Expression Levels Of IFN-γ Research Articles

Immunoexpression of interferon-gamma in the interdental gingiva of chronic periodontitis patients with interferon-gamma (+874A/T) rs62559044 polymorphism.

Interferon-gamma (IFNγ), which is involved in inflammation, has an essential role in the pathogenesis of chronic periodontitis (CP). The association of IFNγ +874A/T gene polymorphisms with a higher incidence of CP has been reported previously. We investigated the immunoexpression of IFNγ in human gingiva from CP patients with IFNγ +874A/T gene polymorphisms compared to a healthy group. Biopsies from interdental gingiva (n=60) were obtained from CP patients (n=38) and individuals who had clinically healthy gingiva (n=22). The specimens were classified according to genotypes of IFNγ +874A/T gene polymorphism using amplification-refractory mutation system-polymerase chain reaction (ARMS-PCR) (10AA, 18AT, and 10TT). After tissue processing and slide preparation, we evaluated the expression levels of IFNγ in the interdental gingiva using the immunohistochemistry technique. Values were compared between two study groups by Mann-Whitney and Kruskal-Wallis analysis. P<0.05 was considered the level of significance. There was a noticeable increase in the immunoexpression of IFNγ in the gingival epithelium, fibroblasts, and capillaries of gingival tissue from CP patients compared to the control group. We found different levels of IFNγ expression in different parts of the gingival epithelium. The highest levels were found in the middle regions of the gingival epithelium in patients with TT genotypes. Immune cells and vascular endothelium of gingiva in the AA group also showed increased expression of IFNγ. Results suggest that IFNγ +874A/T gene polymorphism may change the expression level of IFNγ and subsequently the progression of CP.

404.7: Study on the Regulatory Effect of iPS-Derived Exosomes on the Local Immune Microenvironment in DCD Kidney Transplanted Rats

Introduction: Ischemia reperfusion injury (IRI) of donation After Cardiac death (DCD) transplanted kidney is the key factor of acute renal injury, acute and chronic rejection of renal transplantation and transplanted kidney disease.The latest researches show that mesenchymal stem cells (MSCs) have the function of protecting damaged tissues and organs and strong immune regulation. Induced pluripotent stem (iPS) cells have higher proliferative capacity and exosome secretion potential than MSCs. In addition to the functions of stem cells, exosomes have the unique ability to cross the blood-brain barrier and deliver remotely, which means that exosomes have better advantages in clinical disease treatment compared with the therapeutic potential of stem cells. Therefore, the main purpose of this study is to clarify the role of IPS derived exosomes in protecting the IRI of DCD transplanted kidney and regulating its local immune microenvironment. Methods: The iPS cells were passaged to the third passage and massively expanded. After 48 hours of culture, 100 mL of cell culture supernatant was collected and exosomes were extracted from the supernatant by the kit method, and the morphology and surface membrane marker proteins of the exosomes were identified. The SD rats were established with DCD renal transplantation model and 10, 50, and 100 μg of exosomes were infused into that SD rats respectively, and a PBS control group was set at the same time. On the 3rd, 5th, 7th, 14th and 28th days after transplantation, the content of cytokines in the serum of the recipient rats, the proportion of lymphocyte subsets CD4+CD25+Foxp3+Tregs in the spleen and the relative expression of Foxp3 mRNA were detected. The proliferation of renal tubular cells was detected by PCNA immunofluorescence. The pathological changes of renal tissue were determined by HE staining, and the apoptosis of renal tubular epithelial cells was detected by Tunel staining. The infiltration of inflammatory cells was detected by immunohistochemistry. The expression changes of regulatory T cells in renal tissue and macrophage phenotypic characteristics were analyzed by flow cytometry. Results: The experimental amount of exosomes was successfully obtained, and the morphology was observed by transmission electron microscope. The exosomes were round or oval membranous vesicles with uniform size, and the edges were clearly visible. The TSG101, CD63, and CD9 proteins were highly expressed on the exosome membrane surface by western-blot. A DCD rat kidney transplantation model was successfully established. The expression levels of IL-12, IL-2, IFN-γ, and TNF-α in the serum of each group were increased to varying degrees from the 3rd day after operation. On the 3rd and 7th days, the levels of IL-12, IL-2, IFN-γ, and TNF-α in the 100 μg exosome infusion group were significantly lower than those in the 10 μg, 50 μg and PBS groups (P<0.05), and the expression level of IL-10 in the 100 μg exosome infusion group was significantly higher than 10μg, 50μg and PBS groups (P<0.05). On the 28th day, the expression level of IL-10 in the 100 μg exosome infusion group was significantly higher than other groups (P<0.01). Compared with other groups, the CD4+CD25+Tregs and CD4+CD25+Foxp3+Tregs accounted for the highest proportion of CD4+T cells in the spleen of the 100μg infusion group (P<0.05). On the 28th day, the expression level of Foxp3 mRNA in the 100 μg infusion group was significantly higher than that in the 50 μg group (P<0.05). The stainings of PCNA, HE and Tunel showed that SD rats infused with 100μg exosomes could significantly reduce the pathological damage, inflammatory response and apoptosis of renal tubular epithelial cells in the transplanted kidney, promote the proliferation of renal tubular epithelial cells, and have a protective effect on IRI kidney. On the 28th day, the expression of CD4+CD25+Foxp3+Tregs cells in the transplanted kidney tissue was significantly increased, which further promoted the differentiation of macrophages into the immunomodulatory M2 subset. Conclusions: The infusion of iPS-derived high-dose exosomes is involved in the protection of the DCD kidney transplant during IRI process, and alleviates the acute and chronic injury of the DCD kidney transplant, and promotes the formation of the local immunosuppressive microenvironment in the DCD kidney transplantation. The National Natural Science Foundation of China (No.81900686).

An Integrated Analysis of Inflammatory Endotypes and Clinical Characteristics in Chronic Rhinosinusitis with Nasal Polyps.

ObjectiveChronic rhinosinusitis with nasal polyps (CRSwNP) is mainly characterised by type 1 (T1), type 2 (T2) and type 3 (T3) inflammatory endotypes. However, correlations between inflammatory endotypes and clinical features in CRSwNP have not been demonstrated sufficiently. This study aimed to determine the endotype-phenotype associations in CRSwNP.MethodsClinical data of 31 control subjects and 106 CRSwNP patients were analysed. Interferon (IFN)-γ (T1), Charcot-Leyden crystal galectin (CLC) (T2) and Interleukin (IL)-17A (T3) were used as biomarkers to identify the inflammatory endotypes.ResultsThe mRNA expression level of IFN-γ was positively correlated with IL-17A (r = 0.817; P < 0.0001). Headache/facial pain (P = 0.039) was associated with T1 endotype. Smell loss (P = 0.025) was associated with T2 endotype. Purulent rhinorrhea (P = 0.001) was associated with T3 endotype. Atopy (P = 0.030), asthma (P = 0.005) and recurrence (P = 0.022) were more frequent in T2 endotype. Total Symptom Scores (TSS) of T2 (P < 0.001) and T3 (P = 0.009) endotype were higher than non-T2 and non-T3, respectively. Sino Nasal Outcome Test-22 (SNOT-22) total scores of T3 (P = 0.054) endotype were higher than non-T3.ConclusionIdentifications of endotype-phenotype associations are useful in clinical diagnoses and targeted therapies for patients with CRSwNP.

Expression Pattern of Inflammatory and Anti-Inflammatory Cytokines and Key Differential Transcription Factors in Peripheral Blood Mononuclear Cells of Iranian Coronavirus Disease 2019 Patients with Different Disease Severity.

The purpose of this research was to investigate the gene expression levels of inflammatory cytokines interferon (IFN)γ, tumor necrosis factor (TNF)α, interleukin (IL)1β, IL2, IL6, IL8, and IL17, and anti-inflammatory cytokines IL4, IL10, IFNα, and IFNβ, as well as relevant key transcription factors (TFs), including GATA3, PU1, NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells), IRF3 (interferon regulatory factor 3), BCL6 (B cell lymphoma 6 protein), FOXP3 (forkhead box P3), RORγt, and T-bet (T-box expressed in T cell) in Iranian patients with moderate and severe coronavirus disease 2019 (COVID-19). Fifty-six patients with COVID-19, and 25 healthy controls (HCs) age and sex matched were investigated. Based on the interim guidance of COVID-19 from the World Health Organization, the patients were classified into 33 moderate and 23 severe patients with COVID-19. The gene expression levels of cytokines and relevant TFs were quantified in peripheral blood mononuclear cells by quantitative real-time polymerase chain reaction (qRT-PCR). The gene expression levels of TFs RoRγ (RAR-related orphan nuclear receptor γ), NF-κB, and T-bet were significantly higher in patients with COVID-19 compared with HCs. Furthermore, the gene expression levels of cytokines, including IL2, IFNγ, IL6, TNFα, IL1β, IL8, and IL17, were significantly higher in patients with COVID-19 than HCs. However, there was a significant increase for IL6, TNFα, and IL17 in severe compared with moderate patients with COVID-19. Finally, The Spearman correlation analysis revealed a significantly positive correlation for IL6 and TNFα, IL6 and IL2, IL6, IFNγ, and IL2 and IFNγ. These data suggest that expression of IL6, TNFα, and IL17 as well as the synergic effect of elevated values of IL2 and IFNγ should be considered in the treatment and management of patients with severe COVID-19.

Effect of a combination of oxaliplatin and fluoropyrimidine derivative molecule S-1 on expression levels of IL-9 and CysLTR-1 in a rat model of colon cancer

Purpose: To investigate the effect of a combination of oxaliplatin and fluoropyrimidine derivative molecule S-1 on the expression levels of IL-9 and CysLTR-1 in rats with colon cancer.&#x0D; Methods: One hundred and eighty healthy Spraque Dawley rats were randomly assigned to groups A (healthy control), B (model), and C (treatment) with 60 rats in each group. Western blotting was used to determine contents of CysLTR-1 and related pathway proteins, while the levels of IL-9 and IFN-γ were evaluated by enzyme-linked immunosorbent assay (ELISA). Cell Counting Kit-8 (CCK-8) was used to determine cell proliferation, while cell apoptosis was assessed using flow cytometry.&#x0D; Results: Tumor volume was larger and tumor diameter was longer in group B than in group C (p &lt; 0.05). The expression levels of CysLTR-1, LTD4, β-catenin, COX-2, IL-9 and IFN-γ were significantly up-regulated in groups B and C, relative to group A (p &lt; 0.05). Cell proliferation was also significantly faster in groups B and C than in group A, while apoptotic rate was lower (p &lt; 0.05).&#x0D; Conclusion: Combined use of OX and S-1 inhibits CysLTR-1 expression level in colon cancer rats. Furthermore, it down-regulates IL-9 expression, thereby inhibiting the proliferation and apoptosis of tumor cells. Therefore, the combination treatment has potentials for development as a therapeutic strategy for the management of colon cancer.&#x0D; Keywords: Oxaliplatin; Fluoropyrimidine derivative molecule; Colon cancer; CysLTR-1

Effects of Lactiplantibacillus plantarum 19-2 on immunomodulatory function and gut microbiota in mice.

This study aims to evaluate the effects of Lactiplantibacillus plantarum 19-2 (L. plantarum 19-2) on mice treated with the alkylating agent cyclophosphamide (CTX). Our findings show that L. plantarum 19-2 restored the spleen and thymus index and the number of white blood cells and lymphocytes% in CTX treated mice. Serum immunoglobulin levels in CTX-treated mice were increased by L. plantarum 19-2. In addition, as compared to the model group, L. plantarum 19-2 upregulated the content of SIgA, while L. plantarum 19-2 regulates the mRNA and protein expression levels of GATA-3, T-bet, IFN-γ, and IL-4 in small intestinal tissues, which adjusted mucosal barriers, structural status, and the balance of Helper T-cell 1 and Helper T-cell 2. Lactiplantibacillus plantarum 19-2 regulated the distribution of intestinal flora in mice, promoting the growth of Bacteroides and Proteobacteria. In addition, L. plantarum 19-2 inhibited the growth of several harmful bacteria, including Actinobacteria and Firmicutes.

Integrated strategy of network analysis prediction and experimental validation to elucidate the possible mechanism of compound Turkish gall ointment in treating eczema

BackgroundCompound Turkish gall ointment (CTGO) has a long history of being widely used as a folk medicine in Xinjiang for the treatment of eczema. CTGO is currently in the pre-investigational new drug application stage, but its pharmacological mechanisms of action have not yet been clarified.MethodsFirst, a sensitive and reliable ultra-high performance liquid chromatography-Q exactive hybrid quadrupole-orbitrap high-resolution accurate mass spectrometry (UHPLC-Q-Orbitrap HRMS) technique was established. Second, an integrative strategy of network analysis and molecular docking based on identified and retrieved ingredients was implemented to investigate the potential targets and pathways involved in the treatment of eczema with CTGO. Finally, Sprague–Dawley (SD) rats with eczema were prepared to verify the predicted results. The skin conditions of the rats were observed, evaluated, and scored. Skin tissues were observed by hematoxylin–eosin (HE) staining, and the levels of serum interferon-γ (IFN-γ) and interleukin-4 (IL-4) were determined by enzyme-linked immunosorbent assay (ELISA). The expression levels of toll-like receptor 4 (TLR4), nuclear factor kappa-B p65 (NF-κB p65), interleukin-1β (IL-1β), and tumor necrosis factor-α (TNF-α) were detected by real-time quantitative polymerase chain reaction (RT-qPCR).ResultsA total of 29 compounds were identified. We found 38 active components and 58 targets for the treatment of eczema, which included 118 signaling pathways related to inflammation, immunity, and apoptosis. CTGO significantly improved the skin surface and histopathological characteristics of eczema-affected rats, downregulated the expression of IL-4, TLR4, NF-κB (p65), IL-1β, and TNF-α, and upregulated the expression level of IFN-γ.ConclusionWe predicted and validated our prediction that CTGO may be used to treat eczema by affecting the TLR4/NF-κB signaling pathway, which provides guidance for future experimental studies.

Immune enhancement effects of inactivated vaccine against extracellular products of Aeromonas caviae AC-CY on crucian carp

Aeromonas caviae is a zoonotic pathogen that can cause disease in aquatic organisms and mammals, including humans, and it is widespread in nature, especially in freshwater environments. Previous research has reported that extracellular products (ECPs) secreted by pathogens during growth are effective protective antigens that can induce the host immune response and protect the host from pathogens. However, little is known about how ECPs enhance immunity. Here, we prepared extracellular products by the cellophane plate method, determined the total protein concentration, and analysed the protein composition of the extracellular products by SDS–PAGE. Subsequently, their enzyme activity and pathogenicity were evaluated separately. Crucian carp were randomly divided into four groups to receive formalin-inactivated A. caviae vaccine (FKC), ECPs mixed with the same amount of Freund's complete adjuvant, the same amount of ECPs mixed with an equal volume of A. caviae inactivated vaccine (FKC + ECPs), sterile PBS alone via intraperitoneal injection. On Days 7, 14, 21, and 28 after immunization, the expression levels of IgM, SOD, and CAT and the lysozyme (LYS) activity in the serum were detected by ELISA, and the relative expression levels of the TNF-α, IFN-γ, IL-1β, and IL-10 genes in the liver, kidney, spleen, intestine, and gills were measured by qPCR. The extracellular products generated five clearly visible protein bands and exhibited lipase, protease, amylase, DNase and lysozyme but no urease or lecithinase activities. In addition, the median lethal doses of A. caviae and ECPs to crucian carp were 411.64 μg/fish and 1.6 × 105 CFU/mL, respectively. Compared with those of the control group, the IgM, SOD, and CAT contents and serum LYS activity were significantly increased in the experimental groups, and the qRT–PCR results showed that the relative expression levels of TNF-α, IFN-γ, IL-1β, and IL-10 genes in the liver, kidney, spleen, and intestine were significantly increased after injection immunization. In addition, the relative immune protection rates of the three experimental groups were 60%, 65%, and 45%, all of which were significantly higher than those of the control group. Collectively, our findings show that the extracellular products of A. caviae can be used as a vaccine to significantly improve the immune level of crucian carp and have obvious anti-infection ability. This may represent a promising approach to prevent and control infection by A. caviae and provides strong theoretical support for the development of new inactivated vaccines.

E.faecium QH06 alleviates TNBS-induced colonic mucosal injury in rats

To investigate the effect of Enterococcus faecium QH06 on TNBS-induced ulcerative colitis (UC) in rats and explore the mechanisms in light of intestinal flora and intestinal immunity. Thirty-six male Wistar rats were randomized equally into control group, UC model group, and E.faecium QH06 intervention group. The rats in the latter two groups were subjected to colonic enema with 5% TNBS/ethanol to induce UC, followed by treatment with intragastric administration of distilled water or E.faecium QH06 at the dose of 0.21 g/kg. After 14 days of treatment, the rats were examined for colon pathologies with HE staining. The mRNA and protein expression levels of IL-4, IL-10, IL-12, and IFN-γ in the colon tissues were detected using RT-qPCR and ELISA, and the expression of TLR2 protein was detected with immunohistochemistry and ELISA. Illumina Miseq platform was used for sequencing analysis of the intestinal flora of the rats with bioinformatics analysis. The correlations of the parameters of the intestinal flora with the expression levels of TLR2 and cytokines were analyzed. The rats with TNBS- induced UC showed obvious weight loss (P < 0.01) and severe colon tissue injury with high pathological scores (P < 0.01). The protein expression levels of IFN-γ, IL-12, and TLR2 (P < 0.01) and the mRNA expression levels of IFN-γ, IL-12 and IL-10 (P < 0.05) were significantly increased in the colon tissues of the rats with UC. Illumina Miseq sequence analysis showed that in UC rats, the Shannon index (P < 0.05) ACE (P < 0.01)and Chao (P < 0.05) index for the diversity of intestinal flora both decreased with a significantly increased abundance of Enterobacteriaceae (P < 0.01) and a lowered abundance of Burkholderiaceae (P < 0.05). Compared with the UC rats, the rats treated with E. faecium QH06 showed obvious body weight gain (P < 0.05), lessened colon injuries, lowered pathological score of the colon tissue (P < 0.05), decreased protein expressions of IFN- γ, IL- 12, and TLR2 and mRNA expressions of IFN- γ and IL-12 (P < 0.01 or 0.05), and increased protein expressions of IL- 4 (P < 0.05). The Shannon index ACE (P < 0.05) and Chao (P < 0.05) index of intestinal microflora were significantly increased, the abundance of Enterobacteriaceae was lowered and that of Burkholderiaceae and Rikenellaceae was increased in E.faecium QH06- treated rats (P < 0.01 or 0.05). Correlation analysis showed that IFN-γ was positively correlated with the abundance of Enterobacteriaceae, and IFN-γ was negatively correlated with the abundance of Prevotellaceae, Desulfovibrionaceae, norank_o_Mollicutes_RF39 and Clostridiales_vadinBB60_group; TLR2 was negatively correlated with Clostridiales_vadinBB60_group, norank_o_Mollicutes_RF39 and Prevotellaceae. E.faecium QH06 can alleviate TNBS-induced colonic mucosal injury in rats, and its effect is mediated possibly by increasing the abundance of SCFA-producing bacteria such as Prevotellaceae and inhibiting abnormal immune responses mediated by TLR2.

Dairy Cows Experimentally Infected With Bovine Leukemia Virus Showed an Increased Milk Production in Lactation Numbers 3-4: A 4-Year Longitudinal Study.

Bovine leukemia virus (BLV) is widespread in global cattle populations, but the effects of its infection on milk quantity and quality have not been clearly elucidated in animal models. In this study, 30 healthy first-lactation cows were selected from ≈2,988 cows in a BLV-free farm with the same criteria of parity, age, lactation number, as well as milk yield, SCS, and composition (fat, protein, and lactose). Subsequently, these cows were randomly assigned to the intervention (n = 15) or control (n = 15) group, and reared in different cowsheds. Cows in the intervention group were inoculated with 1 × phosphate-buffered solution (PBS) resuspended in peripheral blood mononuclear cells (PBMC) from a BLV-positive cow, while the controls were inoculated with the inactivated PBMC from the same individual. From June 2016 to July 2021, milk weight (kg) was automatically recorded by milk sensors, and milk SCS and composition were originated from monthly performed dairy herd improvement (DHI) testing. Fluorescence resonance energy transfer (FRET)–qPCR and ELISA showed that cows in the intervention group were successfully infected with BLV, while cows in the control group were free of BLV for the entire period. At 45 days post-inoculation (DPI), the numbers of whole blood cells (WBCs) (P = 0.010), lymphocytes (LYMs) (P = 0.002), and monocytes (MNCs) (P = 0.001) and the expression levels of IFN-γ (P = 0.013), IL-10 (P = 0.031), and IL-12p70 (P = 0.008) increased significantly in the BLV infected cows compared to the non-infected. In lactation numbers 2–4, the intervention group had significantly higher overall milk yield (P < 0.001), fat (P = 0.031), and protein (P = 0.050) than the control group, while milk SCS (P = 0.038) and lactose (P = 0.036) decreased significantly. Further analysis indicated that BLV infection was associated with increased milk yield at each lactation stage in lactation numbers 3–4 (P = 0.021 or P < 0.001), but not with SCS and milk composition. Together, this 4-year longitudinal study revealed that artificial inoculation of BLV increased the milk yield in cows in this BLV challenge model.

Blockage of Galectin-Receptor Interactions Attenuates Mouse Hepatic Pathology Induced by Toxoplasma gondii Infection.

Toxoplasma gondii (T. gondii), one of the most important Apicomplexan protozoa, causes toxoplasmosis in human throughout the world. Galectin (Gal)-9 triggers a series of immune events via binding to its receptors, including T cell immunoglobulin and mucin-containing molecule 3, CD137, CD44, and protein disulfide isomerase. To examine the regulatory role of galectin-receptor interactions in anti-toxoplasmic activities, C57BL/6 mice were infected with T. gondii RH strain and intraperitoneally injected with alpha (α)-lactose to block the interactions of galectins and their receptors. Heatmaps showed upregulated values for Gal-9 and CD137 in the livers of T. gondii-infected mice and T. gondii-infected mice treated with α-lactose. Compared with T. gondii-infected mice, T. gondii-infected mice treated with α-lactose showed significantly increased survival rate, decreased tissue parasite burden, attenuated liver histopathology, increased mRNA expression levels of CD137, IFNγ, IL-4, and IL-10 in the liver, and increased Gal-9 mRNA expression level in the spleen. Correlation analysis showed that significant positive correlations existed between the mRNA expression levels of Gal-9 and CD137, Gal-9 and IFNγ, as well as between CD137 and IFNγ in the liver and spleen of T. gondii-infected mice; between CD137 and IFNγ in the liver of T. gondii-infected mice treated with α-lactose. In addition, blockage of galectin-receptor interactions showed enhanced M2 macrophage polarization in the liver of T. gondii-infected mice. Our data indicate that Gal-9-CD137 interaction may play an important role in T. gondii proliferation and liver inflammation in mice during acute T. gondii infection, through regulating T cell and macrophage immune responses.

Alteration of the Inflammatory and Anti-Inflammatory Cytokine Profiles of Peripheral Blood Mononuclear Cell in Crohn's Disease Patients after Following up.

Background:Crohn’s disease (CD) has a chronic course, which its recurrence varies widely among different patients. In this study we prospectively analyzed blood samples of 19 CD patients. Alteration in transcription of inflammatory and anti-inflammatory cytokines was analyzed compared with household members after three month follow up.Methods:CD patients were diagnosed based on clinical symptoms, endoscopic and histopathologic characteristics. Nineteen CD patients and their households were evaluated from Jun 2019 to Feb 2021 at Tehran university hospitals. CD activity score, biological, clinical and demographic data of the patients were recorded at two time point intervals. Bacteriological tests were done using aerobic and anaerobic blood cultures. To investigate transcriptional alterations, peripheral blood mononuclear cells (PBMCs) were isolated using Ficol centrifugation method and relative quantitative real-time PCR was done to determine the expression level of IFN-γ, TNF-α, IL10, and FOXP3 cytokines.Results:Our results showed a correlation between fecal calprotectin level (709.8 ± 554.6), C-reactive protein concentration (18.1 ± 15.9), and erythrocyte sedimentation rate (30.4 ± 17.9) with disease activity (Flare/remission). IL10 and Foxp3 anti-inflammatory gene’s expression were significantly (P = 0.003 for IL10 and P = 0.008 Foxp3) higher during the flare and remission in patients with active disease respectively. Bacteriological examination showed infection with Streptococcus spp. and Clostridium spp. in two CD patients during flares, which was correlated with upregulation and down-regulation of IL10, TNF-α, IFN-γ and FOXP3 proteins, respectively.Conclusion:Occurrence of bacteremia, and higher amount of CAP, CRP and ESR are correlated with higher level of transcription for inflammatory cytokines, which could effectively reflect the disease activity. Raise in FoxP3 transcription proposed change in Treg sub-population in PBMC or its activity during the CD remission phase.

Moxibustion regulates T-regulatory/T-helper 17 cell balance by modulating the microRNA-221/suppressor of cytokine signaling 3 axis in a mouse model of rheumatoid arthritis

ObjectiveRheumatoid arthritis (RA) progression is associated with the balance of T-regulatory (Treg) and T-helper 17 (Th17) cells, while the role of microRNAs (miRs) in regulating Treg/Th17 cell balance has not been clarified. This study aimed to assess whether moxibustion could regulate Treg/Th17 cell balance by modulating the miR-221/suppressor of cytokine signaling 3 (SOCS3) axis in the RA mouse model. MethodsA mouse model of collagen-induced arthritis (CIA) was established in male DBA/1J mice. Twenty-two days after CIA induction, the mice received daily treatment with moxibustion for 12 times. Pathological scores were assessed according to the levels of synovial hyperplasia. The expression levels of cytokines interleukin (IL)-1β, IL-6, tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), IL-17 and IL-10 were analyzed in serum by enzyme-linked immunosorbent assay. The cluster of differentiation 4 (CD4+) splenocytes was analyzed by fluorescence-activated cell sorting. The expression levels of RA-related miRs and target genes were subsequently detected, and the target of miR-221 was confirmed by the dual-luciferase reporter assay. ResultsIt was revealed that moxibustion treatment decreased the pathological scores and downregulated the expression levels of IL-1β, IL-6, TNF-α, IFN-γ and IL-17, while upregulated the expression level of IL-10. The Treg/Th17 cell balance was regulated by moxibustion treatment. The expression level of miR-221 was suppressed by moxibustion treatment. Furthermore, SOCS3 was found as the direct target of miR-221, which mediated the function of moxibustion by regulating the Treg/Th17 cell balance. ConclusionMoxibustion therapy regulated the Treg/Th17 cell balance by modulating the miR-221/SOCS3 axis in the RA mouse model.

CXCR2 antagonist SB332235 mitigates deficits in social behavior and dysregulation of Th1/Th22 and T regulatory cell-related transcription factor signaling in male BTBR T+ Itpr3tf/J mouse model of autism

Autism spectrum disorders is a complex neurodevelopmental disorder characterized by abnormal social interaction, defective communication, repetitive and stereotyped patterns of behaviors or interests. The BTBR T+ Itpr3tf/J (BTBR) inbred mice are generally used as a model for ASD, display a range of autistic phenotypes. Recent studies suggest that the CXCR2 antagonist is crucial for targets in the treatment of inflammatory and neurodegenerative diseases. In this study, we investigated the potential effects of the CXCR2 antagonist SB332235 on sociability behaviors, marble burying, and self-grooming, we also explored the treatment of SB332235 on Th1 (IFN-γ, Stat1, and T-bet), Th22 (IL-22, TNF-α, and AhR), and T regulatory (Treg, IL-10, Helios and Foxp3) production in CD4+ T cells in male BTBR and C57BL/6 (C57) mice in spleen. We also investigated the effects of SB332235 on IFN-γ, IL-10, IL-22, T-bet, AhR, and Foxp3 mRNA expression levels in the brain tissues. The SB332235-treated mice significantly improve behavioral abnormalities in BTBR mice. In addition, SB332235 administration causes a significantly decreases in IFN-γ, Stat1, T-bet, IL-22, TNF-α, and AhR, and increases in IL-10, Foxp3 and Helios production CD4+ T cells in BTBR mice. We further observed that SB332235 downregulated IFN-γ, IL-10, IL-22, T-bet, and AhR, and upregulated IL-10 and Foxp3 mRNA expression in the brain tissues. Our findings demonstrated that SB332235 treatment attenuated behavior deficits, through inhibiting Th1/Th22 and upregulating Treg cell-related transcription factors signaling pathway. Therefore, CXCR2 antagonist administration may be a promising therapeutic agent to attenuate behavior deficits via its anti-inflammatory effect.

Synergistic effect of acupoint injection and moxibustion or catgut embedding or acupuncture on symptoms and expression of Th1/Th2 related cytokines in nasal mucosa of rats with allergic rhinitis

To observe the effect of moxibustion, catgut embedding and acupuncture on allergic symptoms and expression of interferon-γ (IFN-γ) and interleukin-4 (IL-4) of nasal mucosa in rats with allergic rhinitis (AR) based on acupoint injection, so as to explore their synergistic effect and related mechanism in relieving AR. SD rats (half male half female) were randomly divided into normal control, model, acupoint injection (AI), AI+moxibustion, AI+catgut embedding and AI+acupuncture groups, with 8 rats in each group. The AR model was established by intraperitoneal injection of ovalbumin suspension (once every other day for 7 times), and intranasal drop of 0.5% ovalbumin solution (once daily for 7 days). After successful modeling, rats of the AI group received injection of a mixture solution of equal proportion of 1% lidocaine, dexamethasone and transfer factor into "Yingxiang" (LI20) and "Yintang" (EX-HN3) once every 4 days, 4 times altogether. Mild moxibustion or catgut embedment or manual acupuncture was applied to bilateral "Feishu" (BL13) and "Zusanli" (ST36). Both moxibustion (20 min every time) and acupuncture (with the needles retained for 30 min every time) were conducted once daily for 14 times, and catgut embedding was conducted once a week, twice altogether based on acupoint injection. The rats' nasal allergic reaction score (symptom score, 1-3 points) was given according to the times of nose scratching and sneezing, and the running nose state in 30 min, and histopathological changes of nasal mucosa were observed by H.E. staining. The expression levels of IFN-γ and IL-4 in the nasal mucosa were detected by immunohistochemistry and Western blot, separately. Compared with the normal group, the symptom score and the expression of IL-4 positive cells and protein in nasal mucosa were significantly increased (P<0.05), while the expression of IFN-γ positive cells and protein were considerably decreased in the model group (P<0.05). In comparison with the model group, the symptom score and the expression of IL-4 positive cells and protein were obviously decreased (P<0.05), while the expression of IFN-γ positive cells and protein were remarkably increased in the 4 treatment groups (P<0.05). The effects of AI+moxibustion, AI+catgut embedment, AI+acupuncture were signi-ficantly superior to those of simple AI in up-regulating the expression of IFN-γ positive cells and protein and in down-regulating the expression of IL-4 positive cells and protein (P<0.05). Both the symptom score and the expression of IL-4 were notably lower in the AI+moxibustion group than in the AI+catgut embedment and AI+acupuncture groups (P<0.05), whereas the expression of IFN-γ was apparently higher in the AI+moxibustion group than in the other 3 treatment groups (P<0.05). No significant differences were found between the AI+catgut embedment and AI+acupuncture groups in the levels of symptom score, IFN-γ and IL-4 expressions (P>0.05). Moxibustion or catgut embedment or acupuncture and AI have a synergistic effect in relieving symptoms of AR rats, which may be related to their function in regulating the expression levels of nasal IFN-γ and IL-4 proteins. The therapeutic effect of moxibustion is obviously superior to those of both acupuncture and catgut embedment.

Trichinella spiralis infection ameliorates the severity of Citrobacter rodentium-induced experimental colitis in mice

Trichinellosis is a food-borne zoonotic parasitic disease that causes serious harm to human health and the pig breeding industry. However, there are reports that Trichinella spiralis (T. spiralis) infection can treat autoimmune diseases, including enteritis and experimental autoimmune encephalitis (EAE). However, research on the mechanism of T. spiralis infection in infectious enteritis has not been fully elucidated. Therefore, this experiment used Citrobacter rodentium (C. rodentium) to induce colitis in mouse models and explored its underlying mechanisms. In this experiment, a total of 72 C57BL/6 mice were randomly divided into four groups. Experimental mice in the TS and TS + CR groups were orally inoculated with individual T. spiralis larvae. At 21 days postinfection (dpi) with T. spiralis, experimental animals in the CR and TS + CR groups were inoculated by orogastric gavage with C. rodentium. The control group received PBS only. The results indicated that the weight loss and macroscopic and microscopic colon damage of mice in the TS + CR group were significantly decreased compared with those observed in the CR group. The results of flow cytometry showed that the expression levels of IL-4, IL-10 and CD4+CD25+Foxp3+ Tregs were increased (P < 0.05), while the expression levels of IFN-γ, IL-12 and IL-17 were decreased in the spleens and MLNs of the TS + CR experimental mice compared with the colitis model mice. ELISA results revealed that the TS + CR group not only elicited a strong IgG1 response (P < 0.01) but also a low level of IgG2a response (P < 0.05) relative to the CR group. The above results demonstrated that prior exposure of mice to T. spiralis infection ameliorated the severity of C. rodentium-induced infectious colitis.

Immunotherapy Mechanism of Esophageal Squamous Cell Carcinoma with the Effect of STK11/AMPK Signaling Pathway.

This study was aimed at exploring the mechanism of serine threonine protein kinase 11 (STK11)/Adenosine 5′-monophosphate-activated protein kinase (AMPK) signaling pathway after immunotherapy for esophageal squamous cell carcinoma (ESCC), providing basic information for the clinical treatment of ESCC. In this study, tissue specimens from 100 patients with ESCC who underwent surgical treatment in Taizhou People's Hospital (group A) and 20 patients with recurrent or metastatic ESCC who received second-line immunotherapy (group B) were collected. The real-time fluorescent quantitative polymerase chain reaction (PCR) (RT-qPCR) technology was used to detect the expression levels of STK11, interferon-γ (IFN-γ), interleukin 6 (IL-6), and vascular endothelial growth factor (VEGF) in the tissues. The immunohistochemical staining was used to detect the positive expression levels (PELs) of STK11 and AMPKα in the tissues, and immunofluorescence staining was used to detect the PELs Teff cells (CD3 and CD8), Treg cells (CD4 and FOXP3), and neutrophils (CD68 and CD163). RT-qPCR results showed that the expression levels of STK11 and IFN-γ in group A were obviously lower, and those of IL-6 and VEGF were much higher in contrast to group B (P < 0.05). The results of immunohistochemical staining showed that the number of STK11- and AMPKα-positive staining cells in group A was dramatically less than that in group B (P <0.05). The results of immunofluorescence staining revealed that the number of positive staining cells for Teff cells, Treg cells, and neutrophils in group A was also less dramatically than that in group B (P <0.05). In summary, immunotherapy can play a therapeutic effect on ESCC by regulating STK11/AMPK pathway and immune cell infiltration.

Analysis of Serum Levels of IFN-γ, IL-4, and TNF-α in Patients with Cervicitis Complicated by HPV Infection and Their Clinical Significance.

In order to investigate the expression levels of serum IFN-γ, IL-4, and tumor necrosis TNF-α in patients with cervicitis complicated with human papillomavirus (HPV) infection and clinical significance, a retrospective study was conducted on 90 patients with chronic cervicitis complicated by HP V infection who visited our hospital from June 2020 to June 2021, and they are included in the research group. According to the degree of HPV infection, the patients are divided into low-risk HPV type group (n = 65 cases) and high-risk HPV type group (n = 25 cases); 50 patients with cervicitis (without HPV infection) who received treatment in our hospital are selected as control group 1. Fifty healthy women who underwent physical examination are selected as the control group 2. The general data of the two groups of patients during hospitalization are collected, and HPV-DNA, IFN-γ, IL-4, and TNF-α are detected in all patients. For patients with cervicitis complicated by HPV infection, the IFN-indexes in the body are significantly decreased, IL-4 and TNF-αare significantly increased, and with the degree of HPV infection, IFN-γ, IL-4, and TNF-α have high diagnostic performance with HPV infection, and there is a significant correlation between the three, which can be used in cervicitis complicated with HPV infection. It is widely used in the early diagnosis and screening of infected patients.

Heat shock protein 90 is a new potential target of anti-rejection therapy in allotransplantation.

The critical roles of heat shock protein 90 (HSP90) in immune reactions associated with viral infection and autoimmune disease are well known. To date, however, its roles in the alloimmune response and the immunosuppressive effect of HSP90 inhibitors in allotransplantation have remained unknown. The purpose of this study was to examine the therapeutic efficacy of the HSP90 inhibitor 17-DMAG in allotransplantation models. C57BL/6 (H-2b) and BALB/c (H-2d) mice were used as donors for and recipients of skin and heart transplantation, respectively. Treatment with 17-DMAG (daily i.p.) or a vehicle was initiated 3days before transplantation. Immunological outcomes were assessed by histopathological examinations, flow cytometric analysis, quantitative RT-PCR, ELISA, ELISPOT assay, and MLR. 17-DMAG treatment significantly prolonged the survival of both skin and heart allografts. In 17-DMAG-treated mice, donor-reactive splenocytes producing IFN-γ were significantly reduced along with the intragraft mRNA expression level and serum concentration of IFN-γ. Intragraft mRNA expression of cytokines and chemokines associated with both innate and adaptive immunity was suppressed in 17-DMAG-treated group. MLR showed suppression of the donor-specific proliferation of CD4 + T and CD19 + B cells in the spleens of 17-DMAG-treated mice. 17-DMAG treatment also reduced the number of activated NK cells. Furthermore, the treatment lowered the titers of donor-specific antibodies in the serum and prolonged a second skin allograft in mice sensitized by previous skin transplantation. HSP90 inhibition by 17-DMAG can affect various immune responses, including innate immunity, adaptive immunity, and humoral immunity, suggesting its therapeutic potential against acute rejection in allotransplantation.

Skin Interstitial Fluid and Plasma Multiplex Cytokine Analysis Reveals IFN-γ Signatures and Granzyme B as Useful Biomarker for Activity, Severity and Prognosis Assessment in Vitiligo.

BackgroundThe course of vitiligo is unpredictable, with periods of disease flare-ups and prolonged recovery periods. It is essential to establish a biomarker profile as a substitute marker for disease activity to predict disease activity, severity, and prognosis prediction. The use of localized skin interstitial fluid as biomarkers has recently gained interest, but extensive studies of the association between skin interstitial fluid, plasma, and the disease course is lacking. This study aims to evaluate the cytokine expression profiles in the skin and plasma and the utility of the biomarker panel in assessing disease activity, severity, and prognosis in patients with vitiligo.MethodsIn this prospective cohort study, 86 patients and 34 healthy controls were recruited from the outpatient department of a tertiary medical center from March 2019 to September 2021. All patients were of Asian ethnicity. Two independent investigators evaluated disease activity and severity with longitudinal follow-ups for treatment response for a-12 month period. Ultrasensitive multiplex cytokine panel and single-molecule counting technology immunoassays were used to study the cytokine expression in skin interstitial fluid and plasma.ResultsIFN-γ and its’ signature cytokines, including CXCL9, CXCL10, and GzmB, are most highly expressed in the vitiligo patients’ lesion skin interstitial fluid and plasma compared to healthy control. By way of comparison, no significant changes in IL-1β, IL-13, IL-15, IL-17A, IL-18 were observed. Receiver operating characteristic analysis revealed that IFN-γ is the most sensitive and specific marker in predicting disease activity, followed by CXCL10 and GzmB. CXCL-9 was sensitive and specific in diagnosing vitiligo disease severity. The decrease in IFN-γ expression level is positively correlated with the treatment response.ConclusionIFN-γ, CXCL9, CXCL10, and GzmB are highly expressed in vitiligo patients’ lesion skin and plasma and may serve as biomarkers for the clinical activity, severity, and prognosis prediction in vitiligo patients. Among all, IFN-γ exerts the highest predictive value in disease activity and treatment response, supporting the critical role of IFN-γ in the pathogenesis of vitiligo.