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Abstract Background The interferon‐γ (IFN‐γ) signaling pathway is activated in systemic lupus erythematosus (SLE). This study aimed to assess the causal association between IFN‐γ, IFN‐γ receptor 1 (IFN‐γR1), and IFN‐γR2 and SLE using a bidirectional Mendelian randomization (MR) design. Methods Genetic instruments for exposure to IFN‐γ, IFN‐γR1, and IFN‐γR2 were derived from a large genome‐wide association study (GWAS) that included a sample size of 3301 participants. Instrumental variables for SLE were selected from another independent GWAS analysis comprising 5201 cases and 6099 controls with European ancestry. Bidirectional two‐sample MR was performed using inverse variance weighting, MR‐Egger regression, and weighted median methods. A series of sensitivity analyses were conducted to assess the robustness of the results. Results The inverse variance weighting showed that IFN‐γ had a positive causal association with the risk of SLE (odd ratio [OR] = 1.24, 95% confidence interval [CI]: 1.03–1.47, p = 0.018). IFN‐γR2 levels were not associated with SLE risk after adjustment for multiple comparisons (OR = 0.85, 95% CI: 0.73–0.99, p = 0.034). No genetic association was also detected between IFN‐γR1 and SLE (OR = 0.97, 95% CI: 0.79–1.19, p = 0.768). Evidence from bidirectional MR did not support reverse causality. The weighted median regression also showed directionally similar estimates. Conclusion Higher levels of IFN‐γ are significantly associated with an increased risk of SLE, providing insights into the pathogenesis of SLE.

Abstract Background Evading immune surveillance is necessary for tumor metastasis. Thus, there is an urgent need to better understand the interaction between metastasis and mechanisms of tumor immune evasion. In this study, we aimed to clarify a novel mechanism that link tumor metastasis and immunosuppression in the development of esophageal squamous cell carcinoma (ESCC). Methods The expression of melanoma‐associated antigen C3 (MAGE‐C3) was detected using immunohistochemistry. Transwell assays were used to evaluate the migration and invasion ability of esophageal squamous cell carcinoma (ESCC) cells. Metastasis assays in mice were used to evaluate metastatic ability in vivo. Lymphocyte‐mediated cytotoxicity assays were performed to visualize the immune suppression function on tumor cells. RNA sequencing was performed to identify differentially expressed genes between MAGE‐C3 overexpressing ESCC cells and control cells. Gene ontology (GO) enrichment analyses was performed to identify the most altered pathways influenced by MAGE‐C3. The activation of the interferon‐γ (IFN‐γ) pathway was analyzed using Western blotting, GAS luciferase reporter assays, immunofluorescence, and flow cytometry. The role of MAGE‐C3 in the IFN‐γ pathway was determined by Western blotting and immunoprecipitation. Furthermore, immunohistochemistry and flow cytometry analysis monitored the changes of infiltrated T cell populations in murine lung metastases. Results MAGE‐C3 was overexpressed in ESCC tissues. High expression of MAGE‐C3 had a significant association with the risk of lymphatic metastasis and poor survival in patients with ESCC. Functional experiments revealed that MAGE‐C3 promoted tumor metastasis by activating the epithelial‐mesenchymal transition (EMT). MAGE‐C3 repressed antitumor immunity and regulated cytokine secretion of T cells, implying an immunosuppressive function. Mechanistically, MAGE‐C3 facilitated IFN‐γ signaling and upregulated programmed cell death ligand 1 (PD­L1) by binding with IFN‐γ receptor 1 (IFNGR1) and strengthening the interaction between IFNGR1 and signal transducer and activator of transcription 1 (STAT1). Interestingly, MAGE‐C3 displayed higher tumorigenesis in immune‐competent mice than in immune‐deficient nude mice, confirming the immunosuppressive role of MAGE‐C3. Furthermore, mice bearing MAGE‐C3‐overexpressing tumors showed worse survival and more lung metastases with decreased CD8+ infiltrated T cells and increased programmed cell death 1 (PD‐1)+CD8+ infiltrated T cells. Conclusion MAGE‐C3 enhances tumor metastasis through promoting EMT and protecting tumors from immune surveillance, and could be a potential prognostic marker and therapeutic target.

Objective To observe the gene single nucleotide polymorphisms (SNPs) of interferon-ω1 IFNGR and interferon-γ receptor (IFNGR) in self-limiting hepatitis B virus ( HBV) infection and chronic HBV infection patients. Methods PCR-RFLP was used to detect four SNP sites of IFN-ω1 and four sites of IFNGR in 206 cases of patients with self-limiting HBV infection (control group) and 210 cases of patients with chronic HBV infection (case group). The geno­type and allele frequency of the two groups were compared. Results The rs9376267 gene loci CT genotype frequencies of IFNGR in the case group and control group were respectively 48. 4% and 60.3% (P=0.012). No statistically significant difference was found between the two groups in the genotype and allel frequency of other SNP sites of IFN-ω1 and IFNGR genes. Conclusion The CT genotype frequency on IFNGR rs9376267 site in patients with self-limiting HBV infection was higher than that of the patients with chronic HBV infection. [ABSTRACT FROM AUTHOR]

The effect of Pam3CSK4, a Toll-like receptor 2 ( TLR2) ligand, on interferon-γ ( IFN-γ) -induced nitric oxide ( NO) production in mouse vascular endothelial END-D cells was studied. Pre-treatment or post-treatment with Pam3CSK4 augmented IFN-γ-induced NO production via enhanced expression of an inducible NO synthase ( iNOS) protein and mRNA. Pam3CSK4 augmented phosphorylation of Janus kinase 1 and 2, followed by enhanced phosphorylation of signal transducer and activator of transcription 1 ( STAT1) at tyrosine 701. Subsequently, the enhanced STAT1 activation augmented IFN-γ-induced IFN-regulatory factor 1 expression leading to the iNOS expression. Pam3CSK4 also induced the activation of p38 and subsequent phosphorylation of STAT1 at serine 727. A pharmacological p38 inhibitor abolished the augmentation of IFN-γ-induced NO production by Pam3CSK4. Surprisingly, Pam3CSK4 enhanced a physical association of MyD88 and IFN-γ receptor. Together, these findings suggest that Pam3CSK4 up-regulates IFN-γ signalling in vascular endothelial cells via the physical association between MyD88 and IFN-γ receptor α, and p38-dependent serine 727 STAT1 phosphorylation. [ABSTRACT FROM AUTHOR]

Abstract: Although significant effects of interferon-γ (IFN-γ) on glial cells are well documented, information on the expression level and localization of glial IFN-γ receptors (IFN-γ-R) in the human central nervous system (CNS) is sparse. To examine the glial expression of IFN-γ-R in the human CNS, immunohistochemistry and quantitative analyses were performed on Alzheimer disease hippocampus, Parkinson disease substantia nigra, amyotrophic lateral sclerosis spinal cord and corresponding areas from non-neurological cases. Almost all IFN-γ-R-positive (IFN-γ-R+) cells corresponded to GFAP-positive (GFAP+) astrocytes, while none of IFN-γ-R+ cells corresponded to IBA1-positive (IBA1+) microglia or MBP-positive (MBP+) oligodendrocytes in these neurological cases. We observed a similar pattern of glial IFN-γ-R expression in non-neurological cases. Also, we quantitatively analyzed the IFN-γ-R expression by cultured human glial cells using immunocytochemistry and reverse transcription polymerase chain reaction (RT-PCR). In contrast to in vivo results, almost all IFN-γ-R+ cells were IBA1+ in microglial cultures, GFAP+ in astrocytic cultures and MBP+ in oligodendrocytic cultures. Moreover, no significant difference in IFN-γ-R mRNA expression was found for these glial cell types by RT-PCR. These results suggest that the microglial and oligodendrocytic expression levels of IFN-γ-R are much lower than the astrocytic expression levels in the human CNS in vivo, whereas all three types of glial cells constitutively express IFN-γ-R when cultured in vitro. [ABSTRACT FROM AUTHOR]

Abstract: Astrocytes and microglia become activated in a broad spectrum of inflammatory neurodegenerative diseases. Activated microglia are widely believed to be the principal source of inflammation-induced neuronal degeneration in these disorders. To investigate the neurotoxic potential of human astrocytes, we exposed them and human astrocytic U-373 MG cells to a variety of inflammatory stimulants. We then assessed the effects of their supernatants on human SH-SY5 cells. When astrocytes and U-373 MG cells were stimulated with interferon (IFN)-γ (150U/ml), their supernatants significantly reduced SH-SY5Y cell viability. Other powerful inflammatory stimulants such as lipopolysaccharide (0.5μg/ml), tumor necrosis factor-α (10ng/ml) and interleukin-1β (10ng/ml), alone or in combination, were without effect. These combinations were also unable to enhance the IFN-γ effect. The induced cytotoxicities were reversed by JAK inhibitor I, a potent and specific inhibitor of JAKs. This result indicates that the neurotoxic effect was proceeding through the IFN-γ receptor (IFNGR)-JAK-STAT intracellular pathway. To establish that the IFNGR is expressed on both cultured astrocytes and U-373 MG cells, we performed RT-PCR on total RNA extracts to identify a specific IFNGR product. We showed the protein product on these cultured cells by immunocytochemistry using an antibody to IFNGR. Finally, using human postmortem material, we showed sharp upregulation of the IFNGR on activated astrocytes in affected areas in Alzheimer''s disease, Parkinson''s disease, amyotrophic lateral sclerosis, and multiple sclerosis. These findings suggest that activated astrocytes may become neurotoxic when stimulated by IFN-γ and may therefore exacerbate the pathology in a spectrum of neurodegenerative diseases. [Copyright &y& Elsevier]

Abstract: It has been shown that herpes simplex virus type 1 (HSV-1) blocks specific immune responses by various mechanisms. Cell lines infected with HSV-1 for instance show a severe impairment of the interferon-gamma (IFN-γ)-induced signal transducer and activator of transcription 1 (STAT1) signaling pathway. Thus, we examined the influence of HSV-1 infection on IFN-γ signal transduction in mature dendritic cells (mDCs). In this study, we report the down-regulation of the IFN-γ receptor α chain (IFNGR1) at the mRNA level in HSV-1 infected mDCs. As a consequence, the expression of the IFNGR1 subunit on the cell surface of the infected cell was strongly reduced. Furthermore, we were able to show the inhibition of STAT1 phosphorylation following HSV-1 infection in mDCs, while protein levels of STAT1 remained constant. As a direct downstream effect of STAT1 phosphorylation, the activation of the transcription factor IRF-1 was also clearly inhibited and could no longer be induced by the addition of IFN-γ. Additional experiments using a virus strain lacking the vhs gene suggested that the mutant virus is more sensitive to IFN-γ as STAT1 signaling was inhibited to a lesser extent. Infection with a UV-inactivated, replication incompetent virus did not influence the STAT1 signaling pathway at all. In conclusion, we show that HSV-1 blocks IFN-γ signaling in mDCs, which requires viral gene expression and involves the viral protein vhs. [Copyright &y& Elsevier]

Genetic polymorphism is a difference in DNA sequence among individuals, groups, or populations that give rise to different forms. Differences in DNA sequences that occur naturally in a population. Single nucleotide substitutions, insertions and deletions of nucleotides and repetitive sequences (microsatellites) are all examples of polymorphism. The position at which such a sequence difference is found is a polymorphic site. A single nucleotide substitution is called a single nucleotide polymorphism (SNP). SNPs can occur in coding parts of the gene. If they result in genetic code change, amino-acid polymorphism would occur. The heterodimeric IFN-γ receptor ( IFNGR) complex was made up of two receptor subunits including IFNGR-1 and IFNGR-2. There exist five dbSNP alleles in IFNGR1 exon region and six dbSNP alleles in IFNGR2. Some researchers had found that the greatest risk of the development of systemic lupus erythematosus (SLE) be detected in the individuals who had the Met14/Val14 genotype or the combination of IFNGR1 Met14/Val14 genotype and IFNGR2 Gln64/Gln64 genotype in Japanese patients. So we aimed to assess the association between two polymorphisms within the IFNGR gene (A88G and A839G) and SLE in Chinese patients. This study included 154 patients with SLE and 159 unrelated healthy controls. We examined the IFNGR genotype by the reverse transcription-polymerase chain reaction (RT-PCR)-single-strand conformation polymorphism method, RT-PCR-restriction fragment length polymorphism method and DNA sequencing. Genotype frequencies between SLE patients and controls were compared and relationship between genotype frequencies and clinical manifestations of SLE were evaluated. We found that IFNGR2 Arg64/Arg64 genotype decrease the risk of SLE (OR = 2.326, 95% CI 1.181–4.581, Fisher P = 0.015), and the same as IFNGR2 Arg64/Arg64 genotype and IFNGR1 Val14/Val14 genotype combination (OR = 2.420, 95% CI 1.206–4.854, Fisher P = 0.013). The allelic frequency of Val14/Met14 is significantly higher in the patients with oral ulcer or thrombocytopenia when compared with patient without these clinical feature (OR = 4.630, 95% CI 1.370–15.640, Fisher P = 0.021; or OR = 6.368, 95% CI 2.009–20.191, Fisher P = 0.003). On the contrary, the allelic frequency of Val14/Val14 is lower in the patients with oral ulcer or thrombocytopenia than those without these clinical feature (OR = 0.216, 95% CI 0.064–0.730, Fisher P = 0.021; or OR = 0.157, 95% CI 0.050–0.498, Fisher P = 0.003). And after data analysis, we also find that the allelic frequency of Gln64/Gln64 is lower in the patients with arthritis when compared with patient without arthritis (OR = 0.369, 95% CI 0.166–0.818, Fisher P = 0.017). We can conclude that the IFNGR polymorphisms (Val14Met and Gln64Arg) are protective in SLE in Chinese patients. We describe a novel association between Val14/Met14 carriage and patients with oral ulcer or thrombocytopenia. [ABSTRACT FROM AUTHOR]

Abstract: Mendelian defects in interferon-γ (IFN-γ) signaling most commonly lead to infection with nontuberculous mycobacteria. Mutations have been identified in the genes encoding IFN-γ-receptor-1, IFN-γ-receptor-2 and Stat-1. Partial and complete deficiencies in signaling are found, leading to parallel clinical, pathological, and cellular phenotypes. These rare defects have led to better molecular and mechanistic understanding of the role of IFN-γ in the human immune system. [Copyright &y& Elsevier]

Interferon (IFN)-γ appears to be an important hair cycle modulator in mice. It is unclear whether it has similar hair growth modulatory functions in human hair follicles.To study whether IFN-γ can be exploited to modulate the growth, pigmentation and/or cycling of organ-cultured human anagen scalp hair follicles, as anin vitroindicator system for how IFN-γ affects human hair growthin vivo. This was correlated with the hair follicle expression patterns of IFN-γ receptorsα andβ. In addition, we wanted to establish a new, simple tool for the rapid experimental induction of catagenin vitro.Normal human scalp hair follicles in the anagen VI stage of the hair cycle were cultured according to the method of Philpottet al., with or without IFN-γ (50–1000 IU mL−1). Hair shaft elongation and pigmentation changes were measured, complemented by quantitative histomorphometry to assess changes in hair follicle cycling (hair cycle score), proliferation (Ki-67), melanogenesis (Masson–Fontana) and apoptosis (TUNEL). IFN-γ receptors were also localized by immunofluorescence and EnVision technique. As transforming growth factor (TGF)-β2 is a recognized key inducer of catagen in human hair follicles, TGF-β2 expression was investigated by tyramide signal amplification and reverse transcription-polymerase chain reaction in anagen hair follicles treated with vehicle (phosphate-buffered saline) or IFN-γ.IFN-γ rapidly inhibited hair elongation in cultured human anagen hair follicles and induced morphological signs of catagen transformation after only 4 days of culture, i.e. faster than with other reported catagen-inducers (e.g. TGF-β2). Proliferation was inhibited, apoptosis was increased and follicular melanogenesis was switched off in hair bulb keratinocytes treatedin situwith IFN-γ. Anagen hair follicles displayed strong IFN-γ receptorα-like immunoreactivity, while the immunoreactivity for IFN-γ receptorβ in the hair matrix was only weak. TGF-β2 immunoreactivity and mRNA transcript levels were enhanced in hair follicles treated with IFN-γ.These data suggest that IFN-γ is a potent catagen inducer in normal human scalp hair follicles, which express cognate receptors, and show that IFN-γ administration offers an excellent tool for experimental catagen induction in organ-cultured human hair follicles. This catagen induction probably occurs at least in part via upregulation of the recognized catagen-stimulatory growth factor TGF-β2. [ABSTRACT FROM AUTHOR]

Interferon γ (IFN-γ) plays an important role in immune response, apoptosis, and anti-tumor activity. Its biological activity depends on expression of IFN-γ receptor (IFN-γR). To address whether increased expression of IFN-γR is associated in vivo with a higher biological response by IFN-γ, we constructed an adenovirus vector including murine IFN-γR (Ad-mIFN-γR). We confirmed the appropriate function of mIFN-γR derived from Ad-mIFN-γR based on the observation of signal transduction and transcription. We also found that elevated expression of mIFN-γR increases sensitivity to recombinant murine IFN-γ (rmIFN-γ) in vitro in target cells. Furthermore, we demonstrated that the growth rate of tumors transfected with Ad-mIFN-γR is suppressed in response to rmIFN-γ in vivo and that such growth suppression is partly due to apoptosis. To our knowledge, this is the first report of adenovirus-mediated IFN-γR gene transfer being effective in augmenting the biological activity of IFN-γ, and the strategy employed in the present study will be useful in studying other kinds of cytokine receptors and applications to gene therapy for cancer and infectious diseases. [Copyright &y& Elsevier]

Tyrosine phosphorylation of STAT1α in eosinophils after IFN-γ stimulation has been shown, but the biological significance of eosinophil STAT1α activation in transmitting the signals through the IFN-γ receptor remains unknown. The purpose of this study is to determine whether STAT1 is involved in the regulation of eosinophils by IFN-γ-IFN-γ receptor interaction. rhIL-3- and rhIL-5-induced eosinophils from CD34+ cells of cord blood on day 28 of culture were used. The cells were washed and further incubated in IL-3- and IL-5-free medium for 48 h. The induced eosinophils constitutively expressed CD69 and lost this expression after a further 48-hour incubation without the cytokines. IFN-γ significantly upregulated CD69 expression on the 48-hour incubated cells. In inhibitory experiments on STAT1, a phosphorothioate oligo antisense DNA against STAT1α was added to IL-3- and IL-5-containing medium from day 15 to day 28 of culture. The oligo DNAs altered neither the expressions of myeloid cell marker CD9 and 13 nor the expression of IFN-γ receptor on the cells. The added STAT1α antisense, but not sense, DNA significantly reduced STAT1α mRNA expression in the cells. The STAT1 antisense also significantly inhibited IFN-γ-induced CD69 expression on the 48-hour incubated eosinophils. In conclusion, these results indicate that IFN-γ induces CD69 expression in the induced eosinophils through STAT1α, suggesting that STAT1α may play a significant role in eosinophil regulation by IFN-γ.Copyright © 2001 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

The purpose of this study was to determine whether interferon-γ (IFN-γ) induced CD69 expression by eosinophil precursors. Eosinophil precursors were induced from CD34+ cord blood cells using recombinant human interleukin-3 (IL-3) and interleukin-5 (IL-5). On day 14 of culture, cells constitutively expressed CD69 and the IFN-γ receptor (IFN-γR). Stimulation with IFN-γ for 24 h did not affect IFN-γR expression by the cells. On the other hand, IFN-γ significantly upregulated CD69 expression by the precursors after 24 h of incubation. A specific JAK2 inhibitor (AG-490) caused a concentration-dependent suppression of IFN-γ-induced CD69 expression by the precursors. In conclusion, these results indicate that IFN-γ induces CD69 expression by eosinophil precursors via the activation of JAK2.Copyright © 2000 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background: It has been unclear whether or not the interferon-γ receptor (IFN-γR) on human peripheral eosinophils is completely functional. Accordingly, we examined the expression of IFN-γR α- and β-chains and the association of the two chains after binding of IFN-γ to the receptor. Methods: Peripheral blood eosinophils were obtained from 8 patients (bronchial asthma, n=6, and hypereosinophilic syndrome, n=2), and expression of the α- and β-chain was investigated by flow cytometry with specific antibodies. Expression of mRNA for the α- and β-subunits was also investigated by the reverse transcriptase-polymerase chain reaction. Results: Eosinophils from all the patients were positive for both the α- and β-chains by flow cytometry. In addition, mRNA for both chains was detected by the polymerase chain reaction. Furthermore, coprecipitation of the α- and β-chains was only found after eosinophil stimulation with IFN-γ. Conclusions: Human peripheral eosinophils apparently express both the α- and β-chains of the IFN-γR, and the two chains may only form the receptor complex after ligand binding. [ABSTRACT FROM AUTHOR]

Summary Regulatory T cells (T regs ) are a barrier to anti-tumor immunity. Neuropilin-1 (Nrp1) is required to maintain intratumoral T reg stability and function but is dispensable for peripheral immune tolerance. T reg -restricted Nrp1 deletion results in profound tumor resistance due to T reg functional fragility. Thus, identifying the basis for Nrp1 dependency and the key drivers of T reg fragility could help to improve immunotherapy for human cancer. We show that a high percentage of intratumoral NRP1 + T regs correlates with poor prognosis in melanoma and head and neck squamous cell carcinoma. Using a mouse model of melanoma where Nrp1-deficient ( Nrp1 –/– ) and wild-type ( Nrp1 +/+ ) T regs can be assessed in a competitive environment, we find that a high proportion of intratumoral Nrp1 –/– T regs produce interferon-γ (IFNγ), which drives the fragility of surrounding wild-type T regs , boosts anti-tumor immunity, and facilitates tumor clearance. We also show that IFNγ-induced T reg fragility is required for response to anti-PD1, suggesting that cancer therapies promoting T reg fragility may be efficacious. [ABSTRACT FROM AUTHOR]