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Tumor immunotherapies targeting PD-(L)1 exhibit anti-tumor efficacy in only 10–30% of patients with various cancers. Literature has demonstrated that a "hot tumor" which contains high T lymphocytes in the tumor microenvironment exhibits a better response to immunotherapies than a "cold tumor." This study aimed to investigate whether tumor-intrinsic IFNα and CXCL10 determine the recruitment and activation of CD8+ T cells to become "hot tumor." In this study, we found that CXCL10 overexpressed in a variety of tumors including lung, colon, and liver tumors with a correlation with PD-L1. High PD-L1 and CXCL10 are associated with better survival rates in tumor patients receiving immunotherapies. IFNs-downstream transcriptional factor IRF-1 and STAT1 were correlated with PD-L1 and CXCL10 expression. We demonstrated that IRF-1 and STAT1 were both bound with the promoters of PD-L1 and CXCL10, sharing the same signaling pathway and determining IFNs-mediated PD-L1 and CXCL10 expression. In addition, IFNα significantly increased activation marker IFNγ in PBMCs, promoting M1 type monocyte differentiation, CD4+ T, and CD8+ T cell activation. Particularly, we found that CD8+ T lymphocytes abundantly expressed CXCR3, a receptor of CXCL10, by flow cytometry, indicating that tumor-intrinsic CXCL10 potentially recruited CD8+ T in tumor microenvironment. To demonstrate the hypothesis, immunotherapy-sensitive CT26 and immunotherapy-resistant LL/2 were used and we found that CT26 cells exhibited higher IFNα, IFNγ, CXCL10, and PD-L1 levels compared to LL/2, leading to higher IFNγ expression in mouse splenocytes. Moreover, we found that CD8+ T cells were recruited by CXCL10 in vitro, whereas SCH546738, an inhibitor of CXCR3, inhibited T cell migration and splenocytes-mediated anti-tumor effect. We then confirmed that CT26-derived tumor was sensitive to αPD-L1 immunotherapy and LL/2-tumor was resistant, whereas αPD-L1 significantly increased T lymphocyte activation marker CD107a in CT26-derived BALB/c mice. In conclusion, this study revealed that CXCL10 expression is correlated with PD-L1 in tumors, sharing the same signaling pathway and associating with better immunotherapeutic efficacy. Further evidence in the syngeneic tumor models demonstrated that immunotherapy-sensitive CT26 intrinsically exhibited higher IFNα and CXCL10 compared to immunotherapy-resistant LL/2 to recruit and activate CD8+ T cells in the tumor microenvironment, exhibiting "hot tumor" characteristic of sensitizing αPD-L1 immunotherapies. [ABSTRACT FROM AUTHOR]

The T cell immunoglobulin domain and mucin domain-containing molecule-3 (TIM-3), an inhibitory checkpoint receptor, has been identified as a crucial regulator of cellular immune responses. TIM-3 has been discovered as a receptor involved in the negative regulation of T cells. Recent studies have demonstrated that TIM-3 is expressed on innate immune cells, including dendritic cells (DCs), even at a higher level than T cells. In the tumor microenvironment, the majority of DCs have a monocytic origin. Models for studying such DCs in vitro are DC cultures generated from monocytes in the presence of growth factors. The present study aimed to investigate the expression of TIM-3 in IFNα-induced monocyte-derived DCs (IFN-DCs) and the impact of DC activation on TIM-3 expression. DCs were obtained by culturing the adherent fraction of mononuclear cells from healthy donors for 4 days in the presence of GM-CSF and IFNα, followed by LPS addition for 24 hours. Human double-stranded DNA (dsDNA, 5 μg/mL) was added as an activation stimulus to intact IFN-DCs at the stage of maturation, along with LPS. Expression of the membrane TIM-3 molecule was determined by flow cytometry, and the level of expression of TIM-3 mRNA – by real-time RT-PCR with reverse transcription. Intact donor IFN-DCs expressed the membrane TIM-3 molecule at a high level (more than 70% of cells). The addition of LPS as a maturation stimulus almost halved the expression of TIM-3 (pW < 0.05) without affecting the expression of HAVCR2/TIM-3 mRNA. Exogenous dsDNA (along with LPS) increased the expression of HAVCR2/TIM-3 mRNA by more than three times (pW = 0.05) with a decrease in the number of TIM-3+DCs (pW = 0.003). Our findings indicate the presence of mechanisms that support expression of this inhibitory checkpoint receptor under conditions of DC activation. Further studies of the regulation of TIM-3 expression by monocyte-derived dendritic cells will expand the understanding of the biological significance of inhibitory receptors on DCs from the point of view of the immune response, as well as, in the future, increase the effectiveness of current approaches in cancer immunotherapy using IFN-DCs and inhibitors of checkpoint molecules.

E11 causes acute fulminant hepatitis in newborns. We investigated the pathological changes of different tissues from premature male twins who died due to E11 infection. The E11 expression level was higher in the liver than in other tissues. IP10 was upregulated in liver tissue in the patient group, and might be regulated by IFNAR and IRF7, whereas IFNα was regulated by IFNAR or IRF5. [ABSTRACT FROM AUTHOR]

Probiotic consumption strongly influences local intestinal immunity and systemic immune status. Heyndrickxia coagulans strain SANK70258 (HC) is a spore-forming lactic acid bacterium that has immunostimulatory properties on peripheral tissues. However, few reports have examined the detailed effectiveness of HC on human immune function and its mechanism of action. Therefore, we conducted a randomized, double-blind, placebo-controlled, parallel-group study to comprehensively evaluate the effects of HC on immunostimulatory capacity, upper respiratory tract infection (URTI) symptoms, and changes in intestinal organic-acid composition. Results of a questionnaire survey of URTI symptoms showed that runny nose, nasal congestion, sneezing, and sore throat scores as well as the cumulative number of days of these symptoms were significantly lower in the HC group than in the placebo group during the study period. Furthermore, the salivary secretory immunoglobulin A (sIgA) concentration was significantly higher, and the natural killer (NK) cell activity tended to be higher in the HC group than in the placebo group. In addition, we performed an exposure culture assay of inactivated influenza virus on peripheral blood mononuclear cells (PBMCs) isolated from the blood of participants in the HC and placebo groups. Gene-expression analysis in PBMCs after culture completion showed that IFNα and TLR7 expression levels were significantly higher in the HC group than in the placebo group. In addition, the expression levels of CD304 tended to be higher in the HC group than in the placebo group. On the other hand, the HC group showed a significantly higher increase in the intestinal butyrate concentration than the placebo group. HC intake also significantly suppressed levels of IL-6 and TNFα produced by PBMCs after exposure to inactivated influenza virus. Collectively, these results suggest that HC activated plasmacytoid dendritic cells expressing TLR7 and CD304 and strongly induced IFNα production, subsequently activating NK cells and increasing sIgA levels, and induced anti-inflammatory effects via increased intestinal butyrate levels. These changes may contribute to the acquisition of host resistance to viral infection and URTI prevention. [ABSTRACT FROM AUTHOR]

IntroductionCrohn’s disease (CD) is characterized by chronic inflammation and intestinal fibrosis leading to lifelong complications. However, the disease pathogenesis remains elusive, and the therapeutic options are limited. Here, we investigated the interaction between neutrophils and intestinal fibroblasts in the development of CD immunofibrosis, a disease mechanism predisposing to inflammatory and fibrotic complications.MethodsPeripheral neutrophils, enriched neutrophil extracellular traps (eNETs), serum, primary intestinal fibroblasts (PIFs) and intestinal biopsies from CD, ulcerative colitis (UC) patients, and healthy individuals (HI), were studied. Transcriptome analysis of neutrophils, multi-cytokine profiling and cell-based functional assays at mRNA/protein level were performed.ResultsCompared to UC, PIFs from CD patients, independently to the presence of strictures, displayed a distinct pro-fibrotic phenotype characterized by negative Krüppellike Factor-2 (KLF2) and increased cellular communication network factor-2 (CCN2) expression leading to collagen production. In both UC and CD, PIFs-derived IL-8 acted as a culprit chemoattractant for neutrophils in the intestine, where CD neutrophils were accumulated close to fibrotic lesions. Functionally, only CD neutrophils via eNETs induced a CD-like phenotype in HI PIFs, suggesting their fibrotic plasticity. High IFNa in serum and IFΝ-responsive signature in peripheral neutrophils were observed in CD, distinguishing it from UC. Moreover, CD serum stimulated the release of fibrogenic eNETs from neutrophils in an IFNa-dependent manner, suggesting the priming role of IFNa in circulating neutrophils. Inhibition of eNETs or JAK signaling in neutrophils or PIFs prevented the neutrophil-mediated fibrotic effect on PIFs. Furthermore, both serum IFNa levels and mRNA levels of key IFN signaling components in neutrophils were wellcorrelated with CD severity.ConclusionsThis study reveals the important role of the IFNa/neutrophil/fibroblast axis in CD immunofibrosis, suggesting candidate biomarkers and putative therapeutic targets.

Probiotic consumption strongly influences local intestinal immunity and systemic immune status. Heyndrickxia coagulans strain SANK70258 (HC) is a spore-forming lactic acid bacterium that has immunostimulatory properties on peripheral tissues. However, few reports have examined the detailed effectiveness of HC on human immune function and its mechanism of action. Therefore, we conducted a randomized, double-blind, placebo-controlled, parallel-group study to comprehensively evaluate the effects of HC on immunostimulatory capacity, upper respiratory tract infection (URTI) symptoms, and changes in intestinal organic-acid composition. Results of a questionnaire survey of URTI symptoms showed that runny nose, nasal congestion, sneezing, and sore throat scores as well as the cumulative number of days of these symptoms were significantly lower in the HC group than in the placebo group during the study period. Furthermore, the salivary secretory immunoglobulin A (sIgA) concentration was significantly higher, and the natural killer (NK) cell activity tended to be higher in the HC group than in the placebo group. In addition, we performed an exposure culture assay of inactivated influenza virus on peripheral blood mononuclear cells (PBMCs) isolated from the blood of participants in the HC and placebo groups. Gene-expression analysis in PBMCs after culture completion showed that IFNα and TLR7 expression levels were significantly higher in the HC group than in the placebo group. In addition, the expression levels of CD304 tended to be higher in the HC group than in the placebo group. On the other hand, the HC group showed a significantly higher increase in the intestinal butyrate concentration than the placebo group. HC intake also significantly suppressed levels of IL-6 and TNFα produced by PBMCs after exposure to inactivated influenza virus. Collectively, these results suggest that HC activated plasmacytoid dendritic cells expressing TLR7 and CD304 and strongly induced IFNα production, subsequently activating NK cells and increasing sIgA levels, and induced anti-inflammatory effects via increased intestinal butyrate levels. These changes may contribute to the acquisition of host resistance to viral infection and URTI prevention.

Tyrosine kinase 2 (TYK2) is involved in type I interferon (IFN-I) signaling through IFN receptor 1 (IFNAR1). This signaling pathway is crucial in the early antiviral response and remains incompletely understood on B cells. Therefore, to understand the role of TYK2 in B cells, we studied these cells under homeostatic conditions and following in vitro activation using Tyk2-deficient (Tyk2−/−) mice. Splenic B cell subpopulations were altered in Tyk2−/− compared to wild type (WT) mice. Marginal zone (MZ) cells were decreased and aged B cells (ABC) were increased, whereas follicular (FO) cells remained unchanged. Likewise, there was an imbalance in transitional B cells in juvenile Tyk2−/− mice. RNA sequencing analysis of adult MZ and FO cells isolated from Tyk2−/− and WT mice in homeostasis revealed altered expression of IFN-I and Toll-like receptor 7 (TLR7) signaling pathway genes. Flow cytometry assays corroborated a lower expression of TLR7 in MZ B cells from Tyk2−/− mice. Splenic B cell cultures showed reduced proliferation and differentiation responses after activation with TLR7 ligands in Tyk2−/− compared to WT mice, with a similar response to lipopolysaccharide (LPS) or anti-CD40 + IL-4. IgM, IgG, IL-10 and IL-6 secretion was also decreased in Tyk2−/− B cell cultures. This reduced response of the TLR7 pathway in Tyk2−/− mice was partially restored by IFNα addition. In conclusion, there is a crosstalk between TYK2 and TLR7 mediated by an IFN-I feedback loop, which contributes to the establishment of MZ B cells and to B cell proliferation and differentiation. [ABSTRACT FROM AUTHOR]

Natural killer (NK) cells are important antiviral effector cells and also involved in tumor clearance. NK cells express IFNAR, rendering them responsive to Type I IFNs. To evaluate Type I IFN-mediated modulation of NK cell functions, individual Type I IFNs subtypes were assessed for their ability to activate NK cells. Different Type I IFN subtypes displayed a broad range in the capacity to induce and modulate NK cell activation and degranulation, measured by CD69 and CD107a expression in response to leukemia cell line K562. When including biological sex as a variable in the analysis, transwell co-cultures of NK cells with either male-or female-derived PBMCs or pDCs stimulated with the TLR7/8 agonist CL097 showed that NK cells were more activated by CL097-stimulated cells derived from females. These sexspecific differences were linked to higher CL097-induced IFNa production by pDCs derived from females, indicating an extrinsic sex-specific effect of Type I IFNs on NK cell function. Interestingly, in addition to the extrinsic effect, we also observed NK cell-intrinsic sex differences, as female NK cells displayed higher activation levels after IFNa-stimulation and after co-culture with CL097-stimulated pDCs, suggesting higher activation of IFNa-signaling transduction in female NK cells. Taken together, the results from these studies identify both extrinsic and intrinsic sex-specific differences in Type I IFN-dependent NK cell functions, contributing to a better understanding of sex-specific differences in innate immunity. [ABSTRACT FROM AUTHOR]

Myeloproliferative neoplasms (MPN) are rare hematologic disorders characterized by clonal hematopoiesis. Familial clustering is observed in a subset of cases, with a notable proportion exhibiting heterozygous germline mutations in DNA double-strand break repair genes (e.g., BRCA1). We investigated the therapeutic potential of targeting BRCA1 haploinsufficiency alongside the JAK2V617F driver mutation. We assessed the efficacy of combining the PARP inhibitor olaparib with interferon-alpha (IFNα) in CRISPR/Cas9-engineered Brca1+/− Jak2V617F-positive 32D cells. Olaparib treatment induced a higher number of DNA double-strand breaks, as demonstrated by γH2AX analysis through Western blot (p = 0.024), flow cytometry (p = 0.013), and confocal microscopy (p = 0.071). RAD51 foci formation was impaired in Brca1+/− cells compared to Brca1+/+ cells, indicating impaired homologous recombination repair due to Brca1 haploinsufficiency. Importantly, olaparib enhanced apoptosis while diminishing cell proliferation and viability in Brca1+/− cells compared to Brca1+/+ cells. These effects were further potentiated by IFNα. Olaparib induced interferon-stimulated genes and increased endogenous production of IFNα in Brca1+/− cells. These responses were abrogated by STING inhibition. In conclusion, our findings suggest that the combination of olaparib and IFNα presents a promising therapeutic strategy for MPN patients by exploiting the synthetic lethality between germline BRCA1 mutations and the JAK2V617F MPN driver mutation. [ABSTRACT FROM AUTHOR]

Natural killer (NK) cells are important antiviral effector cells and also involved in tumor clearance. NK cells express IFNAR, rendering them responsive to Type I IFNs. To evaluate Type I IFN-mediated modulation of NK cell functions, individual Type I IFNs subtypes were assessed for their ability to activate NK cells. Different Type I IFN subtypes displayed a broad range in the capacity to induce and modulate NK cell activation and degranulation, measured by CD69 and CD107a expression in response to leukemia cell line K562. When including biological sex as a variable in the analysis, transwell co-cultures of NK cells with either male- or female-derived PBMCs or pDCs stimulated with the TLR7/8 agonist CL097 showed that NK cells were more activated by CL097-stimulated cells derived from females. These sex-specific differences were linked to higher CL097-induced IFNα production by pDCs derived from females, indicating an extrinsic sex-specific effect of Type I IFNs on NK cell function. Interestingly, in addition to the extrinsic effect, we also observed NK cell-intrinsic sex differences, as female NK cells displayed higher activation levels after IFNα-stimulation and after co-culture with CL097-stimulated pDCs, suggesting higher activation of IFNα-signaling transduction in female NK cells. Taken together, the results from these studies identify both extrinsic and intrinsic sex-specific differences in Type I IFN-dependent NK cell functions, contributing to a better understanding of sex-specific differences in innate immunity.

SARS-CoV-2 infection has claimed just over 1.1 million lives in the US since 2020. Globally, the SARS-CoV-2 respiratory infection spread to 771 million people and caused mortality in 6.9 million individuals to date. Much of the early literature showed that SARS-CoV-2 immunity was defective in the early stages of the pandemic, leading to heightened and, sometimes, chronic inflammatory responses in the lungs. This lung-associated ‘cytokine storm’ or ‘cytokine release syndrome’ led to the need for oxygen supplementation, respiratory distress syndrome, and mechanical ventilation in a relatively high number of people. In this study, we evaluated circulating PBMC from non-hospitalized, male and female, COVID-19+ individuals over the course of infection, from the day of diagnosis (day 0) to one-week post diagnosis (day 7), and finally 4 weeks after diagnosis (day 28). In our early studies, we included hospitalized and critically care patient PBMC; however, most of these individuals were lymphopenic, which limited our assessments of their immune integrity. We chose a panel of 30 interferon-stimulated genes (ISG) to evaluate by PCR and completed flow analysis for immune populations present in those PBMC. Lastly, we assessed immune activation by stimulating PBMC with common TLR ligands. We identified changes in innate cells, primarily the innate lymphoid cells (ILC, NK cells) and adaptive immune cells (CD4+ and CD8+ T cells) over this time course of infection. We found that the TLR-7 agonist, Resiquimod, and the TLR-4 ligand, LPS, induced significantly better IFNα and IFNγ responses in the later phase (day 28) of SARS-CoV-2 infection in those non-hospitalized COVID-19+ individuals as compared to early infection (day 0 and day 7). We concluded that TLR-7 and TLR-4 agonists may be effective adjuvants in COVID-19 vaccines for mounting immunity that is long-lasting against SARS-CoV-2 infection.

Human inborn errors of immunity (IEI) affecting the type I interferon (IFN-I) induction pathway have been associated with predisposition to severe viral infections. Hemophagocytic lymphohistiocytosis (HLH) is a life-threatening systemic hyperinflammatory syndrome that has been increasingly associated with inborn errors of IFN-I-mediated innate immunity. Here is reported a novel case of complete deficiency of STAT2 in a 3-year-old child that presented with typical features of HLH after mumps, measles, and rubella vaccination at the age of 12 months. Due to the life-threatening risk of viral infection, she received SARS-CoV-2 mRNA vaccination. Unfortunately, she developed multisystem inflammatory syndrome in children (MIS-C) after SARS-CoV-2 infection, 4 months after the last dose. Functional studies showed an impaired IFN-I-induced response and a defective IFNα expression at later stages of STAT2 pathway induction. These results suggest a possible more complex mechanism for hyperinflammatory reactions in this type of patients involving a possible defect in the IFN-I production. Understanding the cellular and molecular links between IFN-I-induced signaling and hyperinflammatory syndromes can be critical for the diagnosis and tailored management of these patients with predisposition to severe viral infection. [ABSTRACT FROM AUTHOR]

Purpose: Dry eye disease (DED) is characterized by altered ocular surface proinflammatory and antiinflammatory factors. Interferons (IFNs) are a class of pleiotropic cytokines well known for their antimicrobial, inflammatory, and immunomodulatory roles. Hence, this study investigates the ocular surface expression of different types of IFNs in patients with DED. Methods: The cross-sectional, observational study included patients with DED and normal subjects. Conjunctival impression cytology (CIC) samples were obtained from the study subjects (controls, n = 7; DED, n = 8). The mRNA expression levels of type 1 IFN (IFNα, IFNβ), type 2 IFN (IFNγ), and type 3 IFN (IFNλ1, IFNλ2, IFNλ3) were measured by quantitative PCR (polymerase chain reaction) in CIC samples. IFNα and IFNγ expression under hyperosmotic stress was also studied in human corneal epithelial cells (HCECs) in vitro. Results: The mRNA expression levels of IFNα and IFNβ were significantly lower and that of IFNγ was significantly higher in DED patients compared to healthy controls. The mRNA levels of IFNα, IFNβ, and IFNλ were significantly lower compared to IFNγ in DED patients. An inverse association between tonicity-responsive enhancer-binding protein (TonEBP; hyperosmotic stress maker) and IFNα or IFNβ expression and a positive association between TonEBP and IFNγ expression was observed in CIC samples. The expression of IFNα was lower than IFNγ in HCECs undergoing hyperosmotic stress compared to HCECs without the stress. Conclusion: The presence of an imbalance between type 1 and type 2 IFNs in DED patients suggests newer pathogenic processes in DED, plausible ocular surface infection susceptibility in DED patients, and potential therapeutic targets in the management of DED.

BACKGROUND:The dysfunction of type I interferon (IFN) signaling is an important mechanism of immune escape and metastasis in tumors. Increased NOS1 expression has been detected in melanoma, which correlated with dysfunctional IFN signaling and poor response to immunotherapy, but the specific mechanism has not been determined. In this study, we investigated the regulation of NOS1 on the interferon response and clarified the relevant molecular mechanisms. METHODS:After stable transfection of A375 cells with NOS1 expression plasmids, the transcription and expression of IFNα-stimulated genes (ISGs) were assessed using pISRE luciferase reporter gene analysis, RT-PCR, and western blotting, respectively. The effect of NOS1 on lung metastasis was assessed in melanoma mouse models. A biotin-switch assay was performed to detect the S-nitrosylation of HDAC2 by NOS1. ChIP-qPCR was conducted to measure the binding of HDAC2, H4K16ac, H4K5ac, H3ac, and RNA polymerase II in the promoters of ISGs after IFNα stimulation. This effect was further evaluated by altering the expression level of HDAC2 or by transfecting the HDAC2-C262A/C274A site mutant plasmids into cells. The coimmunoprecipitation assay was performed to detect the interaction of HDAC2 with STAT1 and STAT2. Loss-of-function and gain-of-function approaches were used to examine the effect of HDAC2-C262A/C274A on lung metastasis. Tumor infiltrating lymphocytes were analyzed by flow cytometry. RESULTS:HDAC2 is recruited to the promoter of ISGs and deacetylates H4K16 for the optimal expression of ISGs in response to IFNα treatment. Overexpression of NOS1 in melanoma cells decreases IFNα-responsiveness and induces the S-nitrosylation of HDAC2-C262/C274. This modification decreases the binding of HDAC2 with STAT1, thereby reducing the recruitment of HDAC2 to the ISG promoter and the deacetylation of H4K16. Moreover, expression of a mutant form of HDAC2, which cannot be nitrosylated, reverses the inhibition of ISG expression by NOS1 in vitro and decreases NOS1-induced lung metastasis and inhibition of tumor infiltrating lymphocytes in a melanoma mouse model. CONCLUSIONS:This study provides evidence that NOS1 induces dysfunctional IFN signaling to promote lung metastasis in melanoma, highlighting NOS1-induced S-nitrosylation of HDAC2 in the regulation of IFN signaling via histone modification.

Abstract Background Sorafenib is a multi-kinase inhibitor that shows antitumor activity in advanced hepatocellular carcinoma. Sorafenib exerts a regulatory effect on immune cells, including T cells, natural killer cells and dendritic cells. Studies have shown that plasmacytoid dendritic cells (pDCs) are functionally impaired in cancer tissues or produce low type I interferon alpha (IFNα) in cancer microenvironments. However, the effects of sorafenib on the function of pDCs have not been evaluated in detail. Methods Normal and patient PBMCs were stimulated with CpG-A to evaluate IFNα production with Flow cytometry and ELISA. Result We analyzed the production of IFNα by PBMCs in patients with advanced HCC under sorafenib treatment. We found that sorafenib-treated HCC patients produced less IFNα than untreated patients. Furthermore, we demonstrated that sorafenib suppressed the production of IFNα by PBMCs or pDCs from heathy donors in a concentration-dependent manner. Conclusion Sorafenib suppressed pDCs function. Given that sorafenib is a currently recommended targeted therapeutic agent against cancer, our results suggest that its immunosuppressive effect on pDCs should be considered during treatment.

Purpose: Dry eye disease (DED) is characterized by altered ocular surface proinflammatory and antiinflammatory factors. Interferons (IFNs) are a class of pleiotropic cytokines well known for their antimicrobial, inflammatory, and immunomodulatory roles. Hence, this study investigates the ocular surface expression of different types of IFNs in patients with DED. Methods: The cross‑sectional, observational study included patients with DED and normal subjects. Conjunctival impression cytology (CIC) samples were obtained from the study subjects (controls, n = 7; DED, n = 8). The mRNA expression levels of type 1 IFN (IFNα, IFNβ), type 2 IFN (IFNγ), and type 3 IFN (IFNλ1, IFNλ2, IFNλ3) were measured by quantitative PCR (polymerase chain reaction) in CIC samples. IFNα and IFNγ expression under hyperosmotic stress was also studied in human corneal epithelial cells (HCECs) in vitro. Results: The mRNA expression levels of IFNα and IFNβ were significantly lower and that of IFNγ was significantly higher in DED patients compared to healthy controls. The mRNA levels of IFNα, IFNβ, and IFNλ were significantly lower compared to IFNγ in DED patients. An inverse association between tonicity‑responsive enhancer‑binding protein (TonEBP; hyperosmotic stress maker) and IFNα or IFNβ expression and a positive association between TonEBP and IFNγ expression was observed in CIC samples. The expression of IFNα was lower than IFNγ in HCECs undergoing hyperosmotic stress compared to HCECs without the stress. Conclusion: The presence of an imbalance between type 1 and type 2 IFNs in DED patients suggests newer pathogenic processes in DED, plausible ocular surface infection susceptibility in DED patients, and potential therapeutic targets in the management of DED. [ABSTRACT FROM AUTHOR]

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by the production of autoantibodies specific to self-molecules in the nucleus, cytoplasm, and cell surface. The diversity of serologic and clinical manifestations observed in SLE patients challenges the development of diagnostics and tools for monitoring disease activity. Elevated type I interferon signature (IFN- I) in SLE leads to dysregulation of innate and adaptive immune function, resulting in autoantibodies production. The most common method to determine IFN-I signature is measuring the gene expression of several IFN-α-inducible genes (IFIGs) in blood samples and calculating a score. Optimal selection of IFIGs improves the sensitivity, specificity, and accuracy of the diagnosis of SLE. We describe the mechanisms of the immunopathogenesis of IFN-I signature (IFNα production) and its clinical consequences in SLE. In addition, we explore the association between IFN-I signature, the presence of autoantibodies, disease activity, medical therapy, and ethnicity. We discuss the presence of IFN-I signature in some patients with other autoimmune diseases, including rheumatoid arthritis, systemic and multiple sclerosis, Sjogren's syndrome, and dermatomyositis. Prospective studies are required to assess the role of IFIG and the best combination of IFIGs to monitor SLE disease activity and drug treatments. [ABSTRACT FROM AUTHOR]

Background: Sorafenib is a multi-kinase inhibitor that shows antitumor activity in advanced hepatocellular carcinoma. Sorafenib exerts a regulatory effect on immune cells, including T cells, natural killer cells and dendritic cells. Studies have shown that plasmacytoid dendritic cells (pDCs) are functionally impaired in cancer tissues or produce low type I interferon alpha (IFNα) in cancer microenvironments. However, the effects of sorafenib on the function of pDCs have not been evaluated in detail.Methods: Normal and patient PBMCs were stimulated with CpG-A to evaluate IFNα production with Flow cytometry and ELISA.Result: We analyzed the production of IFNα by PBMCs in patients with advanced HCC under sorafenib treatment. We found that sorafenib-treated HCC patients produced less IFNα than untreated patients. Furthermore, we demonstrated that sorafenib suppressed the production of IFNα by PBMCs or pDCs from heathy donors in a concentration-dependent manner.Conclusion: Sorafenib suppressed pDCs function. Given that sorafenib is a currently recommended targeted therapeutic agent against cancer, our results suggest that its immunosuppressive effect on pDCs should be considered during treatment. [ABSTRACT FROM AUTHOR]

Interferon alpha (IFNα) leads to therapeutic effects on various tumors, especially renal cell cancer (RCC), by directly protecting against tumors cell proliferation or indirectly inducing an anti-tumor immune response. However, new combination therapies are needed to enhance the efficacy of IFNα and reduce its adverse effects during long-term treatment. In this study, we found that the anti-proliferative effects of IFNα on RCC cells in vitro and in vivo were greater after the allosteric inhibition of SHP2 by SHP099 than after treatment with enzymatic inhibitors of SHP2. SHP099 increased IFNα-induced pro-caspase-1 expression in RCC cells, activated the NLRP3 inflammasome, and induced pyroptosis. Mechanistically, SHP099 not only increased the expression of NLRP3 inflammasome components via the NF-κB signaling pathway, but also further activated the NLRP3 inflammasome by regulating mitochondrial homeostasis through ANT1-mediated reactive oxygen species modulation. Allosteric inhibition of SHP2 by SHP099 also potently enhanced the anti-tumor immunity induced by IFNα by modulating T cell proliferation and infiltration in vitro and in vivo. These results reveal the new function of SHP2 in NLRP3 inflammasome activation and pyroptosis in RCC and provide a basis for further investigating the combination of allosteric SHP2 inhibitors with IFNα in cancer immunotherapy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Introduction: Crohn's disease (CD) is characterized by chronic inflammation and intestinal fibrosis leading to lifelong complications. However, the disease pathogenesis remains elusive, and the therapeutic options are limited. Here, we investigated the interaction between neutrophils and intestinal fibroblasts in the development of CD immunofibrosis, a disease mechanism predisposing to inflammatory and fibrotic complications., Methods: Peripheral neutrophils, enriched neutrophil extracellular traps (eNETs), serum, primary intestinal fibroblasts (PIFs) and intestinal biopsies from CD, ulcerative colitis (UC) patients, and healthy individuals (HI), were studied. Transcriptome analysis of neutrophils, multi-cytokine profiling and cell-based functional assays at mRNA/protein level were performed., Results: Compared to UC, PIFs from CD patients, independently to the presence of strictures, displayed a distinct pro-fibrotic phenotype characterized by negative Krüppellike Factor-2 (KLF2) and increased cellular communication network factor-2 (CCN2) expression leading to collagen production. In both UC and CD, PIFs-derived IL-8 acted as a culprit chemoattractant for neutrophils in the intestine, where CD neutrophils were accumulated close to fibrotic lesions. Functionally, only CD neutrophils via eNETs induced a CD-like phenotype in HI PIFs, suggesting their fibrotic plasticity. High IFNa in serum and IFΝ-responsive signature in peripheral neutrophils were observed in CD, distinguishing it from UC. Moreover, CD serum stimulated the release of fibrogenic eNETs from neutrophils in an IFNa-dependent manner, suggesting the priming role of IFNa in circulating neutrophils. Inhibition of eNETs or JAK signaling in neutrophils or PIFs prevented the neutrophil-mediated fibrotic effect on PIFs. Furthermore, both serum IFNa levels and mRNA levels of key IFN signaling components in neutrophils were wellcorrelated with CD severity., Conclusions: This study reveals the important role of the IFNa/neutrophil/fibroblast axis in CD immunofibrosis, suggesting candidate biomarkers and putative therapeutic targets., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. The author(s) declared that they were an editorial board member of Frontiers, at the time of submission. This had no impact on the peer review process and the final decision., (Copyright © 2024 Gavriilidis, Divolis, Natsi, Kafalis, Kogias, Antoniadou, Synolaki, Pavlos, Koutsi, Didaskalou, Papadimitriou, Tsironidou, Gavriil, Papadopoulos, Agelopoulos, Tsilingiris, Koffa, Giatromanolaki, Kouklakis, Ritis and Skendros.)

23 children diagnosed with acute infectious mononucleosis were hospitalized and examined after a short prednisolone treatment course. Related interferon status during infection was compared with that in 38 patients with acute infectious mononucleosis receiving no hormone therapy. Interferon status was investigated by Ershov method, allowing to estimate amount of interferon in the blood serum samples or patient blood cell culture by assessing interferon biological activity. Along with measuring IFNα or IFNγ biological activity, their level was quantified by using enzyme immunoassay. Immunological examination conducted on the next day after the end of hormone therapy revealed sharply decreased potential of patient blood cells to produce both IFNα and IFNγ. The multiplicity of IFNα and IFNγ titer reduction in various patients varied by 4–5 and 3–4-fold, respectively. The concentration of IFNα, determined by ELISA, decreased by 4–6-fold, whereas for IFNγ — by 1.5–2-fold. A follow-up examination 1 month after discharge from the clinic showed that mean IFNα titer in children aged 3–6 years and treated with prednisolone was significantly reduced compared to the baseline, whereas most patients receiving no hormone therapy had normal IFNα production. The change in the level of IFNα 1 month after hormone therapy in 7–14-year age group was similar. IFNγ production quickly recovered, and 1 month after discharge from the clinic, its concentration in culture supernatants from patients reached 10–15 ng/ml, exceeding normal values more than twice. The biological activity of IFNγ in these culture supernatants was significantly higher than those immediately after hormone therapy, whereas in 3–6-year-old group of patients it was also higher than baseline level. These results can serve as a laboratory justification for including recombinant IFNα-2b drugs in the therapy of such patients, presumably immediately after the end of hormone course. Overall, laboratory justified administration of interferon preparations seems to be necessary to determine optimal timepoint for applying such drugs to increase effectiveness for achieving a durable patient recovery.

The study of the interferon (IFN) α-induced cell transcriptome has shown altered expression of several long non-coding RNAs (lncRNAs). ISR8/IRF1-AS1 (IFN stimulated RNA 8), located close to IFN regulatory factor 1 (IRF1) coding gene, transcribes a lncRNA induced at early times after IFNα treatment or IRF1 or NF-κB activation. Depletion or overexpression of ISR8 RNA does not lead to detected deregulation of the IFN response. Surprisingly, disruption of ISR8 locus with CRISPR-Cas9 genome editing results in cells that fail to induce several key ISGs and pro-inflammatory cytokines after a trigger with IFNα or overexpression of IRF1 or the NF-κB subunit RELA. This suggests that the ISR8 locus may play a relevant role in IFNα and NF-κB pathways. Interestingly, IFNα, IRFs and NF-κB-responding luciferase reporters are normally induced in ISR8-disrupted cells when expressed from a plasmid but not when integrated into the genome. Therefore, IFNα and NF-κB pathways are functional to induce the expression of exogenous episomic transcripts but fail to activate transcription from genomic promoters. Transcription from these promoters is not restored with silencing inhibitors, by decreasing the levels of several negative regulators or by overexpression of inducers. Transcriptome analyses indicate that ISR8-disrupted cells have a drastic increase in the levels of negative regulators such as XIST and Zinc finger proteins. Our results agree with ISR8 loci being an enhancer region that is fundamental for proper antiviral and proinflammatory responses. These results are relevant because several SNPs located in the ISR8 region are associated with chronic inflammatory and autoimmune diseases including Crohn's disease, inflammatory bowel disease, ulcerative colitis or asthma. [ABSTRACT FROM AUTHOR]

Abstract Background The requirement of promyelocytic leukaemia protein (PML) in interferon (IFN)-induced cell apoptosis is well-established. However, the exact mechanisms by which the multiple isoforms of PML protein participate in this process remain not well-understood. We previously demonstrated that PML isoform II (PML-II) positively regulates induced gene expression during a type I IFN response and evaluate here how PML-II contributes to IFNα-induced cell death. Methods HeLa cells were transiently depleted of PML-II by siRNA treatment and the response of these cells to treatment with IFNα assessed by molecular assays of mRNA and proteins associated with IFN and apoptosis responses. Results In HeLa cells, death during IFNα stimulation was reduced by prior PML-II depletion. PML-II removal also considerably decreased the induced expression of pro-apoptotic ISGs such as ISG54 (IFIT2), and substantially impaired or prevented expression of PUMA and TRAIL, proteins that are associated with the intrinsic and extrinsic apoptotic pathways respectively. Thirdly, PML-II depletion enhanced ERK and AKT pro-survival signaling activation suggesting that PML-II normally suppresses signaling via these pathways, and that lack of PML-II hence led to greater than normal activation of AKT signaling upon IFNα stimulation and consequently increased resistance to IFNα-induced apoptosis. Conclusions The positive contribution of PML-II to the expression of various IFNα-induced pro-apoptotic proteins and its inhibition of pro-survival signaling together provide a mechanistic explanation for reduced apoptosis under conditions of PML deficiency and may account for at least part of the role of PML as a tumor suppressor gene. Video Abstract

The study of the interferon (IFN) α-induced cell transcriptome has shown altered expression of several long non-coding RNAs (lncRNAs). ISR8/IRF1-AS1 (IFN stimulated RNA 8), located close to IFN regulatory factor 1 (IRF1) coding gene, transcribes a lncRNA induced at early times after IFNα treatment or IRF1 or NF-κB activation. Depletion or overexpression of ISR8 RNA does not lead to detected deregulation of the IFN response. Surprisingly, disruption of ISR8 locus with CRISPR-Cas9 genome editing results in cells that fail to induce several key ISGs and pro-inflammatory cytokines after a trigger with IFNα or overexpression of IRF1 or the NF-κB subunit RELA. This suggests that the ISR8 locus may play a relevant role in IFNα and NF-κB pathways. Interestingly, IFNα, IRFs and NF-κB-responding luciferase reporters are normally induced in ISR8-disrupted cells when expressed from a plasmid but not when integrated into the genome. Therefore, IFNα and NF-κB pathways are functional to induce the expression of exogenous episomic transcripts but fail to activate transcription from genomic promoters. Transcription from these promoters is not restored with silencing inhibitors, by decreasing the levels of several negative regulators or by overexpression of inducers. Transcriptome analyses indicate that ISR8-disrupted cells have a drastic increase in the levels of negative regulators such as XIST and Zinc finger proteins. Our results agree with ISR8 loci being an enhancer region that is fundamental for proper antiviral and proinflammatory responses. These results are relevant because several SNPs located in the ISR8 region are associated with chronic inflammatory and autoimmune diseases including Crohn’s disease, inflammatory bowel disease, ulcerative colitis or asthma.

Our report concerns the observations made during the treatment of pneumonia with individually selected bacteriophages in HCAI patients on mechanical ventilation. 19 patients on mechanical ventilation whose condition was complicated by antibiotic-resistant pneumonia were examined. The treatment of patients was supplemented with phage therapy, bacteriophages were selected individually for each patient, taking into account the microbial etiology of the disease (Pseudomonas aeruginosa, Кlebsiella pneumoniae, Acinetobacter baumanii). Immunophenotyping of blood lymphocytes was carried out using 2-3-parameter flow cytometry. The functional activity of blood leukocytes was assessed by their ability to produce IFNα and IFNγ during cultivation. The level of interferons production in supernatants collected after cultivation was quantitatively evaluated both by their concentration (ELISA, reagents from “Vector-Best-Europe”, Russia) and by their biological activity. Statistical processing of the results was carried out using the Statistica 6 program according to the nonparametric Mann-Whitney U-test. In the course of successful phage therapy with individually selected bacteriophages overcoming of lymphopenia (if there was one) and an increase in both the number and functional activity of peripheral blood lymphocytes in all patients with pneumonia observed are noted. The relationship between the microbial load (mono- or mixed infection, the number of CFU pathogens of pneumonia, the need for repeated courses of phage therapy) and the degree of deficiency in one or another subpopulation of lymphocytes was not detected. Activation of the immune system achieved after one course of phage therapy was maintained for at least 3 weeks after phage administration was discontinued.

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease characterized by the production of autoantibodies specific to self-molecules in the nucleus, cytoplasm, and cell surface. The diversity of serologic and clinical manifestations observed in SLE patients challenges the development of diagnostics and tools for monitoring disease activity. Elevated type I interferon signature (IFN- I) in SLE leads to dysregulation of innate and adaptive immune function, resulting in autoantibodies production. The most common method to determine IFN-I signature is measuring the gene expression of several IFN-α-inducible genes (IFIGs) in blood samples and calculating a score. Optimal selection of IFIGs improves the sensitivity, specificity, and accuracy of the diagnosis of SLE. We describe the mechanisms of the immunopathogenesis of IFN-I signature (IFNα production) and its clinical consequences in SLE. In addition, we explore the association between IFN-I signature, the presence of autoantibodies, disease activity, medical therapy, and ethnicity. We discuss the presence of IFN-I signature in some patients with other autoimmune diseases, including rheumatoid arthritis, systemic and multiple sclerosis, Sjogren’s syndrome, and dermatomyositis. Prospective studies are required to assess the role of IFIG and the best combination of IFIGs to monitor SLE disease activity and drug treatments.

Abstract Background The dysfunction of type I interferon (IFN) signaling is an important mechanism of immune escape and metastasis in tumors. Increased NOS1 expression has been detected in melanoma, which correlated with dysfunctional IFN signaling and poor response to immunotherapy, but the specific mechanism has not been determined. In this study, we investigated the regulation of NOS1 on the interferon response and clarified the relevant molecular mechanisms. Methods After stable transfection of A375 cells with NOS1 expression plasmids, the transcription and expression of IFNα-stimulated genes (ISGs) were assessed using pISRE luciferase reporter gene analysis, RT-PCR, and western blotting, respectively. The effect of NOS1 on lung metastasis was assessed in melanoma mouse models. A biotin-switch assay was performed to detect the S-nitrosylation of HDAC2 by NOS1. ChIP-qPCR was conducted to measure the binding of HDAC2, H4K16ac, H4K5ac, H3ac, and RNA polymerase II in the promoters of ISGs after IFNα stimulation. This effect was further evaluated by altering the expression level of HDAC2 or by transfecting the HDAC2-C262A/C274A site mutant plasmids into cells. The coimmunoprecipitation assay was performed to detect the interaction of HDAC2 with STAT1 and STAT2. Loss-of-function and gain-of-function approaches were used to examine the effect of HDAC2-C262A/C274A on lung metastasis. Tumor infiltrating lymphocytes were analyzed by flow cytometry. Results HDAC2 is recruited to the promoter of ISGs and deacetylates H4K16 for the optimal expression of ISGs in response to IFNα treatment. Overexpression of NOS1 in melanoma cells decreases IFNα-responsiveness and induces the S-nitrosylation of HDAC2-C262/C274. This modification decreases the binding of HDAC2 with STAT1, thereby reducing the recruitment of HDAC2 to the ISG promoter and the deacetylation of H4K16. Moreover, expression of a mutant form of HDAC2, which cannot be nitrosylated, reverses the inhibition of ISG expression by NOS1 in vitro and decreases NOS1-induced lung metastasis and inhibition of tumor infiltrating lymphocytes in a melanoma mouse model. Conclusions This study provides evidence that NOS1 induces dysfunctional IFN signaling to promote lung metastasis in melanoma, highlighting NOS1-induced S-nitrosylation of HDAC2 in the regulation of IFN signaling via histone modification.

E11 causes acute fulminant hepatitis in newborns. We investigated the pathological changes of different tissues from premature male twins who died due to E11 infection. The E11 expression level was higher in the liver than in other tissues. IP10 was upregulated in liver tissue in the patient group, and might be regulated by IFNAR and IRF7, whereas IFNα was regulated by IFNAR or IRF5., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Luo, Wang, Chen, Liu, Zhao, Wu, Huang and Wang.)

Probiotic consumption strongly influences local intestinal immunity and systemic immune status. Heyndrickxia coagulans strain SANK70258 (HC) is a spore-forming lactic acid bacterium that has immunostimulatory properties on peripheral tissues. However, few reports have examined the detailed effectiveness of HC on human immune function and its mechanism of action. Therefore, we conducted a randomized, double-blind, placebo-controlled, parallel-group study to comprehensively evaluate the effects of HC on immunostimulatory capacity, upper respiratory tract infection (URTI) symptoms, and changes in intestinal organic-acid composition. Results of a questionnaire survey of URTI symptoms showed that runny nose, nasal congestion, sneezing, and sore throat scores as well as the cumulative number of days of these symptoms were significantly lower in the HC group than in the placebo group during the study period. Furthermore, the salivary secretory immunoglobulin A (sIgA) concentration was significantly higher, and the natural killer (NK) cell activity tended to be higher in the HC group than in the placebo group. In addition, we performed an exposure culture assay of inactivated influenza virus on peripheral blood mononuclear cells (PBMCs) isolated from the blood of participants in the HC and placebo groups. Gene-expression analysis in PBMCs after culture completion showed that IFNα and TLR7 expression levels were significantly higher in the HC group than in the placebo group. In addition, the expression levels of CD304 tended to be higher in the HC group than in the placebo group. On the other hand, the HC group showed a significantly higher increase in the intestinal butyrate concentration than the placebo group. HC intake also significantly suppressed levels of IL-6 and TNFα produced by PBMCs after exposure to inactivated influenza virus. Collectively, these results suggest that HC activated plasmacytoid dendritic cells expressing TLR7 and CD304 and strongly induced IFNα production, subsequently activating NK cells and increasing sIgA levels, and induced anti-inflammatory effects via increased intestinal butyrate levels. These changes may contribute to the acquisition of host resistance to viral infection and URTI prevention., Competing Interests: MA, NT, YA, SS, AS, NU and RY was employed by the Mitsubishi Chemical Corporation. The remaining author declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Aida, Togawa, Mizuyama, Aoki, Suehiro, Sakamoto, Uchida and Yamada.)

Objectives: Adverse pregnancy outcomes are more common in women with systemic lupus erythematosus (SLE) compared with healthy women, but we lack prognostic biomarkers. Plasma interferon alpha (IFNα) protein levels are elevated in a subgroup of pregnant women with SLE, but whether this is associated with pregnancy outcomes is unknown. We investigated the relationship between IFNα, adverse pregnancy outcomes and the presence of autoantibodies in SLE pregnancy., Methods: We followed 76 women with SLE prospectively. Protein levels of IFNα were quantified in plasma collected in the 2nd and 3rd trimester with single-molecule array. Positivity for antinuclear and antiphospholipid antibodies was assessed during late pregnancy with multiplexed bead assay. Clinical outcomes included the adverse pregnancy outcomes small for gestational age (SGA), preterm birth, and preeclampsia., Results: During SLE pregnancy, women with SGA infants compared with those without had higher levels of plasma IFNα protein, and IFNα positivity was associated with lower birth weight of the infant. Preterm birth was associated with autoantibodies against chromatin. IFNα protein levels associated positively with autoantibodies against chromatin, Smith/ribonucleoprotein (SmRNP) and RNP, but negatively with phospholipid antibodies., Conclusion: Elevated IFNα protein in plasma of women with SLE is a potential risk factor for lower birth weight of their infants. The association between IFNα and lower birth weight warrants further investigation regarding the pathophysiological role of IFNα during SLE pregnancy., (© The Author(s) 2024. Published by Oxford University Press on behalf of the British Society for Rheumatology.)

Hepatopathy is frequently observed in patients with severe malaria but its pathogenesis remains unclear. Galectins are evolutionarily conserved glycan-binding proteins with pleiotropic roles in innate and adaptive immune responses, and exhibit pivotal roles during Plasmodium spp. infection. Here, we analyzed the impact of blockage of galectin-receptor interactions by treatment with alpha (α)-lactose on liver immunopathology during the erythrocytic stage of malaria in mice infected with Plasmodium berghei ANKA (Pb ANKA). Our results found that compared with Pb ANKA-infected mice (malarial mice), blockage of galectin-receptor interactions led to decreased host survival rate and increased peripheral blood parasitemia; exacerbated liver pathology, increased numbers of CD68+ macrophages and apoptotic cells, and increased parasite burden in the livers on days 5 and 7 post infection (p.i.) as well as increased mRNA expression levels of galectin-9 (Gal-9) and its receptor, the T cell immunoglobulin domain and mucin domain protein 3 (Tim-3), interferon (IFN)α, IFNγ, and the triggering receptor expressed on myeloid cells (TREM)-1 in the livers or spleens of Pb ANKA-infected mice on day 7 p.i. Observed by transmission electron microscopy, the peritoneal macrophages isolated from malarial mice with α-lactose treatment had more pseudopodia than those from malarial mice. Measured by using quantitative real-time reverse transcription-polymerase chain reaction assay, the mRNA expression levels of Gal-9, IFNα, IFNβ, IFNγ, and TREM-1 were increased in the peritoneal macrophages isolated from malarial mice with α-lactose treatment in comparison of those from malarial mice. Furthermore, significant positive correlations existed between the mRNA levels of Gal-9 and Tim-3/IFNγ/TREM-1 in both the livers and the peritoneal macrophages, and between Gal-9 and Tim-3/TREM-1 in the spleens of malarial mice; significant positive correlations existed between the mRNA levels of Gal-9 and IFNγ in the livers and between Gal-9 and IFNα in the peritoneal macrophages from malarial mice treated with α-lactose. Our data suggest a potential role of galectin-receptor interactions in limiting liver inflammatory response and parasite proliferation by down-regulating the expressions of IFNα, IFNγ, and TREM-1 during Pb ANKA infection. [ABSTRACT FROM AUTHOR]

Hepatopathy is frequently observed in patients with severe malaria but its pathogenesis remains unclear. Galectins are evolutionarily conserved glycan-binding proteins with pleiotropic roles in innate and adaptive immune responses, and exhibit pivotal roles during Plasmodium spp. infection. Here, we analyzed the impact of blockage of galectin-receptor interactions by treatment with alpha (α)-lactose on liver immunopathology during the erythrocytic stage of malaria in mice infected with Plasmodium berghei ANKA (PbANKA). Our results found that compared with PbANKA-infected mice (malarial mice), blockage of galectin-receptor interactions led to decreased host survival rate and increased peripheral blood parasitemia; exacerbated liver pathology, increased numbers of CD68+ macrophages and apoptotic cells, and increased parasite burden in the livers on days 5 and 7 post infection (p.i.) as well as increased mRNA expression levels of galectin-9 (Gal-9) and its receptor, the T cell immunoglobulin domain and mucin domain protein 3 (Tim-3), interferon (IFN)α, IFNγ, and the triggering receptor expressed on myeloid cells (TREM)-1 in the livers or spleens of PbANKA-infected mice on day 7 p.i. Observed by transmission electron microscopy, the peritoneal macrophages isolated from malarial mice with α-lactose treatment had more pseudopodia than those from malarial mice. Measured by using quantitative real-time reverse transcription-polymerase chain reaction assay, the mRNA expression levels of Gal-9, IFNα, IFNβ, IFNγ, and TREM-1 were increased in the peritoneal macrophages isolated from malarial mice with α-lactose treatment in comparison of those from malarial mice. Furthermore, significant positive correlations existed between the mRNA levels of Gal-9 and Tim-3/IFNγ/TREM-1 in both the livers and the peritoneal macrophages, and between Gal-9 and Tim-3/TREM-1 in the spleens of malarial mice; significant positive correlations existed between the mRNA levels of Gal-9 and IFNγ in the livers and between Gal-9 and IFNα in the peritoneal macrophages from malarial mice treated with α-lactose. Our data suggest a potential role of galectin-receptor interactions in limiting liver inflammatory response and parasite proliferation by down-regulating the expressions of IFNα, IFNγ, and TREM-1 during PbANKA infection.

Background: The requirement of promyelocytic leukaemia protein (PML) in interferon (IFN)-induced cell apoptosis is well-established. However, the exact mechanisms by which the multiple isoforms of PML protein participate in this process remain not well-understood. We previously demonstrated that PML isoform II (PML-II) positively regulates induced gene expression during a type I IFN response and evaluate here how PML-II contributes to IFNα-induced cell death. Methods: HeLa cells were transiently depleted of PML-II by siRNA treatment and the response of these cells to treatment with IFNα assessed by molecular assays of mRNA and proteins associated with IFN and apoptosis responses. Results: In HeLa cells, death during IFNα stimulation was reduced by prior PML-II depletion. PML-II removal also considerably decreased the induced expression of pro-apoptotic ISGs such as ISG54 (IFIT2), and substantially impaired or prevented expression of PUMA and TRAIL, proteins that are associated with the intrinsic and extrinsic apoptotic pathways respectively. Thirdly, PML-II depletion enhanced ERK and AKT pro-survival signaling activation suggesting that PML-II normally suppresses signaling via these pathways, and that lack of PML-II hence led to greater than normal activation of AKT signaling upon IFNα stimulation and consequently increased resistance to IFNα-induced apoptosis. Conclusions: The positive contribution of PML-II to the expression of various IFNα-induced pro-apoptotic proteins and its inhibition of pro-survival signaling together provide a mechanistic explanation for reduced apoptosis under conditions of PML deficiency and may account for at least part of the role of PML as a tumor suppressor gene. 97dGYrHGNNqGsLmmVXdge6 Video Abstract [ABSTRACT FROM AUTHOR]

Tight control of transgene expression is key to ensure the efficacy of a wide range of gene therapy interventions, in which the magnitude and duration of gene expression have to be adjusted to therapeutic needs, thereby limiting secondary effects. The development of upgraded strategies to link transgene expression to pathological stress episodes is an unmet need in gene therapy. Here, we propose an expression strategy that associates transgene expression to an intracellular stress coping mechanism, the unfolded protein response. Specifically, we harnessed the cis elements required to sustain the noncanonical splicing of X-box binding protein 1 (XBP1) messenger RNA (mRNA) in response to the dysfunction of the endoplasmic reticulum (ER), a situation commonly known as ER stress, to drive the expression of heterologous genes. Since ER stress features a wide variety of pathological conditions, including viral infections, cancer, or metabolic disorders, this new expression module stimulates the synthesis of therapeutic genes as a response to cellular damage, and ensures their expression only when necessary. Validation of this inducible expression system was performed in vitro and in vivo, and its potential to limit/inhibit viral infections has been shown in proof-of principle experiments. [ABSTRACT FROM AUTHOR]

Subcutaneous panniculitis‐like T cell lymphoma (SPTCL) is an extremely rare subtype of primary cutaneous T cell lymphomas mimicking panniculitis. Clinically, patients are usually presented with subcutaneous nodules, which usually leads to initial misdiagnosis as a benign cutaneous condition. Here, we report a 40‐year‐old female who presented with subcutaneous erythematous nodules on her extremities with fever. On the basis of the clinical presentations, histopathological features and immunohistochemical findings, a diagnosis of SPTCL was made. The patient was treated with the injection of recombinant human interferon α‐1b (30 μg) every other day for 3 months. The lesions gradually regressed. No new erythema nodules reappeared during the 10‐month follow‐up. [ABSTRACT FROM AUTHOR]

SARS-CoV-2 infection has claimed just over 1.1 million lives in the US since 2020. Globally, the SARS-CoV-2 respiratory infection spread to 771 million people and caused mortality in 6.9 million individuals to date. Much of the early literature showed that SARS-CoV-2 immunity was defective in the early stages of the pandemic, leading to heightened and, sometimes, chronic inflammatory responses in the lungs. This lung-associated 'cytokine storm' or 'cytokine release syndrome' led to the need for oxygen supplementation, respiratory distress syndrome, and mechanical ventilation in a relatively high number of people. In this study, we evaluated circulating PBMC from non-hospitalized, male and female, COVID-19+ individuals over the course of infection, from the day of diagnosis (day 0) to one-week post diagnosis (day 7), and finally 4 weeks after diagnosis (day 28). In our early studies, we included hospitalized and critically care patient PBMC; however, most of these individuals were lymphopenic, which limited our assessments of their immune integrity. We chose a panel of 30 interferon-stimulated genes (ISG) to evaluate by PCR and completed flow analysis for immune populations present in those PBMC. Lastly, we assessed immune activation by stimulating PBMC with common TLR ligands. We identified changes in innate cells, primarily the innate lymphoid cells (ILC, NK cells) and adaptive immune cells (CD4+ and CD8+ T cells) over this time course of infection. We found that the TLR-7 agonist, Resiquimod, and the TLR-4 ligand, LPS, induced significantly better IFNα and IFNγ responses in the later phase (day 28) of SARS-CoV-2 infection in those non-hospitalized COVID-19+ individuals as compared to early infection (day 0 and day 7). We concluded that TLR-7 and TLR-4 agonists may be effective adjuvants in COVID-19 vaccines for mounting immunity that is long-lasting against SARS-CoV-2 infection.

Key points: Type I interferon receptors are expressed by the majority of vagal C‐fibre neurons innervating the respiratory tractInterferon alpha and beta acutely and directly activate vagal C‐fibers in the airways.The interferon‐induced activation of C‐fibers occurs secondary to stimulation of type 1 interferon receptorsType 1 interferons may contribute to the symptoms as well as the spread of respiratory viral infections by causing coughing and other defensive reflexes associated with vagal C‐fibre activation We evaluated the ability of type I interferons to acutely activate airway vagal afferent nerve terminals in mouse lungs. Using single cell RT‐PCR of lung‐specific vagal neurons we found that IFNAR1 and IFNAR2 were expressed in 70% of the TRPV1‐positive neurons (a marker for vagal C‐fibre neurons) and 44% of TRPV1‐negative neurons. We employed an ex vivo vagal innervated mouse trachea‐lung preparation to evaluate the effect of interferons in directly activating airway nerves. Utilizing 2‐photon microscopy of the nodose ganglion neurons from Pirt‐Cre;R26‐GCaMP6s mice we found that applying IFNα or IFNβ to the lungs acutely activated the majority of vagal afferent nerve terminals. When the type 1 interferon receptor, IFNAR1, was blocked with a blocking antibody the response to IFNβ was largely inhibited. The type 2 interferon, IFNγ, also activated airway nerves and this was not inhibited by the IFNAR1 blocking antibody. The Janus kinase inhibitor GLPG0634 (1 μm) virtually abolished the nerve activation caused by IFNβ. Consistent with the activation of vagal afferent C‐fibers, infusing IFNβ into the mouse trachea led to defensive breathing reflexes including apneas and gasping. These reflexes were prevented by pretreatment with an IFN type‐1 receptor blocking antibody. Finally, using whole cell patch‐clamp electrophysiology of lung‐specific neurons we found that IFNβ (1000 U ml−1) directly depolarized the membrane potential of isolated nodose neurons, in some cases beyond to action potential threshold. This acute non‐genomic activation of vagal sensory nerve terminals by interferons may contribute to the incessant coughing that is a hallmark of respiratory viral infections. Key points: Type I interferon receptors are expressed by the majority of vagal C‐fibre neurons innervating the respiratory tractInterferon alpha and beta acutely and directly activate vagal C‐fibers in the airways.The interferon‐induced activation of C‐fibers occurs secondary to stimulation of type 1 interferon receptorsType 1 interferons may contribute to the symptoms as well as the spread of respiratory viral infections by causing coughing and other defensive reflexes associated with vagal C‐fibre activation [ABSTRACT FROM AUTHOR]

Background: Psoriasis is a pro-inflammatory disease with unknown etiology, that is characterized by skin inflammation and keratinocytes hyperproliferation. Specific inhibition of inflammation has shown positive effects avoiding the progression of the psoriatic lesions in different animal models of the disease, turning this strategy as a remarkable therapeutic alternative. Objective: To screen the effectiveness of a novel IFN-α/β signalling inhibitor in the development reduction of skin lesions in IMQ and TPA mice models of psoriasis. Methods: We used a Phage-peptide library for the screening of a peptide with inhibitory effects on the development of psoriasis-like lesions in mice. To evaluate the in vivo effect of the phage-peptides (Phpep3D) and the derived peptide (Pep3D), we administered Phpep3D or Pep3D intradermally in mice with imiquimod (IMQ)-induced psoriasis and 12-O-tetradecanoyl phorbol-13-acetate (TPA)-induced psoriasis. We scored the lesions, and we determined the number of neutrophils and the production of some pro-inflammatory cytokines in the lesions. Results: In this work, we describe how the Ph3pepD and Pep3D reduced skin thickness, redness, and acanthosis despite the presence of the psoriasis inducers, IMQ or TPA. We also found that Pep3D reduced the number of GR1+ infiltrated cells and decreased the production of IL-17A and TNFα in the psoriatic skin of mice. In-silico, docking analysis showed that Pep3D may interact with the interferon-alpha receptor, but further analyses should be performed to uncover the mechanism of action of this peptide. Conclusion: Our results suggest that Pep3D could be used as a new treatment for psoriasis.

Introduction: Ovarian cancer is one of the most lethal gynecologic cancers. Relapses after remission are common, hence novel strategies are urgently needed. Our group has previously developed a vaccination approach based on dendritic cells pulsed with HOCl-oxidized tumor lysates. Here we investigate the improvement of this vaccine strategy using squaric acid treatment of cancer cells during tumor lysate preparation and by differentiating dendritic cells in the presence of GM-CSF and IFNα. Methods: Induction of cell death by squaric acid treatment was assessed with propidium iodide (PI) and Annexin V in ID8 tumor cells. High mobility group box 1 (HMGB1) immunogenic status was analyzed using a western blot-based method, as previously described. For immunological tests, ID8 cells expressing ovalbumin (ova-ID8) were treated with squaric acid before cell lysis. DCs prepared with the canonical GM-CSF and IL-4 differentiation cocktail or IFNα and GM-CSF were pulsed with tumor cell lysates and further matured in the presence of IFNγ and LPS (4-DCs and α-DCs respectively). DCs were then used in co-culture assays with ova-specific T cells and IFNγ and IL-4 secretion measured by ELISA. DC phenotypes were characterized by FACS. Finally, DCs were tested in an ovarian cancer mouse model measuring body weight and animal survival. Results: Squaric acid treatment of mouse ovarian cancer cells induced tumor cell death as well as preserve HMGB1, a crucial Damage-associated molecular pattern (DAMP) signal, in its active reduced form. Squaric acid treatment of ID8-ova cells increased IFNγ and decreased IL-4 production from ova-specific T cells in co-culture experiments, promoting a more immunogenic cytokine secretion pattern. DCs differentiated in the presence of IFNα induced a considerable decrease in IL-4 production compared to canonical 4-DCs, without affecting IFNγ release. DC phenotyping demonstrated a more mature and immunogenic phenotype for IFNα-differentiated DCs. Vaccination in tumor-bearing mice showed that IFNα-differentiated DCs pulsed with squaric acid-treated lysates were the most potent at delaying tumor growth, improving animal survival. Conclusion: We identified squaric acid as a novel immunogenic treatment of tumor cells for cancer vaccines particularly efficient in prolonging animal survival when used in combination with IFNα-differentiated DCs. These promising results support future efforts for the clinical translation of this approach.

Hepatitis B virus (HBV) is a major risk factor for liver diseases, in which HBV covalently closed circular DNA (cccDNA), as the genomic form that templates viral transcription, plays crucial roles in sustaining viral persistence. Clinically, the excessive ethanol intake accelerates the progression of liver diseases with HBV infection. Here, we supposed that ethanol might trigger HBV cccDNA in the liver. Interestingly, we observed that the ethanol remarkably elevated the levels of HBeAg, HBsAg, HBV DNA and cccDNA in HBV-expressing hepatoma cells. Mechanically, the ethanol increased the levels of HBx and MSL2 in vivo and in HBV-expressing HepG2 cells, but not in HBV-free HepG2 cells. Moreover, the down-regulation of MSL2 by small interference RNA could block the ethanol-promoted HBV cccDNA in HepG2.2.15 cells. As a commonly administered treatment for HBV, the effect of IFNα on ethanol-triggered HBV cccDNA remains poorly understood. Strikingly, we showed that the treatment with IFN-α2b inhibited the ethanol-promoted cccDNA through depressing MSL2 in the cells. Thus, we conclude that IFN-α2b inhibits the ethanol-enriched HBV cccDNA through blocking a positive feedback loop of HBx/MSL2/cccDNA/HBV/HBx. Our finding provides new insights into the mechanism by which IFN-α2b inhibits ethanol-enhanced HBV cccDNA. Therapeutically, IFNα may contribute to the cccDNA induced by ethanol in liver. Image 1 • Ethanol triggers the HBV cccDNA. • Ethanol enriches cccDNA via driving a feedback loop of HBx/MSL2/HBV/cccDNA/HBV/HBx. • IFN-α2b down-regulates MSL2 in HBV-free and/or HBV-expressing cells. • IFN-α2b depresses the ethanol-enriched cccDNA through inhibiting MSL2. [ABSTRACT FROM AUTHOR]

Epicutaneous (EC) sensitization with protein allergens is the most important sensitization route for atopic dermatitis. Plasmacytoid dendritic cells (pDCs) are characterized by massive secretion of interferon‐α (IFNα). B6 mice are T helper type 1 (Th1)‐prone and are representative of non‐atopic humans, whereas BALB/c mice are Th2‐prone and are representative of atopic humans. Here, we show that naïve BALB/c mice contain a greater number of nonactivated pDCs in peripheral lymph nodes (LNs) than do naïve B6 mice. Naïve BALB/c mice also have more of the CD8α– subset in LNs than naïve B6 mice. Moreover, in vivo depletion of pDCs during EC sensitization results in enhanced Th2 responses in BALB/c mice, but not in B6 mice. Mechanistically, when BALB/c mice undergo EC sensitization, there is an increase in pDCs entering draining LNs. These cells exhibit modest activation including comparable costimulation expression but increased cytokine expression compared with those of naïve mice. In vivo depletion of pDCs during EC sensitization significantly increases the activation of dermal dendritic cells (dDCs) suggesting a regulatory effect on these cells. To this end, a suppressive effect of pDCs on conventional dendritic cells was also demonstrated in vitro. Further, in vivo blockade of IFNα by an anti‐IFNAR antibody (Ab) or in vivo reduction of IFNα production of pDCs by anti‐siglec‐H Ab both resulted in enhanced activation of dDCs. Collectively, our results demonstrate that pDCs suppress Th2 responses induced by EC sensitization via IFNα‐mediated regulation of dDCs. [ABSTRACT FROM AUTHOR]

The presence of naturally occurring cytokine-specific autoantibodies (c-aAb) in humans is well established, as well as associations to selected pathologies. However, the overall influence of c-aAb on immunocompetence remains largely unknown. In this paper, we performed a large-scale investigation of c-aAb association with infection risk. A cohort of healthy Danish blood donors was screened for c-aAb against IL-1α, IL-6, IL-10, IFNα, and GM-CSF using a Luminex-based multiplex assay, and results were linked to data from the Danish National Prescription Registry. The filing of an antimicrobial prescription following c-aAb measurement was used as a proxy for impaired immunocompetence. We found that c-aAb against pro-inflammatory cytokines IFNα and GM-CSF tended to associate with increased risk of prescription filings in women, whereas antibodies against anti-inflammatory IL-10 were associated with a lower predicted risk of antimicrobial prescriptions, as well as higher self-perceived health scores. We also observed an association of cumulative c-aAb presence with prescription risk. Our data show that cytokine autoantibodies in healthy individuals associate with various proxies for immunomodulation, with the exact association dependent on the pattern of pro- or anti-inflammatory cytokines targeted. This suggests that c-aAb may express cytokine-modulatory properties in healthy individuals and may be critical to further investigate as biomarkers of immunodeficiency. [ABSTRACT FROM AUTHOR]

Treatment of chronic hepatitis B with pegylated-interferon-α-2a (PegIFNα) in pediatric patients can lead to a higher rate of hepatitis B virus (HBV) surface antigen (HBsAg) loss than in adults. However, the mechanism of underlying immune response is not clear. The aim of this study was to explore innate and adaptive immunity, especially HBV-specific T cell responses in hepatitis B e antigen (HBeAg)-positive pediatric patients, who have experienced HBsAg loss. Isolated lymphocytes of 20 HBeAg-positive pediatric patients were collected every 12 weeks until treatment was stopped. The phenotype of T/natural killer (NK) cells and function of HBV-specific T cells were analyzed by flow cytometry. The frequency of CD69 and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) expressed on T cells and TRAIL on CD56hi NK cells in patients with HBsAg loss was remarkably higher compared with nonresponse patients. Furthermore, in vitro peptide stimulation of HBV-specific T cell responses was increased in patients with HBsAg loss when compared with week 0 and 48, and correlated with decline of viral load. The PegIFNα therapy in pediatric patients triggered T/NK cell activation and HBV-specific T cell responses, thereby contributing to successful viral control. [ABSTRACT FROM AUTHOR]

• Both exogenous and endogenous lactate reduced the protein level and activation of the signal transducer and activator of transcription 1(STAT1) in ovarian cancer cells following inhibited the expression of IFNα-STAT1 regulated genes. • LDHA knockdown did not result in the downregulation of STAT1 mRNA level in ovarian cancer cells. Instead, we observed that lactate triggered the degradation of STAT1 through the proteasomal pathway. • Lactate reduced the stability of STAT1 by promoting the expression of F-box only protein 40 (Fbxo40). • Ectopic over-expression of the Fbxo40 gene substantially suppressed the expression of ISGs in LDHA knockdown cells. The emergence of lactate, produced by lactate dehydrogenase A (LDHA), as an important regulator of the immune response in tumor development has garnered attention in recent research. But, many questions still need to be clarified regarding the relationship between lactate and anti-tumor immunity. Here, we reported that both exogenous and endogenous lactate reduced the protein level and activation of the signal transducer and activator of transcription 1(STAT1) in ovarian cancer cells. As a consequence, the expression of IFNα-STAT1 regulated genes was weakened. This, in turn, weakened the antitumor effect of IFNα by impeding NKT and CD8+T cells recruitment. Strikingly, we found that LDHA knockdown did not result in the downregulation of STAT1 mRNA level in ovarian cancer cells. Instead, we observed that lactate triggered the degradation of STAT1 through the proteasomal pathway. Notably, we identified that lactate reduced the stability of STAT1 by promoting the expression of F-box only protein 40 (Fbxo40). This protein interacts with STAT1 and potentially acts as an E3 ubiquitin ligase, leading to the induction of STAT1 polyubiquitination and degradation. Importantly, ectopic over-expression of the Fbxo40 gene significantly inhibited the expression of ISGs in LDHA knockdown cells. In the TCGA tumor data, we observed that high expression of Fbxo40 negatively correlates with overall survival in ovarian cancer patients. Collectively, our findings reveal lactate as a negative regulator of the IFNα-STAT1 signaling axis in ovarian cancer. This discovery suggests that strategies aimed at targeting lactate for ovarian cancer prevention and treatment should consider the impact on the IFNα-STAT1 response. [ABSTRACT FROM AUTHOR]

Irradiation-broken DNA fragments increase type I interferon and chemokines secretion in tumor cells. Since radiotherapy may augment tumor immunotherapy, we hypothesize that the chemokines increased by irradiation could recruit CD8+ T cells to suppress tumor proliferation. This study intended to unveil the secreted factors activating and recruiting CD8+ T cells in non-small-cell lung cancer (NSCLC). EGFR-positive A549 was selected and treated by X-irradiation (IR) to identify the overexpression of chemokines associated to CD8+ T cell cytotoxicity and recruitment. A transwell assay with Alexa 488-labeled CD8+ T cells was used to evaluate CD8+ T cell motility in vitro. A nuclear imaging platform by In111-labeled nivolumab was used to track CD8+ T cells homing to tumors in vivo. The activation markers GZMB, PRF-1, and IFNγ, migration marker CD183 (CXCR3), and inhibitory marker CD274 (PD-1), were measured and compared in CD8+ T cells with A549 co-cultured, chemokines treated, and patients with late-stage lung cancer. We found that IR not only suppressed A549 proliferation but also induced IFNα and CXCL9 expression (p < 0.05). IFNα majorly increased IFNγ levels in CD8+ T cells (p < 0.05) and synergistically with CXCL9 enhanced CD8+ T cell migration in vitro (p < 0.05). We found that CXCR3 and PD-1 were down-regulated and up-regulated, respectively, in the peripheral blood CD8+ T cells in patients with lung cancer (n = 4 vs. healthy n = 3, both p < 0.05), which exhibited reduction of cell motility (p < 0.05). The in vivo nuclear imaging data indicated highly CD8+ T cells migrated to A549-induced tumors. In addition, we demonstrated that healthy PBMCs significantly suppressed the parallel tumor growth (p < 0.05) and the radioresistant tumor growth in the tumor xenograft mice (p < 0.05), but PBMCs from patients with lung cancer had lost the anti-tumor capacity. We demonstrated that IR induced IFNα and CXCL9 expression in A549 cells, leading to CD8+ T cell migration. This study unveiled a potential mechanism for radiotherapy to activate and recruit CD8+ T cells to suppress lung tumors.

Abstract Background Type I interferons are major players against viral infections and mediate their function by the induction of Interferon regulated genes (IRGs). Recently, it became obvious that these cytokines have a multitude of additional functions. Due to the unique features of the chickens’ immune system, available data from mouse models are not easily transferable; hence we performed an extensive analysis of chicken IRGs. Results A broad database search for homologues to described mammalian IRGs (common IRGs, cIRGs) was combined with a transcriptome analysis of spleen and lung at different time points after application of IFNα. To apply physiological amounts of IFN, half-life of IFN in the chicken was determined. Interestingly, the calculated 36 min are considerably shorter than the ones obtained for human and mouse. Microarray analysis revealed many additional IRGs (newly identified IRGs; nIRGs) and network analysis for selected IRGs showed a broad interaction of nIRGs among each other and with cIRGs. We found that IRGs exhibit a highly tissue and time specific expression pattern as expression quality and quantity differed strongly between spleen and lung and over time. While in the spleen for many affected genes changes in RNA abundance peaked already after 3 h, an increasing or plateau-like regulation after 3, 6 and 9 h was observed in the lung. Conclusions The induction or suppression of IRGs in chickens is both tissue and time specific and beside known antiviral mechanisms type I IFN induces many additional cellular functions. We confirmed many known IRGs and established a multitude of so far undescribed ones, thus providing a large database for future research on antiviral mechanisms and additional IFN functions in non-mammalian species.

Plasmacytoid dendritic cells (pDCs) are a minority subset of dendritic cells that despite their tiny quantity play an important role in the immune system, especially in antiviral immunity. They are known mostly as the major producers of type I IFN, which they secrete upon stimulation of endosomal Toll-like receptors 7 and 9 with viral RNA and DNA. However, the functionality of pDCs is more complex, as they were shown to be also involved in autoimmunity, inflammation, and cancer. In the context of the tumor microenvironment, pDCs mostly show substantial functional defects and thus contribute to establishing immunosuppressive micromilieu. Indeed, tumor-infiltrating pDCs were shown to be predominantly pro-tumorigenic, with reduced ability to produce IFNα and capacity to prime regulatory T cells via the ICOS/ICOS-L pathway. Here we describe in detail a method to assess the functional capacity of pDCs upon exposure to tumor-derived cell culture supernatants. The same technique can be implemented with minimal variations to test any soluble factor's impact on pDC phenotype and function., (Copyright © 2024 Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

Sensing self-nucleic acids through toll-like receptors in plasmacytoid dendritic cells (pDCs), and the dysregulated type I IFN production, represent pathogenic events in the development of the autoimmune responses in systemic lupus erythematosus (SLE). Production of high-mobility group box-1 protein (HMGB1) promotes type I IFN response in pDCs. To better understand the active pathogenic mechanism of SLE, we measured serum levels of HMGB1, thrombomodulin, and cytokines (IL-1β, IL-2, IL-4, IL-5, IL-6, IL-10, IL-13, IL-17A, IL-17F, IFNα, IFNγ, TNFα) in 35 patients with SLE. Serum HMGB1 and IFNα were significantly higher in patients with active SLE (SLE Disease Activity Index (SLEDAI) score ≥ 6) compared with healthy donors or patients with inactive SLE. Furthermore, the HMGB1 levels were significantly correlated with IFNα levels. By qualitative analysis, the detection of serum IFNα or HMGB1 suggests active SLE and the presence of SLE-related arthritis, fever, and urinary abnormality out of SLEDAI manifestations. Collectively, HMGB1 and IFNα levels are biomarkers reflecting disease activity, and qualitative analysis of IFNα or HMGB1 is a useful screening test to estimate SLE severity and manifestations. Our results suggest the clinical significance of type I IFNs and HMGB1 as key molecules promoting the autoimmune process in SLE. [ABSTRACT FROM AUTHOR]