Th17 Frequencies Research Articles

P052 IL-12p70 as a potential driver of pathogenic IL-17+IFNγ+ mucosal memory TIGITneg T cells in a subgroup of severe CD patients

Abstract Background Crohn’s disease (CD) is a chronic disease driven by inflammatory memory CD4+ T cells. Deciphering heterogeneity in underlying immune disease is required for predicting the individual patients’ disease course. We have shown that analysis of circulating CD38+ memory CD4+ T cells, which are enriched for gut-homing, identifies therapy-naïve pediatric CD patients with a more severe disease course. At diagnosis, these patients have increased frequencies of circulating CD38+ memory T cells that do not express TIGIT (TIGITneg). TIGIT (T-cell immunoreceptor with immunoglobulin and ITIM domains), is a coinhibitory receptor selectively induced on CD4+ T cells with a memory phenotype. In healthy controls, TIGIT+CD38+ memory T cells have an inhibitory function while TIGITnegCD38+ memory T cells are enriched in inflammatory cells. In our cohort of CD patients, circulating inflammatory TIGITnegCD38+ memory T cells had increased frequencies of pathogenic IFNγ +IL-17+-double-producing or Th1* cells. We hypothesize that in these severe CD patients, CD-related inflammatory cytokines selectively increase frequencies of Th1* TIGITnegCD38+ memory T cells. Methods We cultured peripheral blood memory CD4+ T cells from healthy individuals in vitro with TCR- and co-stimulation together with different cocktails of CD-related inflammatory cytokines during 72h. Results Of all cytokines tested, only cocktails containing IL-12p70 increased TIGITneg cell frequencies within gut-homing CD38+ memory T cells, but not CD38neg memory T cells that are enriched in skin-homing. As hypothesized, IL-12p70-stimulated cultures contained higher frequencies of Th1* cells, expressing both hallmark transcription factors T-bet and RORgt, in TIGITnegCD38+ T cells but not within TIGIT+CD38+ T cells. This selective IL-12p70 responsiveness in TIGITneg cells agreed with the higher expression of the IL-12 receptor b2 subunit in TIGITnegCD38+ compared to TIGIT+CD38+ memory T cells. These data argue that IL12p70 initiates reprogramming of the TIGITnegCD38+ memory T cells to the Th1* phenotype. To assess whether IL-12p70 causes expansion of TIGITnegCD38+ cells or downregulates TIGIT expression on TIGIT+CD38+ cells, we traced fluorescently-labelled TIGIT+ cells during culture. TIGIT+ cells did not downregulate TIGIT, indicating that IL-12p70 increased proliferation of TIGITnegCD38+ cells. Conclusion The finding that IL-12p70 drives expansion of TIGITnegCD38+ memory T cells with a pathogenic Th1* phenotype argues that this cytokine may enforce chronic inflammation in these severe CD patients. These analyses pave the way toward deciphering key actors involved in CD inflammation at an individual patient level, needed to tailor novel IL-12/IL-23 targeting therapeutic strategies.

P053 Ustekinumab decreases circulating Th1, Th17, and MAIT cells in Crohn’s disease

Abstract Background Ustekinumab is a humanized antibody to a p40 subunit shared by the cytokines IL-12 and IL-23, which has been approved for the treatment of Crohn’s disease. While it is presumed to derive clinical efficacy by blocking the ability of these cytokines to stimulate specific T cell subsets, its actual effect upon the human immune system has not been well-described. Methods Peripheral blood mononuclear cells (PBMC) were cryopreserved from 20 Crohn’s patients before and during ustekinumab therapy. Thawed PBMC were then analyzed by flow cytometry for cell surface markers and, after overnight stimulation, intracellular cytokines. Success (n=9) or failure (n=11) of ustekinumab therapy was at the discretion of the treating physician, blinded to flow cytometry data. Results A significant decrease in IL-17A-producing Th17 (p=0.001) and CD161+/TCRVα7.2+ MAIT (p=0.005) cells was seen overall in response to ustekinumab. While no significant effect was seen overall in IFN-γ -producing CD4 (Th1) and CD8 T cells, there was a dichotomy between treatment responders and non-responders, with Th1 cells decreasing in responders (p=0.004) and increasing in non-responders (p=0.03) after ustekinumab initiation. IFN-γ+ CD8 T cells likewise increased in non-responders (p=0.01), although their decrease in responders was less significant (p=0.12). While there was no difference in the pre-treatment frequency of Th17’s or IFN-γ+ T cells between responders and non-responders, MAIT cells were significantly more common in the blood of responders (p=0.03). Conclusion While the IL-23-blocking effect of ustekinumab on Th17 and MAIT cells was seen across all recipients, its IL-12-blocking effect on IFN-γ-producing T cells was restricted to treatment-responsive patients, suggesting that its mechanism of action may differ from newer, IL-23-specific agents. Whether the frequency of MAIT cells before or a rise in IFN-γ-producing T cells during therapy can predict treatment failure warrants additional study.

Delayed Microbial Maturation Durably Exacerbates Th17-driven Asthma in Mice.

Microbial maturation disrupted by early-life dysbiosis has been linked with increased asthma risk and severity; however, the immunological mechanisms underpinning this connection are poorly understood. We sought to understand how delaying microbial maturation drives worsened asthma outcomes later in life and its long-term durability. Drinking water was supplemented with antibiotics on Postnatal Days 10-20. To assess the immediate and long-term effects of delaying microbial maturation on experimental asthma, we initiated house dust mite exposure when bacterial diversity was either at a minimum or had recovered. Airway hyperresponsiveness, histology, pulmonary leukocyte recruitment, flow cytometric analysis of cytokine-producing lymphocytes, and assessment of serum IgG1 (Immunoglobulin G1) and IgE (Immunoglobulin E) concentrations were performed. RT-PCR was used to measure IL-13 (Interleukin 13)-induced gene expression in sequentially sorted mesenchymal, epithelial, endothelial, and leukocyte cell populations from the lung. Delayed microbial maturation increased allergen-driven airway hyperresponsiveness and Th17 frequency compared with allergen-exposed control mice, even when allergen exposure began after bacterial diversity recovered. Blockade of IL-17A (Interleukin 17A) reversed the airway hyperresponsiveness phenotype. In addition, allergen exposure in animals that experienced delayed microbial maturation showed signs of synergistic signaling between IL-13 and IL-17A in the pulmonary mesenchymal compartment. Delaying microbial maturation in neonates promotes the development of more severe asthma by increasing Th17 frequency, even if allergen exposure is initiated weeks after microbial diversity is normalized. In addition, IL-17A-aggravated asthma is associated with increased expression of IL-13-induced genes in mesenchymal, but not epithelial cells.

Selenium Supplementation Alter the Frequency of Th17 but not Th1/Th2 Lymphocytes in Diffuse Large B Cell Lymphoma Patients.

Selenium (Se) is a micronutrient, which has recently been proven to have a positive effect on the immune system of cancer patients, but the underlying mechanism is not clearly defined. In this randomized controlled trial, we evaluated the effect of three-month Se supplementation on the profile of CD4+ T-helper subsets including IFN-γ+/IL-4- Th1, IFN-γ-/IL-4+ Th2, and CD4+IL-17+ Th17 cells in sixteen diffuse large B cell lymphoma (DLBCL) patients at stable remission phase who consumed Se (Se+) compared to the fourteen control patients who did not receive Se (Se-). The frequency of IFN-γ+/IL-4- Th1, IFN-γ-/IL-4+ Th2, and CD4+IL-17+ Th17 lymphocytes was determined using a four-color flow cytometry method. The results revealed that three-month Se supplementation significantly decreased the proportion of CD4+IL-17+ Th17 lymphocytes but not IFN-γ+/IL-4- Th1 and IFN-γ-/IL-4+ Th2 subtypes in DLBCL patients at stable remission. Change in the percentage of IFN-γ+/IL-4- Th1, IFN-γ-/IL-4+ Th2, and CD4+IL-17+ Th17 cells did not significantly differ between Se+ and Se- groups. No positive correlation was observed between changes in different Th subpopulations in both Se+ and Se- groups. Taken together, three-month Se supplementation can reduce the proportion of CD4+IL-17+ Th17 cells in DLBCL patients at stable remission phase. Larger population and longer follow-up of patients is necessary to specify the clinical significance of Se supplementation on the popularity of T-helper cells in DLBCL patients.

TH1/Treg ratio may be a marker of autism in children with immune dysfunction

Evidence suggests a link between autism spectrum disorder (ASD), behavioral symptoms in the context of ASD, and presence of an altered immune function. Several studies have highlighted differences in T-lymphocyte subpopulations, their activation status, and their response to stimulation in children and adults with ASD. These T cell abnormalities have often been associated with more impaired behaviors. However, few studies have attempted to address whether T cell subsets have the potential to serve as biomarkers in ASD. Moreover, although many studies have been performed in Western populations, few (if any) have been performed in Asian populations in mainland China. In this study we used intracellular cytokine flow cytometry to assess the frequencies of CD4+ T-cell subpopulations (T-helper (TH) 1, TH2, TH17, Foxp3+ regulatory T cells (Treg) as well as CD8+, subpopulations of T cytotoxic (TC) 1, TC2, and TC17 in 82 children with ASD and 50 healthy typical developing children from the Second Affiliated Hospital of Kunming Medical University of Yunnan province. To further elucidate immune status cytokine levels were also measured in the plasma and serum using a bead-based cytokine assay. Our results showed that the frequency of circulating Treg cells and the levels of active TGF-β1 in plasma were lower in children with ASD than in healthy controls. In contrast, the frequencies of TH1, TH2, TH17 and TC1 cells were increased. Proinflammatory cytokine levels of TNF-α, IL-4, IL-5 and IL-17A were higher in the plasma of children with ASD compared to typical controls. We also found an association between the severity of behavior impairments in ASD children and altered immune responses as measured using the effector T cell responses and regulatory responses (using Teff/Treg ratios). Higher the Teff/Treg ratios, were associated with more severe problematic behavioral symptoms. Further, the potential biomarker relevance of Teff/Treg ratio was evaluated by the receiver operating characteristics curve. Data suggests that high TH1/Treg cell ratios could also be used as a potential marker for the diagnosis of children with ASD. Overall, our data suggest an imbalance in inflammatory and regulatory immune responses in ASD. Ratios of inflammatory/regulatory cells or cell frequencies such as Teff/Treg cells may be useful biomarkers for children with ASD with immune dysfunction.

Exenatide regulates Th17/Treg balance via PI3K/Akt/FoxO1 pathway in db/db mice

BackgroundThe T helper 17 (Th17)/T regulatory (Treg) cell imbalance is involved in the course of obesity and type 2 diabetes mellitus (T2DM). In the current study, the exact role of glucagon-like peptide-1 receptor agonist (GLP-1RA) exenatide on regulating the Th17/Treg balance and the underlying molecular mechanisms are investigated in obese diabetic mice model.MethodsMetabolic parameters were monitored in db/db mice treated with/without exenatide during 8-week study period. The frequencies of Th17 and Treg cells from peripheral blood and pancreas in db/db mice were assessed. The phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/Forkhead box O1 (FoxO1) pathway in Th17 and Treg cells from the spleens of male C57BL/6J mice was detected by western blotting. In addition, the expression of glucagon-like peptide-1 receptor (GLP-1R) in peripheral blood mononuclear cells (PBMCs) of male C57BL/6J mice was analyzed.ResultsExenatide treatment improved β-cell function and insulitis in addition to glucose, insulin sensitivity and weight. Increased Th17 and decreased Treg cells in peripheral blood were present as diabetes progressed while exenatide corrected this imbalance. Progressive IL-17 + T cell infiltration of pancreatic islets was alleviated by exenatide intervention. In vitro study showed no significant difference in the level of GLP-1R expression in PBMCs between control and palmitate (PA) groups. In addition, PA could promote Th17 but suppress Treg differentiation along with down-regulating the phosphorylation of PI3K/Akt/FoxO1, which was reversed by exenatide intervention. FoxO1 inhibitor AS1842856 could abrogate all these effects of exenatide against lipid stress.ConclusionsExenatide could restore systemic Th17/Treg balance via regulating FoxO1 pathway with the progression of diabetes in db/db mice. The protection of pancreatic β-cell function may be partially mediated by inhibiting Th17 cell infiltration into pancreatic islets, and the resultant alleviation of islet inflammation.

Perturbations of immune landscape in COVID-19 associated mucormycosis.

A rise in secondary fungal infections during the COVID-19 pandemic necessitates a deeper understanding of the associated immunological perturbations. To evaluate the clinical and immunological characteristics observed in patients with COVID-19 associated mucormycosis (CAM) infection. Cases of mucormycosis with or post-COVID-19 infection were compared with cases of acute COVID-19 and convalescent COVID-19. Lymphocyte subsets, cytokines and other laboratory markers were compared between the groups. The frequency of proposed risk factors for CAM was diabetes mellitus (77%), recent history of steroid use (69%) and hypoxia during COVID-19 infection (52%). Iron metabolism was dysregulated in CAM patients with low TIBC and total iron. Further, CAM was accompanied with lymphopenia with drastic reduction in B cell counts; however, plasmablasts were not altered. Further, CAM patients had low immunoglobulin levels and antibodies specific to mucor peptide did not increase in CAM suggesting dysfunction in B-cell response. There was increase in activated effector cytotoxic CD8 T cells and NK cells in CAM compared with COVID-19 infection and healthy controls. Among T helper cells, Tregs were reduced and Th-1 frequency was increased in CAM compared with COVID-19 infection. A distinct cytokine signature was evident in CAM with increase in IL-1β, IFN-γ, IL-6, IL-22, IL-17A, IL-10, IL-2, IL-8, IL-7, IL-21 and GM-CSF. This is the first study on immunophenotyping in CAM suggesting the need for long-term monitoring of B-cell function after SARS-CoV-2 in patients with dysregulated glycaemic control and the possible benefit of therapeutic supplementation with intravenous immunoglobulins in CAM.

Anticeramide Improves Sjögren’s Syndrome by Blocking BMP6-Induced Th1

T-cell dysfunction has been shown to play an important role in the pathogenesis of Sjögren’s syndrome (SS). In recent studies, the increased expression of BMP6 has been reported to be related to SS. However, the roles that BMP6 plays in immune homeostasis in the development of SS as well as the downstream signals activated by BMP6 remain unclear. In this study, we investigated the effects and molecular mechanisms of BMP6 on naive CD4+ T cells, showing that BMP6 could upregulate interferon (IFN)–γ secretion from CD4+ T cells through a ceramide/nuclear factor–κB pathway, with no effect on T-cell activation or proliferation. Moreover, an in vivo study showed that anticeramide treatment (myriocin) for an SS animal model (NOD/LtJ mice) could significantly decrease the IFN-γ expression and Th1 frequency in the salivary glands and suppress the inflammation infiltration in salivary glands and maintain the salivary flow rates, both of which reflect SS-like symptoms. This study identifies a promising target that could effectively attenuate the abnormal state of CD4+ T cells and reverse the progression of SS.

Gancao Fuzi decoction regulates the Th17/Treg cell imbalance in rheumatoid arthritis by targeting Foxp3 via miR-34a

Ethnopharmacological relevanceDuring the Eastern Han Dynasty, Zhang Zhongjing first recorded the Gancao Fuzi decoction (GCFZD) formula in the “Synopsis of the Golden Chamber”, which is reportedly an effective and safe treatment for rheumatoid arthritis (RA). However, the mechanism underlying the observed improvement in the T helper 17 (Th17)/regulatory T (Treg) cell imbalance in RA obtained with GCFZD has not been reported. Aim of the studyThis study aimed to demonstrate whether GCFZD ameliorated RA by modulating the Th17/Treg imbalance in RA mice. Materials and methodsCollagen was used to induce a model of collagen-induced arthritis (CIA) in mice. GCFZD was administered by gavage, and the arthritis index score, imaging and histopathological changes of the ankle joints, and the levels of the immunoglobulin G (IgG) class antibodies and proinflammatory factors in serum were determined. In addition, the frequencies of Th17 and Treg cells, the levels of relevant transcription factors and functional factors and the miR-34a gene in the spleen and the levels of interleukin-17A (IL-17A) and IL-10 in serum were determined. ResultsGCFZD significantly reduced the arthritis score, improved joint swelling and bone damage, reduced the pathological score, and decreased the serum levels of IgG class antibody (IgG and IgG2a) and proinflammatory factor [tumour necrosis factor-alpha (TNF-α), IL-1β and IL-6]. Moreover, the Th17-cell proportion, the expression level of the Th17-specific transcription factor retinoic acid-related orphan receptor γt (RORγt) and functional factor IL-17A in the spleen, and the serum IL-17A level were decreased, whereas the Treg cell proportion, expression levels of the Treg-specific transcription factor forkhead box P3 (Foxp3) and functional factor IL-10 in the spleen, and the serum IL-10 level were increased. Furthermore, GCFZD inhibited miR-34a gene expression while promoting Foxp3 protein expression. ConclusionsThe findings of this study demonstrate the therapeutic effect of GCFZD on mice with CIA, and the mechanism is related to an improvement in the Th17/Treg cell imbalance by targeting Foxp3 via miR-34a.

Cd4+ T cell phenotypes, autoantibodies and transcriptional signatures overlap between myocarditis, ischemic and nonischemic heart failure, suggesting shared inflammatory pathogenic mechanisms

Abstract Background Myocarditis may progress to heart failure (HF) often as a result of viral myocardial infection where cardiomyocyte damage exposes sequestered self-antigens including cardiac myosin (CM) leading to autoimmunity. CM acts as an adjuvant via activation of Toll-like receptor (TLR)2 and TLR8, and is a potent autoantigen promoting HF in myocarditis through elevated IL-6, IL-17A and Th17 differentiation concomitant with autoantibody responses against CM. Since cardiomyocyte damage also occurs in ischemic and nonischemic cardiomyopathies, we investigated the hypothesis that ischemic and nonischemic HF would have similar inflammatory phenotypes. Purpose We defined CD4+ T helper cell phenotypes, autoantibodies, and peripheral blood transcriptome of the inflammatory phenotypes in myocarditis, compared to ischemic, and nonischemic HF. The identification of immune phenotypes in myocarditis and HF are important for consideration of inflammatory interventions and immunotherapies in myocarditis and HF. Methods Peripheral blood was collected from myocarditis patients within 6 months of disease onset, and from clinically stable, chronic HF patients. CD4+ T helper cells (Th1, Th2, and Th17) were evaluated by flow cytometry after density gradient separation, and human CM autoantibody titers were determined by ELISA. Whole blood RNA sequencing, Ingenuity Pathway Analysis (IPA), as well as multi-contrast pathway enrichment were performed. Results (Fig. 1A) Significantly elevated human CM autoantibodies were observed in myocarditis, ischemic and nonischemic HF patients, and correlated with nonrecovered patients and poor outcomes. (Fig. 1B) Myocarditis and nonischemic HF patients had significantly elevated Th17 and Th2 frequencies. Ischemic HF patients had a trend toward elevated Th 17 and Th 2 CD4+ T cell frequencies. None of the cohorts demonstrated an elevation in Th1 (not shown) (Fig 2A), IPA canonical pathway enrichment for all cohorts together showed elevated pathways concordant with our proposed pathogenic mechanisms in myocarditis and HF, including TLR and inflammasome signaling and IL17A and IL13 pathways (Fig. 2B). Myocarditis and nonischemic HF shared more inflammatory pathway enrichment in common than ischemic HF (Fig. 2C) Multi contrast pathway enrichment demonstrated that males in each cohort shared TLR and IL1b signaling that was not observed in females. Conclusion Nonischemic HF patients have overlapping immunological features with myocarditis including elevated anti-CM autoantibodies, as well as Th17, and Th2 frequencies. Ischemic HF demonstrated elevated CM autoantibodies and fewer inflammatory genes and pathways. The transcriptome supported the immune phenotypes in myocarditis and heart failure suggesting pathogenic autoimmune and inflammatory mechanisms shared in myocarditis and ischemic HF. The results have implications for immunomodulatory therapies in myocarditis and HF. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): National Institutes of Health

T-cell deficiency and hyperinflammatory monocyte responses associate with Mycobacterium avium complex lung disease.

Immunological mechanisms of susceptibility to nontuberculous mycobacterial (NTM) disease are poorly understood. To understand NTM pathogenesis, we evaluated innate and antigen-specific adaptive immune responses to Mycobacterium avium complex (MAC) in asymptomatic individuals with a previous history of MAC lung disease (MACDZ). We hypothesized that Mav-specific immune responses are associated with susceptibility to MAC lung disease. We measured MAC-, NTM-, or MAC/Mtb-specific T-cell responses by cytokine production, expression of surface markers, and analysis of global gene expression in 27 MACDZ individuals and 32 healthy controls. We also analyzed global gene expression in Mycobacterium avium-infected and uninfected peripheral blood monocytes from 17 MACDZ and 17 healthy controls. We were unable to detect increased T-cell responses against MAC-specific reagents in MACDZ compared to controls, while the responses to non-mycobacteria derived antigens were preserved. MACDZ individuals had a lower frequency of Th1 and Th1* T-cell populations. In addition, MACDZ subjects had lower transcriptional responses in PBMCs stimulated with a mycobacterial peptide pool (MTB300). By contrast, global gene expression analysis demonstrated upregulation of proinflammatory pathways in uninfected and M. avium-infected monocytes, i.e. a hyperinflammatory in vitro response, derived from MACDZ subjects compared to controls. Together, these data suggest a novel immunologic defect which underlies MAC pathogenesis and includes concurrent innate and adaptive dysregulation which persists years after completion of treatment.

Leflunomide combined with low-dose prednisone inhibits proinflammatory T cells responses in myasthenia gravis patients.

We previously found that leflunomide combined with low-dose prednisone rapidly improved the clinical symptoms of myasthenia gravis (MG), but we had not investigated the mechanism of this phenomenon. This study documents the effect of leflunomide combined with low-dose prednisone on pro-inflammatory T cells in MG patients. We compared 32 treated MG patients with 18 controls. We collected peripheral blood before treatment and 4, 8, and 12 weeks after treatment. We extracted peripheral blood mononuclear cells (PBMCs) and stimulated them with phorbol 12-myristate 13-acetate (PMA) + ionomycin and quantified IFN-γ, IL-4, IL-17, and IL-9 secretion through ELISA. We quantified T helper (Th) cells Th1 (CD3+CD4+IFN-γ+), Th2 (CD3+CD4+IL-4+), Th17 (CD3+CD4+IL-17A+) and Th9 (CD3+CD4+IL-9+) among PBMCs. The treatment significantly reduced IL-17 and IL-9 secretion in peripheral blood but did not affect IFN-γ levels. Significant decreases in IL-17 and IL-9 appeared at week 12, and the trend of change was similar to that of the MG composite score. Flow cytometry indicated that leflunomide combined with low-dose prednisone significantly reduced the frequency of Th1 and Th17 cells. These findings demonstrate the potential of this treatment as an alternative immunosuppressive therapy for MG.

A mouse model of irradiation and spleen-thymus lymphocyte infusion induced aplastic anemia

ABSTRACT Objectives The immune-induced aplastic anemia (AA) mouse model has been used for the study of AA. However, there were no uniform conditions for establishing a model and no assessment of immunological homeostasis. Our study aimed to identify the conditions of establishing a model and assess the AA model in immunology and pathology. Methods We induced an AA mouse model by the combination between sublethal irradiation and spleen-thymus lymphocyte infusion. The success of establishing the AA model was identified by blood routine tests and pathology of bone marrow. The frequency of Th17 and Treg cells was measured by flow cytometry. The frequency of CD34+ and CD41+ cells was detected by immunohistochemical technique.IL-6, IL-8, IL-17, TNF-α and IFN-γ were evaluated by ELISA. Results The 137Cs sublethal irradiation (5 Gy) and spleen-thymus lymphocyte infusion (5 106) induced the AA mouse model successfully. The AA mice had a long lifetime and manifested pancytopenia and bone marrow failure. The percentage of Th17 cells increased and the percentage of Treg cells decreased distinctly in AA mice. The area of hematopoietic tissues and count of CD34+ cells and CD41+ cells were significantly reduced in AA mice.The level of cytokines, IL-6, IL-8, IL-17, TNF-α and IFN-γ, was increased significantly in peripheral blood and bone marrow. Conclusion Our data suggest that the improved AA mouse model conforms to the diagnosis standard of AA and simulates the immune internal environment of human AA. The AA mouse model has a longer lifetime and unbalances of Th17/Treg cells caused the destruction of CD34+ cells and CD41+ cells, which was immune-mediated pathogenesis to adapt to long-term research.

Gamma-Delta T-Cell Phenotype and Function in DAA-Treated HIV-HCV Co-Infected and HCV-Mono-Infected Subjects

HIV-HCV co-infected subjects are at risk of liver fibrosis which may be linked to immune imbalances. Direct-acting antivirals (DAAs) represent the mainstay of HCV treatment in co-infected individuals, yet their effects on immune cell populations playing a role in fibrogenesis is unknown. We assessed γδ T-cell phenotype and function, Treg and Th17 frequencies, as well as γ-globulins and B-cell activation in 47 HIV-HCV co-infected and 35 HCV mono-infected individuals prior to and following DAA treatment (SVR12). Γδ T-cell activation decreased in both groups yet persisted at higher levels in the HIV-HCV co-infected subjects. No differences were registered in terms of γδT-cell function. Of note, the Vδ2/Th17 ratio, inversely linked to liver damage, increased significantly in the two groups upon treatment, yet a negative correlation between the Vδ2/Th17 ratio and liver function enzymes was found in the co-infected subjects alone. B-cell activation and γ-globulin levels decreased in both settings, yet B-cell activation remained higher in the HIV-HCV co-infected individuals. In HIV-HCV co-infected and HCV mono-infected participants, the effect of DAA was limited to γδ T- and B-cell activation as well as γ-globulin concentrations and the Vδ2/Th17 ratio, with no changes in γδ T-cell function and Treg frequencies. Importantly, γδ T- and B-cell activation remained at higher levels in the co-infected individuals than in those with HCV mono-infection alone. The persistence of such alterations within these cell subsets may be associated with the risk of hepatic and extrahepatic complications.

A comprehensive evaluation of the immune system response and type-I Interferon signaling pathway in hospitalized COVID-19 patients

BackgroundThe COVID-19 pandemic has become the world’s main life-threatening challenge in the third decade of the twenty-first century. Numerous studies have been conducted on SARS-CoV2 virus structure and pathogenesis to find reliable treatments and vaccines. The present study aimed to evaluate the immune-phenotype and IFN-I signaling pathways of COVID-19 patients with mild and severe conditions.Material and methodsA total of 100 COVID-19 patients (50 with mild and 50 with severe conditions) were enrolled in this study. The frequency of CD4 + T, CD8 + T, Th17, Treg, and B lymphocytes beside NK cells was evaluated using flow cytometry. IFN-I downstream signaling molecules, including JAK-1, TYK-2, STAT-1, and STAT-2, and Interferon regulatory factors (IRF) 3 and 7 expressions at RNA and protein status were investigated using real-time PCR and western blotting techniques, respectively. Immune levels of cytokines (e.g., IL-1β, IL-6, IL-17, TNF-α, IL-2R, IL-10, IFN-α, and IFN-β) and the existence of anti-IFN-α autoantibodies were evaluated via enzyme-linked immunosorbent assay (ELISA).ResultsImmune-phenotyping results showed a significant decrease in the absolute count of NK cells, CD4 + T, CD8 + T, and B lymphocytes in COVID-19 patients. The frequency of Th17 and Treg cells showed a remarkable increase and decrease, respectively. All signaling molecules of the IFN-I downstream pathway and IRFs (i.e., JAK-1, TYK-2, STAT-1, STAT-2, IRF-3, and IRF-7) showed very reduced expression levels in COVID-19 patients with the severe condition compared to healthy individuals at both RNA and protein levels. Of 50 patients with severe conditions, 14 had anti-IFN-α autoantibodies in sera. Meanwhile, this result was 2 and 0 for patients with mild symptoms and healthy controls, respectively.ConclusionOur results indicate a positive association of the existence of anti-IFN-α autoantibodies and immune cells dysregulation with the severity of illness in COVID-19 patients. However, comprehensive studies are necessary to find out more about this context.5mo_nwLJpK7s-uZity9h4-Video abstract

IL-17A aggravates asthma-induced intestinal immune injury by promoting neutrophil trafficking.

With the concept of the gut-lung axis reinforced in recent years, emerging evidence has shown that intestinal homeostasis is vital for lung health. Nevertheless, the impacts of lung homeostasis on intestinal tracts and their mechanism are rarely studied. Our results showed that papain-induced asthmatic mice exhibited apparent colonic injuries compared with controls, including increased intestinal permeability, neutrophil and Th17 infiltration in the colonic lamina propria. Moreover, the intranasal administration of papain aggravated such colonic injuries in mice with dextran sulfate sodium-induced colitis, as evidenced by increased occult blood scores, shortened colon length, and accumulated neutrophils. The level of IL-17A was also higher in the serum of asthmatic mice than wild-type mice. Interestingly, the pathologic scores, the proportion of Th17 cells, and neutrophil infiltration in the colon were markedly reduced after IL-17A blocking. Similarly, longer length, lower pathologic scores, and fewer neutrophils were also observed in the colon of IL-17-deficient asthmatic mice. More importantly, we demonstrated that severe gastrointestinal symptoms could accompany clinical asthmatics. The frequencies of Th17 cells and the mRNA expression of IL-17A in the peripheral blood of these patients were significantly enhanced. Besides, the gastrointestinal symptom rating scale scores positively correlated with the frequencies of Th17 in asthmatics. These findings enlighten that IL-17A aggravates asthma-induced intestinal immune injury by promoting neutrophil trafficking, which facilitates the exploration of new potential biomarkers to treat asthma.

T helper profile in pregnant women recovered from COVID-19

T helper (Th) cell subsets play distinct and important roles during pregnancy. This work was focused on investigating the Th and cytokine profile in pregnant women recovered from COVID-19. To this aim, the frequency of Th1, Th2, Th17 subsets and the level of associated cytokines were analysed in pregnant women recovered from COVID-19 and in matched non-pregnant women. Principal component analysis highlighted a significant impact of pregnancy on Th profile with an increase of ex-Th17 subset and a parallel decrease of Th1 population. These modulations may participate in both preserving the pregnancy and reducing the risk of severe infection.

Cimetidine and Ibuprofen Modulate T Cell Responses in a Mouse Model of Breast Cancer.

Cimetidine and ibuprofen exhibit immunomodulatory effects as an antagonist of histamine H2 receptor, and a cyclooxygenase inhibitor, respectively. Here, the effects of cimetidine and ibuprofen on some effector T cell-related parameters were investigated using a breast cancer (BC) model. BC was established in Balb/c mice using the 4T1 cell line. On day 10 after tumor induction, the BC-bearing mice were classified into four groups and treated with PBS, cimetidine (20 mg/kg), ibuprofen (20 mg/kg) or a combination of “cimetidine + ibuprofen” via intraperitoneal injection (daily from days 11 to 30). The mice were sacrificed on day 31 and the frequency of splenic Th1 and Treg cells, plasma IFN-γ and TGF-β levels, and intra-tumoral T-bet, GATA3, FOXP3 and RORγt expressions were detected using flowcytometry, ELISA and real-time-PCR, respectively. In untreated cancerous mice, the percentage of splenic Th1 cells and plasma IFN-γ levels were lower (P<0.003 and P<0.01, respectively), whereas the percentage of splenic Treg cells and plasma TGF-β levels were higher than in healthy mice (P<0.04 and P<0.005, respectively). Treatment of BC-bearing mice with cimetidine, ibuprofen or both drugs promoted the frequency of Th1 cells (P<0.05, P<0.007 and P<0.005, respectively) as well as IFN-γ levels (P<0.004, P<0.0001 and P<0.03, respectively), while reduced the frequencies of Treg cells (P<0.02, P<0.03 and P<0.01, respectively), TGF-β levels (P<0.006, P<0.02 and P<0.002, respectively), intra-tumoral expression of FOXP3 (P<0.006, P<0.005 and P<0.005, respectively), and intra-tumoral expression of RORγt (P<0.04, P<0.03 and P<0.05, respectively) compared with untreated BC mice. The “cimetidine + ibuprofen”-treated mice displayed greater T-bet expression than the un-treated mice (P<0.006). Cimetidine and/or ibuprofen-treated BC-bearing mice exhibited reduced intra-tumoral expression of GATA3 compared with the untreated BC mice, but the differences were not significant. Cimetidine and ibuprofen correct some effector T cell-related parameters in cancerous mice. Immunotherapeutic potentials cimetidine and ibuprofen in cancers need investigations.

7-DAYS HIGH SALT DIET AFFECTS THE PRODUCTION OF PRO- AND ANTI-INFLAMMATORY CYTOKINES IN YOUNG HEALTHY INDIVIDUALS

Objective: This study aimed to investigate the earliest immunomodulatory effects of NaCl during pre-hypertensive phase of acute salt loading in humans. Thus we have investigated serum levels of systemic pro- and anti-inflammatory cytokines in relation to peripheral Th17 and regulatory T cell abundance following 7-day high-salt (HS) loading in young salt-resistant healthy individuals. Design and method: 30 young healthy individuals adhered to a 7-day low-salt (LS) diet (3.5 g salt/day) followed by a 7-day high-salt (HS) diet (~14.7 g salt/day) protocol. Serum pro- and anti-inflammatory cytokines (INF-gamma, TNF-alpha, IL-6, IL-9, IL-17A, IL-23, IL-10, IL-21, IL-22, TGF-beta1), complement component C3a, as well as frequencies of peripheral blood Th17 and regulatory T cells (Treg), Th17/Treg ratio and the level of enzymes involved in their activation and differentiation (SGK1 and p38/MAP kinase) were measured before and after 7-day HS diet protocol. Results: Short-term HS diet significantly increased serum level of pro-inflammatory cytokines INF-gamma, TNF-alpha, IL-9 and IL-17A level, but also serum concentration of anti-inflammatory cytokines IL-10 and TGF-beta1. In addition, serum INF-gamma and IL-9 levels positively correlated to 24-h urine natriuresis. Relative amount of total and phosphorylated p38/MA kinase remained unchanged, while relative amount of total SGK1 significantly increased following 7-day HS diet compared to LS values. Conclusions: Result of the present study provided evidence of acute salt-loading induced changes in the immune status of young healthy individuals. The increase in relative amount of total SGK1 in peripheral mononuclear cells suggests lymphocyte (re)activation in response to hyperosmotic stress resulting in enhanced production of proinflammatory (IL-17, INF-gamma), but also anti-inflammatory cytokines (IL-10 and TGF-beta1). Still, which effect is prevailing remains elusive and warrants further studies.

The pro-inflammatory effect of triglyceride on human CD4+ T cells and experimental autoimmune uveitis

Aberrant lipid metabolism plays a role in inflammation and progression of autoimmune diseases but the definite mechanism remains unclear. In this study we investigate lipidomic profiles in Behçet's disease (BD) and the role of triglyceride (TAG) in the pathogenesis of autoimmune uveitis. Lipidomics revealed a distinct lipid metabolite profile including increased TAG metabolites in plasma of active BD patients. TAG could stimulate the proliferation, IL-17 and IFN-γ expression by CD4+ T cells and Th1, Th17 cell differentiation in vitro, but did not influence neutrophils. A922500 inhibited the TAG generation, ameliorated the EAU severity, decreased Th17 frequency and IL-17 expression by CD4+ T cells in vivo. The proteomocis analysis showed an up-regulation of apoptosis-related protein, Pik3r2, in CD4+ T cells from A922500-treated mice. In conclusion, TAG can stimulate human CD4+ T cells and the inhibition of its generation could significantly ameliorate EAU activity in association with down-regulated Th17 cell response.