Interferon-gamma Expression Research Articles

Newcastle disease virus promotes pyroptosis in medulloblastoma cells by regulating interferon-gamma-mediated guanylate-binding protein 1 expression and activating caspase-4

Objective: The literature has reported that Newcastle disease virus (NDV) can have inhibitory effects on various tumors. This study aims to investigate the mechanism by which NDV induces pyroptosis in medulloblastoma (MB) cells. Material and Methods: We treated MB cell lines Daoy and D283 with NDV or recombinant interferon-gamma (IFN-g) proteins. Guanylate-binding proteins (GBPs) were measured using real-time quantitative polymerase chain reaction. Small interfering RNA-specific targeting GBP1 was transfected into MB cells. Apoptosis was assessed using Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nucleoside nick end labeling and flow cytometry assays. Pyroptosis-related proteins, including caspase-4, caspase-1, and gasdermin D (GSDMD), were detected using Western blotting. Results: Bioinformatics analysis revealed that GBP family genes and interferon-related genes might be responsive to NDV stimulation in MB cells. Treatment with NDV resulted in increased IFN-g levels and upregulated GBP expression, particularly GBP1. In addition, IFN-g treatment induced GBP1 expression and enhanced cell apoptosis. GBP1 knockdown attenuated the decreased cell proliferation and increased cell apoptosis induced by NDV in MB cells. GBP1 overexpression upregulated the expression of pyroptosis-related proteins, including caspase-4, caspase-1, and GSDMD, subsequently leading to inhibition of cell proliferation and an increase in cell apoptosis levels. The silencing of caspase-4 confirmed the regulatory role of GBP1 in MB cell pyroptosis. Conclusion: Our findings suggest that NDV elevates IFN-g and GBP1 expression in MB cells, potentially contributing to caspase-4-mediated pyroptosis activation.

Nrf2 deficiency in muscle attenuates experimental autoimmune myositis-induced muscle weakness.

Idiopathic inflammatory myopathies (IIMs) are systemic autoimmune diseases characterised by muscle weakness. Although multiple physiological and pathological processes are associated with IIMs, T-lymphocyte infiltration into muscle plays a key role in the development and exacerbation of IIMs. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a key transcription factor that regulates inflammatory responses; therefore, muscle Nrf2 may serve an important role in the development of IIMs. In this study, we demonstrated that experimental autoimmune myositis (EAM) causes loss of muscle mass and function in oxidative and glycolytic muscles in C57BL/6 mice. EAM increased CD4+ and CD8+ T-lymphocyte infiltration, as well as interferon-gamma (IFN-γ) and tumour necrosis factor-alpha (TNF-α) mRNA expression in oxidative soleus and glycolytic extensor digitorum longus muscles, along with elevated chemokine mRNA levels (i.e. CCL3, CCL5, CXCL9, CXCL10 and CXCL16). IFN-γ and TNF-α treatments increased the mRNA expression levels of these chemokines in C2C12 myotubes. EAM also increased phosphorylated Nrf2 at Ser40 in soleus and glycolytic white vastus lateralis muscle. Although the expression of several chemokines was affected by Nrf2 activation following tert-butylhydroquinone treatment or Keap1 knockdown, CCL5 mRNA expression significantly increased in C2C12 myotubes and mouse skeletal muscle. Moreover, muscle-specific Nrf2 knockout in mice attenuates EAM-induced loss of muscle mass and function, which was associated with the inhibition of CCL5 mRNA expression, CD8+ T-lymphocyte infiltration and IFN-γ mRNA expression. Collectively, these findings reveal that regulating Nrf2 activity is a promising therapeutic approach for treating IIM-mediated muscle weakness. KEY POINTS: Experimental autoimmune myositis (EAM) causes loss of muscle mass and function. Loss of muscle mass and function in EAM were associated with increased chemokine mRNA expression (i.e. CCL3, CCL5, CXCL9, CXCL10 and CXCL16), T-lymphocyte infiltration and inflammatory cytokine mRNA expression (i.e. IFN-γ and TNF-α) in the skeletal muscle. EAM activated Nrf2 in muscle and increased Nrf2 activity in vivo and in vitro increased CCL5 mRNA expression. Muscle-specific Nrf2 knockout in mice attenuated EAM-induced muscle weakness by inhibiting CCL5 mRNA expression, CD8+ T-lymphocyte migration and IFN-γ mRNA expression in muscles. These results provide further evidence for the potential therapeutic targeting of Nrf2 to mitigate EAM-induced muscle weakness.

Impact of immunosuppressive drugs on efficacy of mesenchymal stem cell therapy for suppressing renal fibrosis.

Preemptive regenerative medicine using mesenchymal stem cells (MSCs) may provide a novel therapeutic approach to prevent the progression from organ damage to organ failure. Although immunosuppressive drugs are often used in patients with organ disorder, their impact on MSC therapy remains unclear. We investigated the effects of immunosuppressive drugs on the therapeutic efficacy of MSCs. We created unilateral ureteral obstruction models, as a well-established model of renal fibrosis, a preliminary stage of organ failure. Three immunosuppressive drugs (methylprednisolone, cyclosporine, and cyclophosphamide) were intraperitoneally administered 3 days after surgery, and MSCs were injected via tail vein the following day. Preadministration of methylprednisolone or cyclophosphamide interfered with MSC activation by reducing expression of interferon-gamma (IFN-γ) and high-mobility group box-1 protein, thus significantly attenuating the therapeutic efficacy of MSCs. Preadministration of cyclophosphamide downregulated the expression of stromal cell-derived factor-1/C-X-C motif ligand 12, which is a potent migration factor for MSCs, resulting in reduced MSC engraftment in the renal cortex. IFN-γ-preconditioned activated MSCs were unaffected by these drugs and maintained their beneficial therapeutic effects. Cyclosporine preadministration had no effect on the therapeutic efficacy of MSCs. Our study demonstrated that the administration of certain immunosuppressive drugs interfered with MSC activation and engraftment at the site of injury, resulting in a significant attenuation of their therapeutic efficacy. These findings provide crucial information for selecting patients suitable for MSC therapy. Use of MSCs preactivated with IFN-γ or other means is preferred for patients on methylprednisolone or cyclophosphamide.

Diagnostic value of IFN-gamma in tuberculous pleural effusion

BackgroundSimple, rapid, and accurate diagnosis of tuberculous pleural effusion (TPE) remains challenging. This study aimed to determine the accuracy of IFN-γ in diagnosing TPE. MethodsWe quantified the expression of interferon-gamma (IFN-γ) in blood (B), adenosine deaminase (ADA), and IFN-γ in pleural effusions (PE) from 25 TPE patients and 31 non-TPE patients using a combination of immunological assays and flow cytometric analysis. The diagnostic performance of these three biomarkers was evaluated using receiver operating characteristic (ROC) curves. ResultsWe found that IFN-γ levels in blood and pleural fluid were higher in the TPE group than in the non-TPE group. The mean concentration of IFN-γ in pleural fluid of the TPE group was 3140.90 (1817.94, 6611.05) pg/mL, while that of the non-TPE group was 4.91 (0.69, 8.6) pg/mL), and the difference was statistically significant (z = 6.39, P < 0.001). The mean blood IFN-γ was 40.19 (16.45, 59.08) pg/mL in the TPE group and 2.76 (1.96, 6.02) pg/mL in the non-TPE group, which was statistically different (z = 5.12, P < 0.001). The area under the ROC curve (AUC) for pleural fluid IFN-γ, blood IFN-γ, and ADA were 0.999 (95 % CI: 0.994–1.00), 0.901 (95 % CI: 0.798–1.00) and 0.996 (95 % CI: 0.987–1.00), respectively. ConclusionThis study confirms that IFN-γ has high diagnostic validity in patients with TPE and can potentially be an excellent biomarker.

Dietary glutamine supplementation improves both Th1 and Th17 responses via CARD11-mTORC1 pathway in murine model of atopic dermatitis

Glutamine (GLN) is considered an immunomodulatory nutrient, while caspase recruitment domain 11 (CARD11) is a susceptibility locus for atopic dermatitis (AD). T-cell antigen receptor (TCR)-stimulated GLN uptake requires CARD11. However, the specific pathogenesis of AD via GLN uptake remains unclear. This study aimed to elucidate the association between dietary GLN supplementation and the CARD11 pathway in the pathogenesis of AD, focusing on T helper type 1 (Th1) and Th17 cell expression in AD. Herein, wild-type (WT) mice with house dust mite epidermal-sensitized skin exhibited increased expression of interferon-gamma (IFN-gamma) and interleukin (IL)-17, whereas CARD11 deficiency impaired Th1 and Th17 responses at the same site. CARD11 is a key mediator of Th1 and Th17 expression in AD. Additionally, we suppressed mammalian target of rapamycin complex 1 (mTORC1) signaling, downstream of CARD11, to underscore the critical role of CARD11 in mediating Th1 and Th17 expression in AD. Further, dietary supplementation of GLN to CARD11−/− mice restored Th1 and Th17 responses, whereas inflammatory expression was reduced in WT mice, and p-CARD11 expression and mTORC1 signaling activity were increased in JPM50.6 cells and CARD11−/− mice. Upon inhibiting the GLN transporter, alanine-serine-cysteine transporter carrier 2 (ASCT2), we observed that the Th1 and Th17 response in AD was reduced. Conclusively, ASCT2-mediated GLN uptake improves the expression of Th1 and Th17 cells via CARD11-mTORC1 signaling pathway in AD, suggesting the potential of glutamine supplementation for AD treatment.

Effect of diode pumping solid state laser with wavelength 589 nm on gene expression of interlukine-2 and interferon-γ in human T-lymphocytes.

To evaluate the impact of laser irradiation of human T-cell lymphocyte in culture to determine the photobiomodulatory effects of such irradiation on gene expression, and the release of growth factors and cytokines. The experimental study was conducted from November 2021 to April 2022 at the Postgraduate Anatomy Laboratory, Mustansiriyah University, Baghdad, Iraq, and comprised human blood samples T-cell lymphocytes were extracted and irradiated, followed by the extraction of ribonucleic acid to assess the effect of laser on gene expression of cytokines, interlukine-2 and interferon-gamma. A 589nm yellow light low-level laser beam from a diode pumping solid state laser was used at doses 30J/cm2, 50J/cm2 and 70J/cm2 for an exposure time of 15 minutes. Reverse transcriptase-polymerase chain reaction was used to measure gene expression. Data was analysed using GraphPad Prism 9. Cellular gene expression of the mediators changed significantly in response to laser irradiation at a variety of dosage parameters, and these effects depended on the wavelength and radiation exposure (p<0.05). The laser light may photobiomodulate the production of cytokines by human cells in vitro.

Unlocking the potential: Cinnamomum cassia (L.) D. Don aqueous extracts as natural defenders against MSRV in largemouth bass aquaculture

Aquatic viral diseases pose significant threats to aquaculture, necessitating the exploration of effective preventive measures. In this study, we evaluated the antiviral potential of Cinnamomum cassia (L.) D. Don (CCD) aqueous extracts against Micropterus salmoides rhabdovirus (MSRV) in largemouth bass aquaculture. Initially, twelve plant extracts were screened for antiviral activity against MSRV, revealing that CCD extracts at 100 mg/L concentration significantly inhibited MSRV proliferation in vitro. Further dose-response analysis revealed a concentration-dependent antiviral effect of CCD extracts, with higher concentrations demonstrating increased efficacy against MSRV. Subsequent investigations focused on the protective effects of CCD extracts on MSRV-infected cells. At a concentration of 63 mg/L, CCD extracts significantly reduced MSRV titers, preventing the onset of cytopathic effects (CPE) in cells. Fluorescence and transmission electron microscopy further illustrated the protective effects of CCD extracts on cellular morphology. MSRV-infected cells treated with CCD exhibited preserved cell morphology akin to normal cells, contrasting with the typical apoptotic features observed in untreated infected cells. Additionally, dietary supplementation of CCD extracts in largemouth bass demonstrated notable benefits. Fish fed with CCD extracts exhibited increased survival rates when infected with MSRV, with a 26 % higher survival rate compared to the virus-only group. Furthermore, CCD supplementation enhanced the expression of interferon-gamma (IFNγ), indicating improved immune responses in largemouth bass. Moreover, immersion of CCD extracts in water showed partial inhibition of horizontal MSRV transmission, effectively reducing viral spread and mitigating disease risks in fish populations. These findings highlight the potential of CCD aqueous extracts as feed additives for combating MSRV in aquaculture, showcasing their promising application prospects in disease prevention and control strategies.

NOSTRIN is involved in benign prostatic hyperplasia via inhibition of proliferation, oxidative stress, and inflammation in prostate epithelial cells.

Benign prostatic hyperplasia (BPH) is a common disease among older men characterized by non-malignant proliferation of epithelial cells and inflammation. Nitric oxide synthase traffic inducer (NOSTRIN) is a pleiotropic regulator of endothelial cell function and signaling and exerts anti-inflammatory, anti-proliferation, and modulating nuclear factor-kappa B (NF-κB) signaling effects. Its expression and function in BPH tissues and prostate epithelial cells are unknown. The study aims to investigate the expression and functions of NOSTRIN in BPH, and its possible molecular mechanism. The BPH model was constructed in male Institute of Cancer Research (ICR) mice using 5 mg/kg/day testosterone propionate (TP) for 30 days, and the model was evaluated by detecting prostate index, prostate epithelial thickness, and prostate-specific antigen (PSA) expression. Dihydrotestosterone (DHT, 10 nM)-induced in vitro model of human prostate epithelial cells (RWPE-1) was established. We generated lentivirus-harboring human NOSTRIN. The mRNA expression was detected by real-time quantitative polymerase chain reaction (PCR) assay; the protein expression or localization was detected by western blot assay, immunohistochemistry, or immunofluorescence staining. Cell proliferation was assayed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and 5-ethynyl-2'-deoxyuridine (EdU) staining. Reactive oxygen species (ROS) production was observed by dihydroethidium staining. Nitric oxide (NO) and malondialdehyde (MDA) levels and superoxide dismutase (SOD) activity were detected using commercial kits. Enzyme-linked immunosorbent assay (ELISA) was used to determine levels of interleukin 1 beta (IL1B), interleukin 6 (IL6), interferon gamma (IFNG), and tumor necrosis factor (TNF). NOSTRIN expression was significantly inhibited in the TP-induced ICR mouse BPH model and DHT-induced model of RWPE-1 proliferation. Protein expression of the BPH-related and proliferation markers PSA and proliferating cell nuclear antigen (PCNA) was suppressed in NOSTRIN-overexpressing RWPE-1 cells exposed to DHT. NOSTRIN overexpression notably inhibited the RWPE-1 cell proliferation in vitro, as evidenced by MTT and EdU staining. NOSTRIN overexpression significantly decreased the expression of cell cycle-related proteins cyclin dependent kinase 4 (CDK4) and cyclin D1 (CCND1) in vitro. The production of ROS, NO, and lipid peroxidation products MDA was inhibited by NOSTRIN overexpression in vitro, while the SOD activity was increased. NOSTRIN overexpression reduced the mRNA expression of inflammatory mediator nitric oxide synthase 2 (NOS2) and inhibited the mRNA expression and secretion of pro-inflammatory cytokines IL1B, IL6, IFNG, and TNF in vitro. The mechanistic studies revealed an increased phosphorylation of NF-κB p65 in vivo and in vitro. Remarkably, NOSTRIN overexpression notably inhibited the protein expression of phospho-NF-κB p65 in vitro. NOSTRIN is involved in BPH by inhibiting proliferation, oxidative stress, and inflammation in prostate epithelial cells. These functions may act through the inhibition of NF-κB signaling.

Effects of oligomeric proanthocyanidins on growth performance, antioxidant capability, immunity, liver and intestinal health of Channa argus

To determine the effects of proanthocyanidins on the growth, antioxidant capability, immunity, intestinal morphology, intestinal microbiota, and liver health of Channa argus (10.12 ± 0.12 g), four experimental diets with various oligomeric proanthocyanidin (OPC) concentrations (0, 200, 400, and 800 mg/kg) were fed to triplicate groups for 8 weeks. The final body weight (FBW), weight gain (WG), feeding rate (FER), protein efficiency ratio (PER), and specific growth rate (SGR) were significantly improved in the 400 and 800 mg/kg OPC-supplemented diets (P < 0.05). The superoxide dismutase (SOD), glutathione-S-transferase (GST), glutathione (GSH), catalase (CAT), and glutathione peroxidase (GSH-Px) activities in the liver and intestines were increased significantly, whereas the malondialdehyde (MDA) content exhibited the opposite pattern (P < 0.05). The GST, Cu/Zn superoxide dismutase (Cu/Zn SOD), nuclear factor erythroid 2-related factor 2 (Nrf2), NAD(P)H quinone dehydrogenase 1 (NQO-1), and heme oxygenase 1 (HO-1) mRNA expression levels in the intestines and liver were significantly increased, whereas kelch-like ECH-associated protein 1 (Keap1) expression showed the opposite pattern (P < 0.05). Significant increases were noted in the serum lysozyme (LYS), immunoglobulin M (IgM), complement 3 (C3), and complement 4 (C4) levels, and significant decreases were noted in the serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bile acid (TBA), and total cholesterol (TC) levels (P < 0.05). The interferon gamma (IFN-γ), interleukin-8 (IL-8), interleukin-1 beta (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) mRNA expression levels in the liver and intestines were significantly decreased, whereas the expression of interleukin-10 (IL-10) showed the opposite trend (P < 0.05). Compared with the control diet, the 400 and 800 mg/kg OPC-supplemented diets markedly improved liver and intestinal morphology. With increasing OPC concentration, the relative abundance of the beneficial bacteria, namely, Firmicutes increased, whereas the opposite trend was observed for the potential pathogenic bacteria, namely, Proteobacteria, Actinobacteria, Chlamydiae, Bacteroidetes, Cyanobacteria, and Acidobacteria. Therefore, dietary OPC supplementation facilitates growth performance and enhances antioxidant capability and immunity, as well as improves liver health, intestinal morphology, and microbiota composition of C. argus, of which the optimal level is 400 mg/kg.

Cathepsin B-Activatable Bioactive Peptide Nanocarrier for High-Efficiency Immunotherapy of Asthma.

Asthma, a chronic respiratory disease closely associated with inflammation, presents ongoing treatment challenges. IALLIPF (le-Ala-Leu-Leu-Ile-Pro-Phe) is one of millet prolamins peptides (MPP) which shows anti-oxidant bioactivity by reducing the production of reactive oxygen species (ROS). Tryptophan (Trp, W) is an amino acid that has been demonstrated to possess anti-inflammatory effects. We introduce a novel cathepsin B-activatable bioactive peptides nanocarrier, PEG-IALLIPF-GFLG-W (MPP-Trp), designed for immunotherapy of asthma. MPP-Trp is synthesized, purified, and its characteristics are investigated by dynamic light scattering (DLS) and transmission electron microscopy (TEM). The yield of nitric oxide (NO) and pro-inflammatory cytokines (TNF-α, IL-6 and IL-1β) are examined to evaluate anti-inflammatory effects of IALLIPF, Trp and MPP-Trp. The immunomodulatory effects of IALLIPF, Trp and MPP-Trp on Th1/Th2 cell populations and cytokines are investigated by flow cytometry, qRT-PCR and ELISA assays. We explore the therapeutic effect of MPP-Trp in the mouse model of asthma by the analysis of lung histology and ELISA. It is necessary to study the biocompatibility of MPP-Trp by CCK8 assay and histopathologic analysis using hematoxylin and eosin (HE) staining. In asthmatic peripheral blood mononuclear cells (PBMCs), IALLIPF, Trp and MPP-Trp are able to significantly alleviate inflammation by inhibiting the yield of nitric oxide (NO) and pro-inflammatory cytokines (TNF-α, IL-6 and IL-1β), especially MPP-Trp. MPP-Trp significantly upregulates Th1 cell levels while notably reducing Th2 cell levels. Furthermore, MPP-Trp effectively elevates the expression and production of interferon-gamma (IFN-γ), an essential cytokine from Th1 cells. Additionally, MPP-Trp markedly diminishes the mRNA expression and levels of key asthma pathogenesis cytokines, such as interleukin-4 (IL-4), interleukin-13 (IL-13), and interleukin-5 (IL-5), in asthma PBMCs. MPP-Trp ameliorates pulmonary pathological alterations and significantly inhibits OVA-induced inflammation in mice with asthma. It has little influence on the cell viability in Asthma-PBMCs treated with various concentrations or durations of MPP-Trp. No pathological changes, including in the heart, liver, spleen, lung, and kidney tissues, are observed in non-sensitized and non-challenged mice treated with MPP-Trp (20 mg/kg). Our research demonstrates that MPP-Trp has immunomodulatory effects on Th1/Th2 cell populations, essential in managing asthma. It considerably alleviates OVA-induced asthma by shifting the immune response towards a Th1-dominant profile, thereby reducing Th2-driven inflammation. Therefore, this novel bioactive peptide nanocarrier, MPP-Trp, holds promise as a candidate for asthma immunotherapy.

Inflammation and keratoconus: A comprehensive bidirectional Mendelian randomization analysis.

An increasing body of evidence supports the involvement of inflammation and immune responses in the occurrence and development of keratoconus (KC). However, the causal relationship between inflammatory factors and KC remains unclear. We employed a 2-way Mendelian randomization (MR) approach to investigate the interaction between KC and inflammatory factors. Instrumental variables for 41 circulating inflammatory regulators and 12 matrix metalloproteinases (MMPs) were selected from genome-wide association studies of European ancestry. Summary statistics for KC were obtained from a genome-wide association study comprising 2116 cases and 24,626 controls of European ancestry. The primary analytical method for assessing causality was the inverse-variance weighted method. Two additional MR methods (MR-Egger and weighted median) were employed to complement the inverse-variance weighted results. In addition, several sensitivity analyses were conducted to evaluate heterogeneity, horizontal pleiotropy, and stability. Our findings indicated that genetically predicted higher levels of macrophage inflammatory protein-1β (odds ratio = 1.126, 95% confidence interval: 1.029-1.232, P = .01) and MMP-13 (odds ratio = 1.211, 95% confidence interval: 1.070-1.371, P = .003) were positively associated with an elevated risk of KC. Conversely, genetically predicted KC was associated with increased levels of interferon-gamma, interleukin-4, and MMP-1. Our current study provided suggestive evidence supporting causal associations of macrophage inflammatory protein-1β and MMP-13 with the risk of KC. In addition, KC appeared to affect the expression of interferon-gamma, interleukin-4, and MMP-1.

Tumoral programmed cell death 1 (PD1) expression in endometrial carcinoma is a prognostic marker for patient outcome

ObjectiveImmune checkpoint inhibitors have recently demonstrated benefit in patients with advanced and recurrent endometrial carcinoma. This retrospective study investigated immune checkpoint molecules in endometrial carcinoma as they pertain to the...

Evaluating the ability of different chaperones in improving soluble expression of a triple-mutated human interferon gamma in Escherichia coli

Human interferon gamma (hIFN-γ) plays a pivotal role as a soluble cytokine with diverse functions in both innate and adaptive immunity. In a previous investigation, we pinpointed three critical amino acid residues, i.e., threonine (T) 27, phenylalanine (F) 29, and leucine (L) 30, on the IFN-γ structure, which are integral to the epitope recognized by anti-IFN-γ autoantibodies. It is crucial to impede the interaction between this epitope and autoantibodies for effective therapy in adult-onset immunodeficiency (AOID). However, the challenge arises from the diminished solubility of the T27AF29L30A mutant in Escherichia coli BL21(DE3). This study delves into a targeted strategy aimed at improving the soluble expression of IFN-γ T27AF29AL30A. This is achieved through the utilization of five chaperone plasmids: pG-KJE8, pKJE7, pGro7, pG-Tf2, and pTf16. These plasmids, encoding cytoplasmic chaperones, are co-expressed with the IFN-γ mutant in E.coli BL21(DE3), and we meticulously analyze the proteins in cell lysate and inclusion bodies using SDS-PAGE and Western blotting. Our findings reveal the remarkable efficacy of pG-KJE8, which houses cytoplasmic chaperones DnaK-DnaJ-GrpE and GroEL-GroES, in significantly enhancing the solubility of IFN-γ T27AF29AL30A. Importantly, this co-expression not only addresses solubility concerns but also preserves the functional dimerized structure, as confirmed by sandwich ELISA. This promising outcome signifies a significant step forward in developing biologic strategies for AOID.

Identification of broad-spectrum B-cell and T-cell epitopes of H9 subtype avian influenza virus HA protein using polypeptide scanning1

The H9N2 subtype avian influenza virus (AIV) hemagglutinin (HA) protein is a major immunogen in which HA1 is a genetic variant and HA2 is relatively conserved. Identifying broad-spectrum antigen epitopes targeting HA1 is crucial for vaccine design and detection. Based on the phylogenetic and serological analyses, we identified 2 antigenic groups and 3 representative viruses: A/chicken/Jiangsu/JY040218C/2019, A/pigeon/Jiangsu/JY020616/2019, and A/chicken/Jiangsu/WX090312/2018. An overlapping peptide library was synthesized using HA1 amino acid sequences of the viruses as templates. Through peptide scanning of the sera against different strains of H9N2 subtype AIV, we identified peptides from 4 regions (H9-2/3, H9-20/21, H9-26, and H9-29/30/31) that demonstrated broad-spectrum reactivity. Immunological assay results demonstrated that H9-21 (219RIFKPLIGPRPLVNGLMGRI239), H9-26 (269SGESHGRILKTDLKMGSCTV289), and H9-30 (309YAFGNCPKYI GVKSLKLAVG329) effectively induced antibody generation and conferred partial protective efficacy against the parent virus JY040218C. The results of lymphocyte proliferation and ELISpot assays indicated that peptides H9-15 (159MRWLTQKNNAYPTQDAQYTN179), H9-22 (229PLVNGLMGRINYYWSVLKP G249), and H9-23 (239NYYWSVLKPGQTLRIKSDGN259) could effectively stimulate the expression of interferon-gamma in peripheral blood lymphocytes of chickens immunized against different strains of H9N2 AIV. Collectively, 5 novel cell epitopes H9-15, H9-22, H9-23, H9-26, and H9-30, including the best B cell epitope H9-26 and the best T cells epitope H9-22, were identified that could be targeted for vaccine design or detection approaches against H9N2 AIVs.

Possible role of intestinal fungal dysbiosis in dectin-1 and cytokines expression in patients with ulcerative colitis.

Dysregulation of cytokines and intestinal mycobiome has been surveyed in the progression of inflammatory bowel diseases (IBDs), including ulcerative colitis (UC) and Crohn's disease (CD). On the other hand, the intestinal fungal flora and its main receptor, Dectin-1, induce immune-derived cytokines. Total 64 individuals comprising 32 patients with UC (case group) and 32 healthy subjects (HS group) were assessed. The type and prevalence of fecal yeast species were determined by deoxyribonucleic acid (DNA) sequencing through polymerase chain reaction (PCR) amplification usingITS4 and ITS5 primers. Furthermore, the ribonucleic acid (RNAs) of IL-4, IL-10, IL-17, IL-22 and IFN-γ were extracted. The expression of Dectin-1 gene was then measured in the excised tissue samples. A higher global fungal load in UC-affectedpatients (75%) was found in comparison with the HS group (25%), especially Candida albicans. Saccharomyces cerevisiae was significantly reduced in the fecal samples of UC-affected patients compared to HS (15.04% vs. 1.93% UC). The expression level of Dectin-1 was significantly elevated in patients with active UC (7.37 ± 0.81) than in patients with non-active UC (5.01 ± 77.25) and healthy controls (0.97 ± 0.24) (p < 0.05). The expression levels of IL-4, IL-10, especially both IL-17 and IL-22, were higher in the active UC group compared to the HS group (p = 0.0101, p = 0.0155, p < 0.0001, p < 0.0001, respectively). Similar expression level of IL-4, IL-10, IL-17, IL-22 (p > 0.999) and lower expression of interferongamma (IFN-γ) (p = 0.0021) were found in the non-active UC group compared to the HS group. A significant weak to moderate correlation was detected between Dectin-1 and IL-17 (r = 0.339, p = 0.019), as well as Dectin-1 and IL-22 (r = 0.373, p = 0.015). Furthermore, the expression levels of Dectin-1, IL-17 and IL-22 displayed significant associations with disease activity (p < 0.001, p = 0.029 and p = 0.003, respectively), regardless of the participant group. The current study revealed a possible role for intestinal fungi to promote colonic inflammation and increase UC activity through Dectin-1 stimulation. A positive correlation was detected between intestinal fungal richness with UC susceptibility and activity. IL-4 and IL-10 were associated with disease activity. Besides, the expression levels of Dectin-1, IL-17 and IL-22 were independently associated with disease activity.

Immunomodulatory Synthetic Glycocluster Molecule Prevents Melanoma Growth In Vivo.

Triacedimannose (TADM) is a synthetic trivalent acetylated glycocluster and a transmembrane macrophage activator independent of the mannose receptor. TADM induces Th1-type immune responses and suppresses Th2-type cytokines in acute and chronic allergic inflammation models in vivo. We, therefore, wanted to test whether TADM could also facilitate anti-tumour tissue responses similar to what has been observed for the immune checkpoint inhibitors, such as anti-PD-1 and anti-CTLA-4. A syngeneic mouse melanoma model was selected since metastatic melanoma has been successfully targeted by checkpoint inhibitors in the clinic. TADM inhibited the growth of B16 mouse melanoma tumours at levels comparable to an anti-PD-1 antibody. TADM-treated tumours encompassed significantly more apoptotic cells as measured by TUNEL staining, and interferon-gamma (IFN-γ) expression was increased in the spleens of TADM-treated mice compared to untreated controls. TADM-treated mice also demonstrated increased Ly6 C low monocytes and neutrophils in the spleens. However, TADM-treated tumours showed no discernible differences in infiltrating immune cells. TADM can alone suppress the growth of melanoma tumours. TADM likely activates M1 type macrophages, type N1 neutrophils, and CD8+ and Th1 T cells, suppressing the type 2 immune response milieu of melanoma tumour with a strong type 1 immune response.

Disease Severity and Cytokine Expression in the Rhinovirus-Induced First Wheezing Episode.

Wheezing children infected with rhinovirus (RV) have a markedly increased risk of subsequently developing recurrencies and asthma. No previous studies have assessed the association between cytokine response and the severity of acute illness in the first wheezing episode in children infected with RV. Forty-seven children treated both as inpatients and as outpatients infected with RV only, aged 3-23 months, with severe first wheezing episodes were recruited. During acute illness, peripheral blood mononuclear cells (PBMCs) were isolated and stimulated with anti-CD3/anti-CD28 in vitro. A multiplex ELISA was used to quantitatively identify 56 different cytokines. The mean age of the children was 17 months, 74% were males, 79% were hospitalized, and 33% were sensitized. In adjusted analyses, the inpatient group was characterized by decreased expressions of interferon gamma (IFN-γ), interleukin 10 (IL-10), macrophage inflammatory protein 1 alpha (MIP-1α), RANTES (CCL5), and tumor necrosis factor-alpha (TNF-α) and an increased expression of ENA-78 (CXCL5) compared to the outpatient group. The cytokine response profiles from the PBMCs were different between the inpatient and outpatient groups. Our results support that firmly controlled interplay between pro-inflammatory and anti-inflammatory responses are required during acute viral infection to absolve the initial infection leading, to less severe illness.

Inhibition of interferon gamma impairs induction of experimental epidermolysis bullosa acquisita.

Epidermolysis bullosa acquisita (EBA) is a muco-cutaneous autoimmune disease characterized and caused by autoantibodies targeting type VII collagen (COL7). The treatment of EBA is notoriously difficult, with a median time to remission of 9 months. In preclinical EBA models, we previously discovered that depletion of regulatory T cells (Treg) enhances autoantibody-induced, neutrophil-mediated inflammation and blistering. Increased EBA severity in Treg-depleted mice was accompanied by an increased cutaneous expression of interferon gamma (IFN-γ). The functional relevance of IFN-γ in EBA pathogenesis had been unknown. Given that emapalumab, an anti-IFN-γ antibody, is approved for primary hemophagocytic lymphohistiocytosis patients, we sought to assess the therapeutic potential of IFN-γ inhibition in EBA. Specifically, we evaluated if IFN-γ inhibition has modulatory effects on skin inflammation in a pre-clinical EBA model, based on the transfer of COL7 antibodies into mice. Compared to isotype control antibody, anti-IFN-γ treatment significantly reduced clinical disease manifestation in experimental EBA. Clinical improvement was associated with a reduced dermal infiltrate, especially Ly6G+ neutrophils. On the molecular level, we noted few changes. Apart from reduced CXCL1 serum concentrations, which has been demonstrated to promote skin inflammation in EBA, the expression of cytokines was unaltered in the serum and skin following IFN-γ blockade. This validates IFN-γ as a potential therapeutic target in EBA, and possibly other diseases with a similar pathogenesis, such as bullous pemphigoid and mucous membrane pemphigoid.

A CTCF-binding site in the Mdm1-Il22-Ifng locus shapes cytokine expression profiles and plays a critical role in early Th1 cell fate specification

Cytokine expression during Tcell differentiation is a highly regulated process that involves long-range promoter-enhancer and CTCF-CTCF contacts at cytokine loci. Here, we investigated the impact of dynamic chromatin loop formation within the topologically associating domain (TAD) in regulating the expression of interferon gamma (IFN-γ) and interleukin-22 (IL-22); these cytokine loci are closely located in the genome and are associated with complex enhancer landscapes, which are selectively active in type 1 and type 3 lymphocytes. In situ Hi-C analyses revealed inducible TADs that insulated Ifng and Il22 enhancers during Th1 cell differentiation. Targeted deletion of a 17bp boundary motif of these TADs imbalanced Th1- and Th17-associated immunity, both invitro and invivo, upon Toxoplasma gondii infection. In contrast, this boundary element was dispensable for cytokine regulation in natural killer cells. Our findings suggest that precise cytokine regulation relies on lineage- and developmental stage-specific interactions of 3D chromatin architectures and enhancer landscapes.

High Efficacy of Green Synthesized Silver Nanoparticles for Treatment of Toxoplasma Gondii Infection Through Their Immunomodulatory, Anti-Inflammatory, and Antioxidant Potency.

The present experimental survey designed to green synthesis, characterization, as well as in vitro and in vivo anti-Toxplasma gondii activity of silver nanoparticles (SLN) green synthesized by Lupinus arcticus extract. SLN were green synthesized based on the reducing by L. arcticus extract through the precipitation technique. In vitro lethal effects of SLN on T. gondii tachyzoites, infectivity rate, parasites inside of the human macrophage cells (THP-1 cells), nitric oxide (NO) triggering, and iNOS and interferon gamma (IFN-γ) expression genes were evaluated. In vivo, after establishment of toxoplasmosis in BALB/c mice via T. gondii ME49 strain, mice received SLN at 10 and 20mg/kg/day alone and combined to pyrimethamine at 5mg/kg for 14 days. SLN exhibited a spherical form with a size ranging from 25 to 90nm. The 50% inhibitory concentration (IC50) value of SLN and pyrimethamine against tachyzoites was 29.1 and 25.7µg/mL, respectively. While, the 50% cytotoxic concentration (CC50) value of SLN and pyrimethamine against THP-1 cells was 412.3µg/mL and 269.5µg/mL, respectively. SLN in combined with pyrimethamine obviously (p < 0.05) decreased the number and size of the T. gondii cysts in the infected mice. The level of NO, iNOS and IFN-γ genes was obviously (p < 0.001) upregulated. SLN obviously (p < 0.05) decreased the liver level of oxidative stress and increased the level of antioxidant factors. The findings displayed the promising beneficial effects of SLN mainly in combination with current synthetic drugs against latent T. gondii infection in mice. But we need more experiments to approve these findings, clarifying all possible mechanisms, and its efficiency in clinical phases.