12 months and 22 months of age, and IHC staining at 12 and 18 months of age demonstrated significant increases of IFNα, IFNβ, and pSTAT1 expression in B6.EDA+/+ mice compared to C57BL/6J controls. Our study characterizes glaucomatous changes to the retina, ON, and ONH over the course of 2 years and identifies novel molecular pathways associated with these pathophysiological changes. These data illustrate the effects of FN+EDA on the fibro-inflammatory response in the aging ONH in a novel mouse model of glaucoma.

The Type 1 Diabetes-Associated Single Nucleotide Polymorphism rs1990760 in IFIH1 Is Associated with Increased Basal Type I IFNs and IFN-stimulated Gene Expression.

Type 1 diabetes (T1D) is a chronic autoimmune disease that is caused by a combination of genetic and environmental risk factors. In this study, we sought to determine whether a known genetic risk factor, the rs1990760 single nucleotide polymorphism (SNP) (A946T) in IFIH1, resulted in a gain of function in the MDA5 protein and the effects of this mutation on the regulation of type I IFNs during infection with the diabetogenic virus coxsackievirus B3. We found that in cell lines overexpressing the risk variant IFIH1946T there was an elevated level of basal type I IFN signaling and increased basal IFN-stimulated gene expression. An investigation into the mechanism demonstrated that recombinant MDA5 with the A946T mutation had increased ATPase activity invitro. We also assessed the effect of this SNP in primary human PBMCs from healthy donors to determine whether this SNP influenced their response to infection with coxsackievirus B3. However, we observed no significant changes in type I IFN expression or downstream induction of IFN-stimulated genes in PBMCs from donors carrying the risk allele IFIH1946T. These findings demonstrate the need for a deeper understanding of how mutations in T1D-associated genes contribute to disease onset in specific cellular contexts.

Physiology-based pharmacokinetic model with relative transcriptomics to evaluate tissue distribution and receptor occupancy of anifrolumab.

Type I interferons contribute to the pathogenesis of several autoimmune disorders, including systemic lupus erythematosus (SLE), systemic sclerosis, cutaneous lupus erythematosus, and myositis. Anifrolumab is a monoclonal antibody that binds to subunit 1 of the type I interferon receptor (IFNAR1). Results of phase IIb and phase III trials led to the approval of intravenous anifrolumab 300 mg every 4 weeks (Q4W) alongside standard therapy in patients with moderate-to-severe SLE. Here, we built a population physiology-based pharmacokinetic (PBPK) model of anifrolumab by utilizing the physiochemical properties of anifrolumab, binding kinetics to the Fc gamma neonatal receptor, and target-mediated drug disposition properties. A novel relative transcriptomics approach was employed to determine IFNAR1 expression in tissues (blood, skin, gastrointestinal tract, lungs, and muscle) using mRNA abundances from bioinformatic databases. The IFNAR1 expression and PBPK model were validated by testing their ability to predict clinical pharmacokinetics over a large dose range from different clinical scenarios after subcutaneous and intravenous anifrolumab dosing. The validated PBPK model predicted high unbound local concentrations of anifrolumab in blood, skin, gastrointestinal tract, lungs, and muscle, which exceeded its IFNAR1 dissociation equilibrium constant values. The model also predicted high IFNAR1 occupancy with subcutaneous and intravenous anifrolumab dosing. The model predicted more sustained IFNAR1 occupancy ≥90% with subcutaneous anifrolumab 120 mg once-weekly dosing vs. intravenous 300 mg Q4W dosing. The results informed the dosing of phase III studies of anifrolumab in new indications and present a novel approach to PBPK modeling coupled with relative transcriptomics in simulating pharmacokinetics of therapeutic monoclonal antibodies.

PBLD promotes IRF3 mediated the type I interferon (IFN-I) response and apoptosis to inhibit viral replication

Recent studies have implicated the phenazine biosynthesis-like domain-containing protein (PBLD) in the negative regulation of the development and progression of various cancers. However, its function in viral infection remains unknown. In this study, we found that PBLD plays important roles in multiple virus infections including BPIV3, SeV, VSV, and HSV-1. Our study revealed that PBLD enhances the expression of type I interferon (IFN-I) and ISGs through interferon regulatory factor 3 (IRF3). Further study indicated that PBLD promotes transcriptional phosphorylation of IRF3 (S385/386), thereby facilitating virus-induced IFN-I production. Interestingly, PBLD mediates virus-triggered mitochondrial apoptosis through its dependence on IRF3 (K313/315). Mechanistically, PBLD facilitated virus-induced apoptosis by recruiting the Puma protein to the mitochondria via IRF3. Additionally, we performed mutational analyses of IRF3, showing that its loss of either transcriptional or apoptotic function markedly increased viral replication. Moreover, macrophages with PBLD deficiency during viral infection exhibited decreased the IFN-I and ISGs expression, exacerbating viral infection. Importantly, mice deficient in PBLD exhibited increased viral replication and susceptibility to SeV infection, leading to decreased survival. Notably, Cedrelone, a chemical activator of PBLD, has the ability to reduce SeV replication. Collectively, we first discovered the new function of PBLD in viral infection, broadening our understanding of potential therapeutic targets and offering new insights for antiviral drug development.

Expression of interferon-stimulated genes, but not polymorphisms in the interferon α/β receptor 2 gene, is associated with coronavirus disease 2019 mortality

Excessive inflammatory response is a hallmark of severe COVID-19. This study investigated the associations between interferon-stimulated genes (ISGs) expression, genetic variation in the interferon α/β receptor 2 (IFNAR2) gene, and COVID-19 mortality.We investigated 67 patients with moderate-to-severe COVID-19. Of them, 22 patients (32.8 %) died because of COVID-19. We examined the expression of ISGs in total RNA of peripheral whole blood. We observed a significant increase in the expression of all ISGs examined in non-surviving patients, indicating a heightened interferon type I signaling activation in non-survived patients. Subsequently, we analyzed whether the increase in ISGs expression was correlated with polymorphism within the IFNAR2 gene. Intriguingly, no significant association was observed between IFNAR2 gene polymorphism and COVID-19 mortality. Similarly, no association was noted between the IFNAR2 and ISGs expression levels.Overall, our data showed that higher ISGs expression, which presumably indicates heightened interferon type I activation, is associated with COVID-19 mortality.

Tissue Kallikrein-1 Suppresses Type I Interferon Responses and Reduces Depressive-Like Behavior in the MRL/lpr Lupus-Prone Mouse Model.

Excessive production and response to Type I interferons (IFNs) is a hallmark of systemic lupus erythematosus (SLE). Neuropsychiatric lupus (NPSLE) is a common manifestation of human SLE, with major depression as the most common presentation. Clinical studies have demonstrated that IFNα can cause depressive symptoms. We have shown that the kallikrein-kinin system (KKS) [comprised of kallikreins (Klks) and bradykinins] and angiotensin-converting enzyme inhibitors suppressed Type I IFN responses in dendritic cells from lupus-prone mice and human peripheral blood mononuclear cells. Tissue Klk genes are decreased in patients with lupus, and giving exogenous Klk1 ameliorated kidney pathology in mice. We retro-orbitally administered mouse klk1 gene-carrying adenovirus in the Murphy Roths Large lymphoproliferative (MRL/lpr) lupus-prone mice at early disease onset and analyzed immune responses and depressive-like behavior. Klk1 improved depressive-like behavior, suppressed interferon-responsive genes and neuroinflammation, and reduced plasma IFNα levels and proinflammatory cytokines. Klk1 also reduced IFNAR1 and JAK1 protein expression, important upstream molecules in Type I IFN signaling. Klk1 reduced bradykinin B1 receptor expression, which is known to induce proinflammatory response. Together, these findings suggest that Klk1 may be a potential therapeutic candidate to control IFNα production/responses and other inflammatory responses in SLE and NPSLE.

Mitochondrial Hyperactivity and Reactive Oxygen Species Drive Innate Immunity to the Yellow Fever Virus-17D Live-Attenuated Vaccine.

The yellow fever virus 17D (YFV-17D) live attenuated vaccine is considered one of the successful vaccines ever generated associated with high antiviral immunity, yet the signaling mechanisms that drive the response in infected cells are not understood. Here, we provide a molecular understanding of how metabolic stress and innate immune responses are linked to drive type I IFN expression in response to YFV-17D infection. Comparison of YFV-17D replication with its parental virus, YFV-Asibi, and a related dengue virus revealed that IFN expression requires RIG-I-like Receptor signaling through MAVS, as expected. However, YFV-17D uniquely induces mitochondrial respiration and major metabolic perturbations, including hyperactivation of electron transport to fuel ATP synthase. Mitochondrial hyperactivity generates reactive oxygen species (mROS) and peroxynitrite, blocking of which abrogated IFN expression in non-immune cells without reducing YFV-17D replication. Scavenging ROS in YFV-17D-infected human dendritic cells increased cell viability yet globally prevented expression of IFN signaling pathways. Thus, adaptation of YFV-17D for high growth uniquely imparts mitochondrial hyperactivity generating mROS and peroxynitrite as the critical messengers that convert a blunted IFN response into maximal activation of innate immunity essential for vaccine effectiveness.

Platycodin D: A promising anti tilapia lake virus natural compound from Platycodon grandiflorus

Tilapia lake virus (TiLV) poses a significant threat to the worldwide tilapia aquaculture sector, and there currently exists no efficient method of containment for this lethal pathogen. Given the widespread utilization of herbal medicines in the treatment of various aquatic animal diseases, it becomes imperative to evaluate and identify herbs that can be used for developing effective drugs against TiLV. We evaluated the efficacy of 20 herbs to inhibit TiLV proliferation in a TiLV infection model (Oreochromis niloticus) through two rounds of screening, with Platycodon grandiflorus extract showing the strongest inhibitory effect on TiLV replication (IC50 = 72.619 mg/L), survival of TiLV-infected tilapia was also improved in individual experiments. Platycodin D (PD), the principal bioactive compound found in Platycodon grandiflorus, demonstrated a dose-dependent reduction in viral load when used for treatment and enhanced the survival rate of tilapia infected with TiLV (42.3 %). The results of serum enzyme activity showed that PD treatment at a concentration of 5 mg/kg significantly enhance the non-specific parameters in tilapia (GSH, ACP, and AKP) (**p < 0.01). Additionally, immune-related gene analysis indicated that PD up-regulated IFN-I, CC2 and other genes expression while down-regulated the expression of pro-inflammatory factors (TNF-α, IL-1β) and TLR2 in the spleen and kidney of tilapia, implying that PD could induce IFN-I expression resulting in potential antiviral effects and inhibit some pro-inflammatory factors to alleviate oxidative stress and inflammatory responses in tilapia. The aforementioned findings suggest that Platycodon grandiflorus could potentially serve as a promising candidate for the development of drugs targeting TiLV. In particular, the bioactive component PD showed a strong anti-TiLV effect, which needs to be further explored.

Crystal Structures of DNA-bound Fish IRF10 and IRF11 Reveal the Determinants of IFN Regulation.

IFN regulatory factors (IRFs) are transcription factors that mediate homeostatic mechanisms of host defense against pathogens. In addition to IRF1-9, which are conserved across vertebrates, teleost fishes have two other IRFs, IRF10 and IRF11. In zebrafish (Danio rerio), IRF10 represses the expression of IFNφ1 and IFNφ3, whereas IRF11 exerts the opposite effect. In this study, we found IRF10 could significantly inhibit the expression of IFNφ1 and IFNφ3 induced by IFN11 to synergistically regulate type I IFN expression. To clarify the synergistically regulatory mechanism of IRF10 and IRF11 in type I IFN expression, we determined and analyzed the crystal structures of the DNA-binding domains (DBDs) of zebrafish IRF10 and IRF11 bound to DNA, as well as IRF11 DBD in apo form. The interactions of IRF10-DBD and IRF11-DBD with DNA backbone were elaborated in detail. Further analysis showed that IRF10 and IRF11 have the same binding patterns and comparable affinities with the IFN-sensitive response elements of IFNφ1 and IFNφ3 promoters. Therefore, IRF10 could function as a controlling factor for IRF11 by competitive binding of the IFN-sensitive response elements to coregulate the host IFN response. Accordingly, similar to IRF1 and IRF2 in mammals, IRF10 and IRF11 act as another pair of negative and positive regulators to balance the antiviral responses in fish.

Abstract P380: ISG15 is responsible for endothelial inflammation and vascular disfunction induced by Spike protein 1 of SARS-CoV-2.

Introduction: COVID19-associated immunopathology is associated with increased production of interferon (IFN)-alpha (IFNα) and lambda3 (IFNL3). Effects of IFNs are mediated by interferon-stimulated genes (ISGs) and influence expression of angiotensin-converting enzyme 2 (ACE2), the receptor for S-protein (S1P) of SARS-CoV-2. We hypothesized that S1P-induced immune/inflammatory responses in endothelial cells (EC) are mediated via IFNs and may be important in vascular dysfunction associated with hypertension. Methods: Human microvascular ECs (MEC) were stimulated with S1P (1µg/mL), IFNα (100ng/mL) or IFNL3 (100IU/mL). Because ACE2, metalloproteinase domain-17 (ADAM17) and type-II transmembrane serine protease (TMPRSS2) are important for SARS-CoV-2 infection, we used inhibitors of ADAM17 (marimastat), ACE2 (MLN4760), and TMPRSS2 (camostat). Gene and protein expression was investigated by real-time PCR and immunoblotting. Vascular function was assessed in mesenteric arteries from wild-type (WT) normotensive and hypertensive mice and in ISG15KO mice. Results: MEC stimulated with S1P increased expression of IFNα (3-fold), IFNL3 (4-fold) and ISGs (2-fold) (p&lt;0.05). IFNα and IFNλ3 increased protein expression of TMPRSS2 and ADAM17, but not ACE2. MEC exhibited higher responses to IFNα (ISG15: 16-fold) than to IFNL3 (ISG15: 1.7-fold) (p&lt;0.05). ISG15 was also increased by S1P, effects that were enhanced by MLN4760 (2-fold) and reduced by marimastat (50%). S1P increased IL-6 mRNA (1.3-fold), TNFα (6.2-fold) and IL-1β (3.3-fold). These responses were increased by by IFNs. IL-6 was increased by IFNα (1,230pg/mL) and IFNL3 (1,124pg/mL) vs control (591pg/mL). This was associated with increased phosphorylation of Stat1 (134%), Stat2 (102%), ERK1/2 (42%). Nitric oxide production and eNOS phosphorylation (Ser1177) were reduced by IFNα and (40%) and IFNL3 (40%). Reduced endothelium-relaxation maximal response (%Emax) was observed in vessels from WT-mice stimulated with IFNα (67%) and IFNL3 (71%) vs control (82%) (p&lt;0.05) but not in vessels from ISG15KO mice. Increased contraction was observed only in vessels from hypertensive mice treated with IFNα (9.1±0.5mN vs control: 7.3±0.3mN, p&lt;0.05). Conclusions: In ECs, S1P, IFNα and IFNL3 increased ISG15 and IL-6, processes that involve ADAM17 and are independent on ACE2. Inflammation induced by S1P was amplified by IFNs. IFNs induce vascular dysfunction through ISG15-dependent mechanisms, with augmented effects in hypertension.

Developing a mouse model of human coronavirus NL63 infection: comparison with rhinovirus-A1B and effects of prior rhinovirus infection.

Human coronavirus (HCoV)-NL63 causes respiratory tract infections in humans and uses angiotensin-converting enzyme 2 (ACE2) as a receptor. We sought to establish a mouse model of HCoV-NL63 and determine whether prior rhinovirus (RV)-A1B infection affected HCoV-NL63 replication. HCoV-NL63 was propagated in LLC-MK2 cells expressing human ACE2. RV-A1B was grown in HeLa-H1 cells. C57BL6/J or transgenic mice expressing human ACE2 were infected intranasally with sham LLC-MK2 cell supernatant or 1 × 105 tissue culture infectious dose (TCID50) units HCoV-NL63. Wild-type mice were infected with 1 × 106 plaque-forming units (PFU) RV-A1B. Lungs were assessed for vRNA, bronchoalveolar lavage (BAL) cells, histology, HCoV-NL63 nonstructural protein 3 (nsp3), and host gene expression by next-generation sequencing and qPCR. To evaluate sequential infections, mice were infected with RV-A1B followed by HCoV-NL63 infection 4 days later. We report that hACE2 mice infected with HCoV-NL63 showed evidence of replicative infection with increased levels of vRNA, BAL neutrophils and lymphocytes, peribronchial and perivascular infiltrates, and expression of nsp3. Viral replication peaked 3 days after infection and inflammation persisted 6 days after infection. HCoV-NL63-infected hACE2 mice showed increased mRNA expression of IFNs, IFN-stimulated proteins, and proinflammatory cytokines. Infection with RV-A1B 4 days before HCoV-NL63 significantly decreased both HCoV-NL63 vRNA levels and airway inflammation. Mice infected with RV-A1B prior to HCoV-NL63 showed increased expression of antiviral proteins compared with sham-treated mice. In conclusion, we established a mouse model of HCoV-NL63 replicative infection characterized by relatively persistent viral replication and inflammation. Prior infection with RV-A1B reduced HCoV-NL63 replication and airway inflammation, indicative of viral interference.NEW & NOTEWORTHY We describe a mouse model of human coronavirus (HCoV) infection. Infection of transgenic mice expressing human angiotensin-converting enzyme 2 (ACE2) with HCoV-NL63 produced a replicative infection with peribronchial inflammation and nonstructural protein 3 expression. Mice infected with RV-A1B 4 days before HCoV-NL63 showed decreased HCoV-NL63 replication and airway inflammation and increased expression of antiviral proteins compared with sham-treated mice. This research may shed light on human coronavirus infections, viral interference, and viral-induced asthma exacerbations.

Expression of human Interferon Regulatory Factor 3 (IRF-3) in alveolar macrophages relates to clinical and functional traits in COPD.

IntroductionChronic obstructive pulmonary disease (COPD) is a frequent cause of morbidity and mortality. Dysregulated and enhanced immune-inflammatory responses have been described in COPD. Recent data showed impaired immune responses and, in particular, of interferon (IFNs) signaling pathway in these patients.AimTo evaluate in peripheral lung of COPD patients, the expression of some of the less investigated key components of the innate immune responses leading to IFN productions including: IFN-receptors (IFNAR1/IFNAR2), IRF-3 and MDA-5. Correlations with clinical traits and with the inflammatory cell profile have been assessed.MethodsLung specimens were collected from 58 subjects undergoing thoracic surgery: 22 COPD patients, 21 smokers with normal lung function (SC) and 15 non-smoker controls (nSC). The expression of IFNAR1, IFNAR2, IRF-3 and MDA-5, of eosinophils and activated NK cells (NKp46+) were quantified in the peripheral lung by immunohistochemistry.ResultsA significant increase of IRF-3 + alveolar macrophages were observed in COPD and SC compared with nSC subjects. However, in COPD patients, the lower the levels of IRF-3 + alveolar macrophages the lower the FEV1 and the higher the exacerbation rate. The presence of chronic bronchitis (CB) was also associated with low levels of IRF-3 + alveolar macrophages. NKp46 + cells, but not eosinophils, were increased in COPD patients compared to nSC patients (p < 0.0001).ConclusionsSmoking is associated with higher levels of innate immune response as showed by higher levels of IRF-3 + alveolar macrophages and NKp46 + cells. In COPD, exacerbation rates, severe airflow obstruction and CB were associated with lower levels of IRF-3 expression, suggesting that innate immune responses characterize specific clinical traits of the disease.

Exogenous melatonin ameliorates embryo-maternal cross-talk in early pregnancy in sheep.

Melatonin plays a crucial role in enhancing reproductive performance in small ruminants. This paper reveals the effects of exogenous melatonin on the placental and endometrial rearrangement in early pregnancy in sheep. Early pregnancy losses cause 25% of pregnancy failures in small ruminants because of asynchrony between conceptus and uterine signals. In this context, melatonin plays a crucial role in sheep reproductive dynamics, but little is known about its effects during the peri-implantation period. We hypothesized that melatonin supports embryo implantation by modulating the uterine microenvironment. This study aimed to assess the effects of exogenous melatonin on the endometrial and early placental rearrangement. Ten multiparous ewes either did (MEL, n = 5) or did not (CTR, n = 5) receive a subcutaneous melatonin implant (18 mg) 50 days before a synchronized mating. On day 21 of pregnancy, the sheep were euthanized. MEL ewes exhibited a higher prolificity rate (2.8 vs 2.0 embryos/ewe) and plasma progesterone levels (3.84 vs 2.96 ng/mL, P < 0.05) than did CTR ewes. Groups did not differ significantly in embryo crown-rump length. MEL placentas had significantly (P < 0.001) more binucleated trophoblast cells in the chorion region, and ovine placental lactogen expression was significantly (P < 0.05) more strongly upregulated than in CTR. Exogenous melatonin increased significantly (P < 0.05) gene expression of angiogenic factors (VEGFA, VEGFR1, IGF1R), IFNAR2, and PR in the caruncular endometrium. Expression of the MT2 receptor in the endometrium and placenta was significantly (P < 0.05) higher in the MEL group. These results indicate that melatonin implants acted differentially on uterine and placental rearrangement. Melatonin increases differentiation in the placenta and induces changes that could promote vessel maturation in the endometrium, suggesting that it enhances the uterine microenvironment in the early stage of pregnancy in sheep.

TRIM5 inhibits the replication of Senecavirus A by promoting the RIG-I-mediated type I interferon antiviral response

Senecavirus A (SVA) is an emerging virus that poses a threat to swine herds worldwide. To date, the role of tripartite motif 5 (TRIM5) in the replication of viruses has not been evaluated. Here, TRIM5 was reported to inhibit SVA replication by promoting the type I interferon (IFN) antiviral response mediated by retinoic acid-inducible gene I (RIG-I). TRIM5 expression was significantly upregulated in SVA-infected cells, and TRIM5 overexpression inhibited viral replication and promoted IFN-α, IFN-β, interleukin-1beta (IL-1β), IL-6, and IL-18 expression. Conversely, interfering with the expression of TRIM5 had the opposite effect. Viral adsorption and entry assays showed that TRIM5 did not affect the adsorption of SVA but inhibited its entry. In addition, TRIM5 promoted the expression of RIG-I and RIG-I-mediated IFNs and proinflammatory cytokines, and this effect was also proven by inhibiting the expression of TRIM5. These findings expand the scope of knowledge on host factors inhibiting the replication of SVA and indicate that targeting TRIM5 may aid in the development of new agents against SVA.

Toll-Like Receptor 8 is Expressed in Monocytes in Contrast to Plasmacytoid Dendritic Cells and Mediates Aberrant Interleukin-10 Responses in Patients With Systemic Sclerosis.

Systemic sclerosis (SSc) is a severe rheumatic disease causing fibrotic tissue rearrangement. Aberrant toll-like receptor (TLR) 8 transcripts in plasmacytoid dendritic cells (pDCs) were recently linked to SSc pathogenesis, which is at least partially mediated by increased type I interferon (IFN-I) responses. Here, we addressed the functional role of TLR8 signaling in different immune cell subsets of patients with SSc. Monocytes, conventional dendritic cells (cDCs), and pDCs from the blood and skin of patients with SSc were analyzed for TLR8 protein expression. To assess TLR function, cytokine responses upon TLR7 and TLR8 stimulation were studied. To identify relevant alterations specific for patients with SSc (n = 16), patients with primary Sjögren disease (pSS; n = 10) and healthy controls (HCs; n = 13) were included into the study. In all individuals, TLR8 was expressed in monocytes and cDCs but not in pDCs. The TLR8 expression levels were overall similar in patients with SSc and pSS and HCs. Additionally, in all study participants, TLR8 stimulation of pDCs did not induce IFN-I expression. In contrast, monocytes from patients with SSc revealed increased interleukin (IL)-10 responses upon TLR8 (patients with SSc vs HCs, P = 0.0126) and TLR7/8 stimulation (patients with SSc vs HCs, P = 0.0170). TLR8 protein is not expressed in pDCs of patients with SSc. Accordingly, they do not respond to TLR8 stimulation. In contrast, monocytes of patients with SSc respond to TLR8 stimulation with increased IL-10 responses. Therefore, TLR8 signaling in monocytes participates in SSc pathogenesis by conferring aberrant IL-10 expression.

Knock down of transforming growth factor beta improves expressions of co-stimulatory molecules, type I interferon-regulated genes, and pro-inflammatory cytokine in PRRSV-inoculated monocyte-derived macrophages

Porcine reproductive and respiratory syndrome virus (PRRSV) induces a poor innate immune response following infection. This study evaluates the effects of transforming growth factor beta 1 (TGFβ1) up-regulated by PRRSV on gene expressions of co-stimulatory molecules, type I interferon (IFN), type I IFN-regulated genes (IRGs), pattern recognition receptors, and pro-inflammatory cytokines in PRRSV-inoculated monocyte-derived macrophages (MDMs). Phosphorothioate-modified antisense oligodeoxynucleotides (AS ODNs) specific to various regions of porcine TGFβ1 mRNA were synthesized, and those specific to the AUG region efficiently knockdown TGFβ1 mRNA expression and protein translation. Transfection of TGFβAS ODNs in MDMs inoculated with either classical PRRSV-2 (cPRRSV-2) or highly pathogenic PRRSV-2 (HP-PRRSV-2) significantly reduced TGFβ1 mRNA expression and significantly increased mRNA expressions of CD80, CD86, IFNβ, IRGs (i.e. IFN regulatory factor 3 (IRF3), IRF7, myxovirus resistance 1, osteopontin, and stimulator of IFN genes), Toll-like receptor 3, and tumor necrosis factor-alpha. Transfection of TGFβAS ODNs in MDMs inoculated with HP-PRRSV-2 also significantly increased mRNA expressions of IFNα, IFNγ, and 2’-5’-oligoadenylate synthetase 1. The quantity of PRRSV-2 RNA copy numbers was significantly reduced in MDMs transfected with TGFβAS ODNs as compared to untransfected MDMs. Recombinant porcine TGFβ1 (rTGFβ1) and recombinant porcine IFNα (rIFNα) sustained and reduced the yields of PRRSV-2 RNA copy numbers in PRRSV-2 inoculated MDMs, respectively. These findings demonstrate a strategy of PRRSV for innate immune suppression via an induction of TGFβ expression. These findings also suggest TGFβ as a potential parameter that future PRRSV vaccine and vaccine adjuvant candidates should take into consideration.

The immun function of dendritic cells in SFTS patients

ObjectivesSevere fever with thrombocytopenia syndrome (SFTS) is a newly emerging infectious disease caused by a novel bunyavirus in which host immune system suppression is thought to be crucial in disease development. Dendritic cells (DCs) are professional antigen-presenting cells (APCs) critical for initiation and orchestration of the immune response. And it have been suggested that functionally impaired DCs may mediate the suppression of host-specific T-cell immune responses and thus facilitate viral persistence and disease progression.This study was designed to improve the in vitro culture method for DCs and investigate the different immunologic functions of DCs between SFTS patients and healthy people. MethodsAll confirmed SFTS patients (N = 10) were recruited from the Jinan Infectious Diseases Hospital in 2019; routine laboratory parameters were collected. The frequencies, phenotypes were analyzed by flow cytometry. And the levels of 8 cytokines in the cell culture supernatant were detected by flow cytometry. ResultsOn day 8 of the incubation period, cells were harvested and analyzed by flow cytometry. There were significant differences in the rates of CD1a-, CD83-positive cells between SFTS patients and healthy people (all P < 0.05). The detection of 8 cytokines in the culture supernatant showed that the expressions of IFN-α and IFN-γ in the culture supernatant of DC cells in SFTS patients were lower than those in normal people (P < 0.05, P < 0.01). ConclusionsThe present results indicate that DCs may be functionally impaired in SFTS. A decreased level of circulating mDCs was closely correlated with SFTS progression.