Deficient regulatory T cell activity and low frequency of IL-17-producing T cells correlate with the extent of cardiomyopathy in human Chagas' disease

Background

Myocardium damage during Chagas'' disease results from the immunological imbalance between pro- and production of anti-inflammatory cytokines and has been explained based on the Th1–Th2 dichotomy and regulatory T cell activity. Recently, we demonstrated that IL-17 produced during experimental T. cruzi infection regulates Th1 cells differentiation and parasite induced myocarditis. Here, we investigated the role of IL-17 and regulatory T cell during human Chagas'' disease.

Methodology/Principal Findings

First, we observed CD4⁺IL-17⁺ T cells in culture of peripheral blood mononuclear cells (PBMC) from Chagas'' disease patients and we evaluated Th1, Th2, Th17 cytokine profile production in the PBMC cells from Chagas'' disease patients (cardiomyopathy-free, and with mild, moderate or severe cardiomyopathy) cultured with T. cruzi antigen. Cultures of PBMC from patients with moderate and severe cardiomyopathy produced high levels of TNF-α, IFN-γ and low levels of IL-10, when compared to mild cardiomyopathy or cardiomyopathy-free patients. Flow cytometry analysis showed higher CD4⁺IL-17⁺ cells in PBMC cultured from patients without or with mild cardiomyopathy, in comparison to patients with moderate or severe cardiomyopathy. We then analyzed the presence and function of regulatory T cells in all patients. All groups of Chagas'' disease patients presented the same frequency of CD4⁺CD25⁺ regulatory T cells. However, CD4⁺CD25⁺ T cells from patients with mild cardiomyopathy or cardiomyopathy-free showed higher suppressive activity than those with moderate and severe cardiomyopathy. IFN-γ levels during chronic Chagas'' disease are inversely correlated to the LVEF (P = 0.007, r = −0.614), while regulatory T cell activity is directly correlated with LVEF (P = 0.022, r = 0.500).

Conclusion/Significance

These results indicate that reduced production of the cytokines IL-10 and IL-17 in association with high levels of IFN-γ and TNF-α is correlated with the severity of the Chagas'' disease cardiomyopathy, and the immunological imbalance observed may be causally related with deficient suppressor activity of regulatory T cells that controls myocardial inflammation.     

The role of IL-27 in susceptibility to post-influenza Staphylococcus aureus pneumonia

Background

Influenza is a common respiratory virus and Staphylococcus aureus frequently causes secondary pneumonia during influenza infection, leading to increased morbidity and mortality. Influenza has been found to attenuate subsequent Type 17 immunity, enhancing susceptibility to secondary bacterial infections. IL-27 is known to inhibit Type 17 immunity, suggesting a potential critical role for IL-27 in viral and bacterial co-infection.

Methods

A murine model of influenza and Staphylococcus aureus infection was used to mimic human viral, bacterial co-infection. C57BL/6 wild-type, IL-27 receptor α knock-out, and IL-10 knock-out mice were infected with Influenza H1N1 (A/PR/8/34) or vehicle for 6 days followed by challenge with Staphylococcus aureus or vehicle for 24 hours. Lung inflammation, bacterial burden, gene expression, and cytokine production were determined.

Results

IL-27 receptor α knock-out mice challenged with influenza A had increased morbidity compared to controls, but no change in viral burden. IL-27 receptor α knock-out mice infected with influenza displayed significantly decreased IL-10 production compared to wild-type. IL-27 receptor α knock-out mice co-infected with influenza and S. aureus had improved bacterial clearance compared to wild-type controls. Importantly, there were significantly increased Type 17 responses and decreased IL-10 production in IL-27 receptor α knock-out mice. Dual infected IL-10−/− mice had significantly less bacterial burden compared to dual infected WT mice.

Conclusions

These data reveal that IL-27 regulates enhanced susceptibility to S. aureus pneumonia following influenza infection, potentially through the induction of IL-10 and suppression of IL-17.     

Quantiferon-TB Gold in tube assay for the screening of tuberculosis before and during treatment with tumor necrosis factor alpha antagonists

Introduction

The usefulness of interferon-gamma (IFN-γ) release assays for tuberculosis screening before tumor necrosis factor-alpha (TNF-α) antagonists and for monitoring during treatment is a contraversial issue. The aims of this study were to determine whether TNF-α antagonists affect the results of the Quantiferon-TB Gold in-tube assay (QTF); to assess how QTF performs in comparison with the tuberculin skin test (TST) in rheumatoid arthritis (RA) patients who are about to start treatment with TNF-α antagonists, RA patients who are not candidates for treatment with TNF-α antagonists, rheumatology patients with confirmed current or past tuberculosis infection, and healthy controls, and to determine the specificity of the QTF test to differentiate leprosy patients, another group of patients infected with mycobacteria.

Methods

The 38 RA patients who were prescribed TNF-α antagonists, 40 RA patients who were not considered for TNF-α antagonist use, 30 rheumatology patients with a history or new diagnosis of tuberculosis, 23 leprosy patients, and 41 healthy controls were studied. QTF and TST were done on the same day, and both were repeated after a mean of 3.6 ± 0.2 months in patients who used TNF-α antagonists.

Results

Treatment with TNF-α antagonists did not cause a significant change in the QTF or TST positivity rate (34% versus 42%; P = 0.64; and 24% versus 37%; P = 0.22). Patients with leprosy had a trend for a higher mean IFN-γ level (7.3 ± 8.0) and QTF positivity (61%) than did the other groups; however, the difference was not significant (P = 0.09 and P = 0.43).

Conclusions

Treatment with TNF-α antagonists does not seem to affect the QTF test to an appreciable degree. The higher IFN-γ levels in leprosy patients deserves further attention.     

Use of praziquantel as an adjuvant enhances protection and Tc-17 responses to killed H5N1 virus vaccine in mice

Background

H5N1 is a highly pathogenic influenza A virus, which can cause severe illness or even death in humans. Although the widely used killed vaccines are able to provide some protection against infection via neutralizing antibodies, cytotoxic T-lymphocyte responses that are thought to eradicate viral infections are lacking.

Methodology/Principal Findings

Aiming to promote cytotoxic responses against H5N1 infection, we extended our previous finding that praziquantel (PZQ) can act as an adjuvant to induce IL-17-producing CD8⁺ T cells (Tc17). We found that a single immunization of 57BL/6 mice with killed viral vaccine plus PZQ induced antigen-specific Tc17 cells, some of which also secreted IFN-γ. The induced Tc17 had cytolytic activities. Induction of these cells was impaired in CD8 knockout (KO) or IFN-γ KO mice, and was even lower in IL-17 KO mice. Importantly, the inoculation of killed vaccine with PZQ significantly reduced virus loads in the lung tissues and prolonged survival. Protection against H5N1 virus infection was obtained by adoptively transferring PZQ-primed wild type CD8⁺ T cells and this was more effective than transfer of activated IFN-γ KO or IL-17 KO CD8⁺ T cells.

Conclusions/Significance

Our results demonstrated that adding PZQ to killed H5N1 vaccine could promote broad Tc17-mediated cytotoxic T lymphocyte activity, resulting in improved control of highly pathogenic avian influenza virus infection.     

Qualitative immune modulation by interleukin-2 (IL-2) adjuvant therapy in immunological non responder HIV-infected patients

Background

Treatment of HIV-infected patients with interleukin-2 (IL-2) produces significant increases in CD4 T cell counts; however an associated qualitative improvement in cells function has yet to be conclusively demonstrated. By measuring mycobacterial killing activity, we evaluated IL-2-mediated functional immune enhancement ex vivo in immunological non-responders (INRs).

Methods and Findings

PBMC from 12 immunological non-responders (INRs) (CD4+<200/µl, HIV-RNA<50 cp/ml) on combination antiretroviral treatment (cART) were collected at baseline, and after 3 IL-2 cycles. Eight INRs receiving only cART were studied as controls. After 21 days of PBMC incubation with a virulent M. avium suspension, counts of residual colony forming units (CFUs) and concentrations of TNF-α, IL-10 and IFN-γ were determined. In IL-2 treated patients, a significant reduction in mean residual CFUs of PBMC cultures was observed (p<0.01). Moreover, following IL-2 treatment, significant increases in PBMC''s IFNγ production (p = 0.02) and substantial reductions in IL-10 levels were observed.

Conclusions

IL-2 therapy restores the ability of the lympho-monocyte system in eliciting an effective response against mycobacterial infections. Our data indicate the possibility of a clinical role held by IL-2 in enhancing the immune function of subjects unable to achieve immune competence through cART alone.     

TNF,IL6, and IL1B Polymorphisms Are Associated with Severe Influenza A (H1N1) Virus Infection in the Mexican Population

Background

Hypercytokinemia is the main immunopathological mechanism contributing to a more severe clinical course in influenza A (H1N1) virus infections. Most patients infected with the influenza A (H1N1) pdm09 virus had increased systemic levels of pro-inflammatory cytokines; including interleukin IL-6, IL-8, and tumor necrosis factor-alpha (TNF-α). We propose that single-nucleotide polymorphisms (SNPs) in the promoter regions of pro-inflammatory genes are associated with the severity of influenza A (H1N1) pdm09 virus infection.

Methods

145 patients with influenza A (H1N1) (pA/H1N1), 133 patients with influenza-like illness (ILI), and 360 asymptomatic healthy contacts (AHCs) were included. Eleven SNPs were genotyped in six genes (TNF, LT, IL1B, IL6, CCL1, and IL8) using real-time PCR; the ancestral genotype was used for comparison. Genotypes were correlated with 27 clinical severity variables. Ten cytokines (GM-CSF, TNF-α, IL-2, IL-1β, IL-6, IL-8, IFN-γ, IL-10, IL-5, and IL-4) were measured on a Luminex 100.

Results

The IL6 rs1818879 (GA) heterozygous genotype was associated with severe influenza A (H1N1) virus infection (odds ratio [OR] = 5.94, 95% confidence interval [CI] 3.05–11.56), and two IL1B SNPs, rs16944 AG and rs3136558 TC, were associated with a decreased risk of infection (OR = 0.52 and OR = 0.51, respectively). Genetic susceptibility was determined (pA/H1N1 vs. AHC): the LTA rs909253 TC heterozygous genotype conferred greater risk (OR = 1.9), and a similar association was observed with the IL1B rs3136558 CC genotype (OR = 1.89). Additionally, severely ill patients were compared with moderately ill patients. The TNF-238 GA genotype was associated with an increased risk of disease severity (OR = 16.06, p = 0.007). Compared with ILIs, patients with severe pA/H1N1 infections exhibited increased serum IL-5 (p <0.001) and IL-6 (p  =  0.007) levels.

Conclusions

The TNF gene was associated with disease severity, whereas IL1B and IL6 SNPs were associated with influenza A (H1N1) virus infection.     

Akt regulates IL-10 mediated suppression of TNFα-induced cardiomyocyte apoptosis by upregulating Stat3 phosphorylation

Background

We have already reported that TNF-α increases cardiomyocyte apoptosis and IL-10 treatment prevented these effects of TNF-α. Present study investigates the role of Akt and Jak/Stat pathway in the IL-10 modulation of TNF-α induced cardiomyocyte apoptosis.

Methodology/Principal findings

Cardiomyocytes isolated from adult Sprague Dawley rats were exposed to TNF-α (10 ng/ml), IL-10 (10 ng/ml) and TNF-α+IL-10 (ratio 1) for 4 h. Exposure to TNF-α resulted in an increase in cardiomyocyte apoptosis as measured by flow cytometry and TUNEL assay. IL-10 by itself had no effect, but it prevented TNF-α induced apoptosis. IL-10 treatment increased Akt levels within cardiomyocytes and this change was associated with an increase in Jak1 and Stat3 phosphorylation. Pre-exposure of cells to Akt inhibitor prevented IL-10 induced Stat3 phosphorylation. Furthermore, in the presence of Akt or Stat3 inhibitor, IL-10 treatment was unable to block TNF-α induced cardiomyocyte apoptosis.

Conclusion

It is suggested that IL-10 modulation of TNF-α induced cardiomyocyte apoptosis is mediated by Akt via Stat3 activation.     

Comprehensive Analysis of Inflammatory Immune Mediators of the Intraocular Fluid Aspirated from the Foldable Capsular Vitreous Body Filled-Eyes

Purpose

To analyze the level of human inflammatory immune mediators in the intraocular fluid aspirated from foldable capsular vitreous body (FCVB) filled-eyes during FCVB removal surgery, 3 months after implantation.

Methods

8 samples of intra-FCVB fluids (n = 8) were collected from 8 FCVB filled patients in our previous FCVB exploratory clinical trial. The intra-FCVB fluids were aspirated from the FCVB filled-eyes during the FCVB removal surgeries at the third month. For the control groups, the vitreous fluids were collected from patients with idiopathic macular hole (n = 9) or rhegmatogenous retinal detachment (n = 6) during pars plana vitrectomy. A multiplex immunoassay was used to determine levels of 9 cytokines (IL-1β, IL-2, IL-6, IL-8, IL-10, IL-17, TNF-α, IFN-γ and VEGF) in these samples. The VEGF level of some intra-FCVB fluids (n = 6) were re-tested using enzyme-linked immunosorbent assay (ELISA).

Results

In the intra-FCVB fluids, 9 cytokines concentrations of most samples (n = 5) measured by Multiplex immunoassay showed low values, except for Patient 02, 06, and 09. The VEGF concentrations of some intra-FCVB fluids (n = 6) tested by ELISA were in accordance with Multiplex immunoassay results. For all eight patients (n = 8), the concentrations of IL-1β, IL-2, IL-6, TNF-α, IFN-γ and VEGF were slightly higher as compared to the idiopathic macular hole control group. While, the concentrations of IL-8, IL-10, and IL-17 were not statistically significant different compared with the idiopathic macular hole control samples. Most cytokines concentrations (IL-2, IL-6, IL-8, IL-10, IL-17, TNF-α, IFN-γ, VEGF) were not statistically significant different compared to the rhegmatogenous retinal detachment control group except IL-1β.

Conclusions

The FCVB had sufficient porosity to allow cytokines to pass through. This study first discovered that the FCVB possesses favorable permeability of proteins in the human eye.     

Co-administration of a plasmid DNA encoding IL-15 improves long-term protection of a genetic vaccine against Trypanosoma cruzi

Background

Immunization of mice with the Trypanosoma cruzi trans-sialidase (TS) gene using plasmid DNA, adenoviral vector, and CpG-adjuvanted protein delivery has proven highly immunogenic and provides protection against acute lethal challenge. However, long-term protection induced by TS DNA vaccines has not been reported. The goal of the present work was to test whether the co-administration of a plasmid encoding IL-15 (pIL-15) could improve the duration of protection achieved through genetic vaccination with plasmid encoding TS (pTS) alone.

Methodology

We immunized BALB/c mice with pTS in the presence or absence of pIL-15 and studied immune responses [with TS-specific IFN-γ ELISPOT, serum IgG ELISAs, intracellular cytokine staining (IFN-γ, TNF-α, and IL-2), tetramer staining, and CFSE dilution assays] and protection against lethal systemic challenge at 1 to 6 months post vaccination. Mice receiving pTS alone developed robust TS-specific IFN-γ responses and survived a lethal challenge given within the first 3 months following immunization. The addition of pIL-15 to pTS vaccination did not significantly alter T cell responses or protection during this early post-vaccination period. However, mice vaccinated with both pTS and pIL-15 challenged 6 months post-vaccination were significantly more protected against lethal T. cruzi challenges than mice vaccinated with pTS alone (P<0.05). Improved protection correlated with significantly higher numbers of TS-specific IFN-γ producing total and CD8⁺ T cells detected>6 months post immunization. Also, these TS-specific T cells were better able to expand after in vitro re-stimulation.

Conclusion

Addition of pIL-15 during genetic vaccination greatly improved long-term T cell survival, memory T cell expansion, and long-term protection against the important human parasite, T. cruzi.

Author Summary

Over 11 million individuals are infected with Trypanosoma cruzi, the causative agent of Chagas disease, which kills >50,000 people annually. Although recent vector control efforts and increased use and effectiveness of chemotherapeutic drugs including benznidazole have reduced infection rates and mortality, a safe, effective vaccine is needed. Vaccination with the T. cruzi trans-sialidase (TS) has been used effectively in mice to reduce mortality and chronic disease, however, the establishment of vaccine-induced long-term protective immunity remains elusive. Co-immunization strategies utilizing immune regulators such as interleukin-12 (IL-12) and interleukin-15 (IL-15) can be used to enhance antigen-specific T cell responses and prolong protective immunity. In the present report, we show that genetic vaccination of BALB/c mice with plasmid DNA encoding both TS and IL-15 compared with plasmid DNA encoding TS alone significantly enhanced CD4⁺ and CD8⁺ T cell responses including increased TNF-α, IFN-γ, and IL-2 production, and long-term protection against lethal systemic parasite challenge.     

The Association between Serum Biomarkers and Disease Outcome in Influenza A(H1N1)pdm09 Virus Infection: Results of Two International Observational Cohort Studies

Background

Prospective studies establishing the temporal relationship between the degree of inflammation and human influenza disease progression are scarce. To assess predictors of disease progression among patients with influenza A(H1N1)pdm09 infection, 25 inflammatory biomarkers measured at enrollment were analyzed in two international observational cohort studies.

Methods

Among patients with RT-PCR-confirmed influenza A(H1N1)pdm09 virus infection, odds ratios (ORs) estimated by logistic regression were used to summarize the associations of biomarkers measured at enrollment with worsened disease outcome or death after 14 days of follow-up for those seeking outpatient care (FLU 002) or after 60 days for those hospitalized with influenza complications (FLU 003). Biomarkers that were significantly associated with progression in both studies (p<0.05) or only in one (p<0.002 after Bonferroni correction) were identified.

Results

In FLU 002 28/528 (5.3%) outpatients had influenza A(H1N1)pdm09 virus infection that progressed to a study endpoint of complications, hospitalization or death, whereas in FLU 003 28/170 (16.5%) inpatients enrolled from the general ward and 21/39 (53.8%) inpatients enrolled directly from the ICU experienced disease progression. Higher levels of 12 of the 25 markers were significantly associated with subsequent disease progression. Of these, 7 markers (IL-6, CD163, IL-10, LBP, IL-2, MCP-1, and IP-10), all with ORs for the 3^rd versus 1^st tertile of 2.5 or greater, were significant (p<0.05) in both outpatients and inpatients. In contrast, five markers (sICAM-1, IL-8, TNF-α, D-dimer, and sVCAM-1), all with ORs for the 3^rd versus 1^st tertile greater than 3.2, were significantly (p≤.002) associated with disease progression among hospitalized patients only.

Conclusions

In patients presenting with varying severities of influenza A(H1N1)pdm09 virus infection, a baseline elevation in several biomarkers associated with inflammation, coagulation, or immune function strongly predicted a higher risk of disease progression. It is conceivable that interventions designed to abrogate these baseline elevations might affect disease outcome.     

Effect of pregnancy on serum cytokines in SLE patients

Introduction

The aim of this study was to evaluate an extensive panel of cytokines involved in immune regulation during pregnancy in patients with systemic lupus erythematosus (SLE) and in healthy women.

Methods

A total of 47 consecutive successful pregnancies in 46 SLE patients and 56 pregnancies in 56 matched healthy subjects, as controls, were prospectively studied. Serum interleukin (IL)-1-α, IL-1-β, IL-2, IL-6, IL-8, IL-10, IL-12p70, interferon (INF)-γ and tumor necrosis factor (TNF)-α were detected in sera obtained at the first and third trimester of pregnancy by a highly sensitive, multiplexed sandwich ELISA.

Results

Medians (pg/ml) of serum levels of most helper T (Th)1-type cytokines were significantly lower in the third trimester compared with those observed in the first trimester of pregnancy in healthy women: INF-γ 2.0 vs 3.4, TNF-α 10.2 vs 11.5, IL-1-α 0.9 vs 1.1, IL-1-β 0.6 vs 1.0, IL-2 3.0 vs 3.5, and IL-12p70 4.9 vs 5.6 (P-values < 0.02 for all). By contrast, only the IL-1-α serum levels were lower in the third trimester compared with the first trimester in SLE patients (P = 0.006). IFN-γ/IL-6 and IFN-γ/IL-10 ratios were higher in controls than in SLE (P = 0.002, and P = 0.001, respectively); moreover, they were significantly reduced in the third compared to the first trimester of pregnancy in healthy women, but not in SLE.

Conclusions

In SLE patients, Th1/Th2 cytokine serum level ratio does not decrease during pregnancy progression as much as in healthy pregnant women. This could account for the observation of a low frequency of disease flares in the third trimester of gestation.     

Frequency of Th17 CD4+ T Cells in Early Rheumatoid Arthritis: A Marker of Anti-CCP Seropositivity

Objective

To examine the frequency and phenotype of Th17 cells in the peripheral blood of early RA (eRA) patients.

Methods

CD4+ T cells were isolated from the peripheral blood of 33 eRA patients, 20 established RA patients and 53 healthy controls (HC), and from the synovial fluid of 20 established RA patients (RASF), by ficoll-hypaque gradient and magnetical negative selection. After polyclonal stimulation, the frequency of Th17 and Th1 cells was determined by flow cytometry and concentrations of IL-17, IFN-γ, TNF-α and IL-10 were measured by ELISA in cell-free supernatants.

Results

When all of our eRA patients were analyzed together, a significantly lower percentage of circulating Th17 cells and a lower CD4-derived IL-17 secretion were observed in comparison with HC. However, after stratifying by anti-CCP antibody status, circulating Th17 cells were decreased in anti-CCP(+) but not in anti-CCP(-)-eRA. All Th17 cells were CD45RO+CD45RA- and CCR6+. Dual Th17/Th1 cells were also exclusively decreased in anti-CCP(+)-eRA. Circulating Th17 and Th17/Th1 cells were negatively correlated with anti-CCP titres. When anti-CCP(+)-eRA patients were retested one year after initiating treatment with oral methotrexate, their circulating Th17 frequency was no longer different from HC. Of note, the percentage of circulating Th1 cells and the secretion of CD4-derived IFN-γ, TNF-α and IL-10 were not different between eRA patients and HC. In established RA patients, circulating Th17 and T17/Th1 cell frequencies were comparable to HC. In RASF, both Th17 and Th1 cells were increased when compared with blood of eRA patients, established RA patients and HC.

Conclusion

Decreased circulating Th17 levels in eRA seem to be a marker of anti-CCP seropositivity, and return to levels observed in healthy controls after treatment with methotrexate.     

Cytokine response patterns in severe pandemic 2009 H1N1 and seasonal influenza among hospitalized adults

Background

Studying cytokine/chemokine responses in severe influenza infections caused by different virus subtypes may improve understanding on pathogenesis.

Methods

Adults hospitalized for laboratory-confirmed seasonal and pandemic 2009 A/H1N1 (pH1N1) influenza were studied. Plasma concentrations of 13 cytokines/chemokines were measured at presentation and then serially, using cytometric-bead-array with flow-cytometry and ELISA. PBMCs from influenza patients were studied for cytokine/chemokine expression using ex-vivo culture (Whole Blood Assay,±PHA/LPS stimulation). Clinical variables were prospectively recorded and analyzed.

Results

63 pH1N1 and 53 seasonal influenza patients were studied. pH1N1 patients were younger (mean±S.D. 42.8±19.2 vs 70.5±16.7 years), and fewer had comorbidities. Respiratory/cardiovascular complications were common in both groups (71.4% vs 81.1%), although severe pneumonia with hypoxemia (54.0% vs 28.3%) and ICU admissions (25.4% vs 1.9%) were more frequent with pH1N1. Hyperactivation of the proinflammatory cytokines IL-6, CXCL8/IL-8, CCL2/MCP-1 and sTNFR-1 was found in pH1N1 pneumonia (2–15 times normal) and in complicated seasonal influenza, but not in milder pH1N1 infections. The adaptive-immunity (Th1/Th17)-related CXCL10/IP-10, CXCL9/MIG and IL-17A however, were markedly suppressed in severe pH1N1 pneumonia (2–27 times lower than seasonal influenza; P−values<0.01). This pattern was further confirmed with serial measurements. Hypercytokinemia tended to be sustained in pH1N1 pneumonia, associated with a slower viral clearance [PCR-negativity: day 3–4, 55% vs 85%; day 6–7, 67% vs 100%]. Elevated proinflammatory cytokines, particularly IL-6, predicted ICU admission (adjusted OR 12.6, 95%CI 2.6–61.5, per log₁₀unit increase; P = 0.002), and correlated with fever, tachypnoea, deoxygenation, and length-of-stay (Spearman''s rho, P-values<0.01) in influenza infections. PBMCs in seasonal influenza patients were activated and expressed cytokines ex vivo (e.g. IL-6, CXCL8/IL-8, CCL2/MCP-1, CXCL10/IP-10, CXCL9/MIG); their ‘responsiveness’ to stimuli was shown to change dynamically during the illness course.

Conclusions

A hyperactivated proinflammatory, but suppressed adaptive-immunity (Th1/Th17)-related cytokine response pattern was found in severe pH1N1 pneumonia, different from seasonal influenza. Cytokine/immune-dysregulation may be important in its pathogenesis.     

Cytokine and chemokine levels in patients with severe Fever with thrombocytopenia syndrome virus

Background

Severe fever with thrombocytopenia syndrome virus (SFTSV), which can cause hemorrhagic fever–like illness, is a newly discovered bunyavirus in China. The pathogenesis of SFTSV infection is poorly understood. However, it has been suggested that immune mechanisms, including cytokines and chemokines, play an important role in disease pathogenesis. In the present study, we investigated host cytokine and chemokine profiles in serum samples of patients with SFTSV infection from Northeast China and explored a possible correlation between cytokine levels and disease severity.

Methods and Principal Findings

Acute phase serum samples from 40 patients, diagnosed with SFTSV infection were included. Patients were divided into two groups – severe or non-severe – based on disease severity. Levels of tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β, interleukin-6, interferon (IFN)-γ, IFN- γ-induced protein (IP)-10 and RANTES were measured in the serum samples with commercial ELISAs. Statistical analysis showed that increases in TNF-α, IP-10 and IFN-γ were associated with disease severity.

Conclusions

We suggest that a cytokine-mediated inflammatory response, characterized by cytokine and chemokine production imbalance, might be in part responsible for the disease progression of patients with SFTSV infection.     

Localized delivery of interferon-β by Lactobacillus exacerbates experimental colitis

Background

There have been conflicting reports of the role of Type I interferons (IFN) in inflammatory bowel disease (IBD). Clinical trials have shown potent efficacy of systemic interferon-beta (IFN-β) in inducing remission of ulcerative colitis. Likewise, IFNAR1^−/− mice display an increased sensitivity to dextran sulfate sodium (DSS)-induced colitis, suggesting Type I IFN play a protective role during inflammation of the gut. Curiously, however, there have also been reports detailing the spontaneous development of IBD in patients receiving systemic IFN-β therapy for multiple sclerosis or hepatitis.

Methodology/Principal Findings

To investigate the effects of local administration of IFN-β on a murine model of colitis, we developed a transgenic Lactobacillus acidophilus strain that constitutively expresses IFN-β (La-IFN-β). While pretreatment of mice with control Lactobacillus (La-EV) provided slight protective benefits, La-IFN-β increased sensitivity to DSS. Analysis showed colitic mice pretreated with La-IFN-β had increased production of TNF-α, IFN-γ, IL-17A and IL-13 by intestinal tissues and decreased regulatory T cells (Tregs) in their small intestine. Examination of CD103⁺ dendritic cells (DCs) in the Peyer''s patches revealed that IFNAR1 expression was dramatically reduced by La-IFN-β. Similarly, bone marrow-derived DCs matured with La-IFN-β experienced a 3-fold reduction of IFNAR1 and were impaired in their ability to induce Tregs.

Conclusions/Significance

Our IFNAR1 expression data identifies a correlation between the loss/downregulation of IFNAR1 on DCs and exacerbation of colitis. Our data show that Lactobacillus secreting IFN-β has an immunological effect that in our model results in the exacerbation of colitis. This study underscores that the selection of therapeutics delivered by a bacterial vehicle must take into consideration the simultaneous effects of the vehicle itself.     

Multiple cytokines are released when blood from patients with tuberculosis is stimulated with Mycobacterium tuberculosis antigens

Background

Mycobacterium tuberculosis (Mtb) infection may cause overt disease or remain latent. Interferon gamma release assays (IGRAs) detect Mtb infection, both latent infection and infection manifesting as overt disease, by measuring whole-blood interferon gamma (IFN-γ) responses to Mtb antigens such as early secreted antigenic target-6 (ESAT-6), culture filtrate protein 10 (CFP-10), and TB7.7. Due to a lack of adequate diagnostic standards for confirming latent Mtb infection, IGRA sensitivity for detecting Mtb infection has been estimated using patients with culture-confirmed tuberculosis (CCTB) for whom recovery of Mtb confirms the infection. In this study, cytokines in addition to IFN-γ were assessed for potential to provide robust measures of Mtb infection.

Methods

Cytokine responses to ESAT-6, CFP-10, TB7.7, or combinations of these Mtb antigens, for patients with CCTB were compared with responses for subjects at low risk for Mtb infection (controls). Three different multiplexed immunoassays were used to measure concentrations of 9 to 20 different cytokines. Responses were calculated by subtracting background cytokine concentrations from cytokine concentrations in plasma from blood stimulated with Mtb antigens.

Results

Two assays demonstrated that ESAT-6, CFP-10, ESAT-6+CFP-10, and ESAT-6+CFP-10+TB7.7 stimulated the release of significantly greater amounts of IFN-γ, IL-2, IL-8, MCP-1 and MIP-1β for CCTB patients than for controls. Responses to combination antigens were, or tended to be, greater than responses to individual antigens. A third assay, using whole blood stimulation with ESAT-6+CFP-10+TB7.7, revealed significantly greater IFN-γ, IL-2, IL-6, IL-8, IP-10, MCP-1, MIP-1β, and TNF-α responses among patients compared with controls. One CCTB patient with a falsely negative IFN-γ response had elevated responses with other cytokines.

Conclusions

Multiple cytokines are released when whole blood from patients with CCTB is stimulated with Mtb antigens. Measurement of multiple cytokine responses may improve diagnostic sensitivity for Mtb infection compared with assessment of IFN-γ alone.     

The Placental Protein Syncytin-1 Impairs Antiviral Responses and Exaggerates Inflammatory Responses to Influenza

Background

Pregnancy increases susceptibility to influenza. The placenta releases an immunosuppressive endogenous retroviral protein syncytin-1. We hypothesised that exposure of peripheral monocytes (PBMCs) to syncytin-1 would impair responses to H1N1pdm09 influenza.

Methods and Findings

Recombinant syncytin-1 was produced. PBMCs from non-pregnant women (n=10) were exposed to H1N1pdm09 in the presence and absence of syncytin-1 and compared to responses of PBMCs from pregnant women (n=12). PBMCs were characterised using flow cytometry, release of interferon (IFN)-α, IFN-λ, IFN-γ, IL-10, IL-2, IL-6 and IL-1β were measured by cytometric bead array or ELISA. Exposure of PBMCs to H1N1pdm09 resulted in the release of IFN-α, (14,787 pg/mL, 95% CI 7311-22,264 pg/mL) IFN-λ (1486 pg/mL, 95% CI 756-2216 pg/mL) and IFN-γ (852 pg/mL, 95% CI 193-1511 pg/mL) after 48 hours. This was significantly impaired in pregnant women (IFN-α; p<0.0001 and IFN-λ; p<0.001). Furthermore, in the presence of syncytin-1, PBMCs demonstrated marked reductions in IFN-α and IFN-λ, while enhanced release of IL-10 as well as IL-6 and IL-1β.

Conclusions

Our data indicates that a placental derived protein, syncytin-1 may be responsible for the heightened vulnerability of pregnant women to influenza.     

Long-term alterations of cytokines and growth factors expression in irradiated tissues and relation with histological severity scoring

Purpose

Beside its efficacy in cancer treatment, radiotherapy induces degeneration of healthy tissues within the irradiated area. The aim of this study was to analyze the variations of proinflammatory (IL-1α, IL-2, IL-6, TNF-α, IFN-γ), profibrotic (TGF-β1), proangiogneic (VEGF) and stem cell mobilizing (GM-CSF) cytokines and growth factors in an animal model of radiation-induced tissue degeneration.

Materials and Methods

24 rats were irradiated unilaterally on the hindlimb at a monodose of 30 Gy. Six weeks (n = 8), 6 months (n = 8) and 1 year (n = 8) after irradiation the mediators expression in skin and muscle were analyzed using Western blot and the Bio-Plex® protein array (BPA) technology. Additional histological severity for fibrosis, inflammation, vascularity and cellularity alterations scoring was defined from histology and immnunohistochemistry analyses.

Results

A significant increase of histological severity scoring was found in irradiated tissue. Skin tissues were more radio-sensitive than muscle. A high level of TGF-β1 expression was found throughout the study and a significant relation was evidenced between TGF-β1 expression and fibrosis scoring. Irradiated tissue showed a chronic inflammation (IL-2 and TNF-α significantly increased). Moreover a persistent expression of GM-CSF and VEGF was found in all irradiated tissues. The vascular score was related to TGF-β1 expression and the cellular alterations score was significantly related with the level of IL-2, VEGF and GM-CSF.

Conclusion

The results achieved in the present study underline the complexity and multiplicity of radio-induced alterations of cytokine network. It offers many perspectives of development, for the comprehension of the mechanisms of late injuries or for the histological and molecular evaluation of the mode of action and the efficacy of rehabilitation techniques.     

Exercise Training Prevents TNF-α Induced Loss of Force in the Diaphragm of Mice

Rationale

Inflammatory cytokines like tumor necrosis factor alpha (TNF-α) are elevated in congestive heart failure and are known to induce the production of reactive oxygen species as well as to deteriorate respiratory muscle function.

Objectives

Given the antioxidative effects of exercise training, the aim of the present study was to investigate if exercise training is capable of preventing a TNF-α induced loss of diaphragmatic force in mice and, if so, to elucidate the potential underlying mechanisms.

Methods

Prior to intraperitoneal injection of TNF-α or saline, C57Bl6 mice were assigned to four weeks of exercise training or sedentary behavior. Diaphragmatic force and power generation were determined in vitro. Expression/activity of radical scavenger enzymes, enzymes producing reactive oxygen species and marker of oxidative stress were measured in the diaphragm.

Main Results

In sedentary animals, TNF-α reduced specific force development by 42% concomitant with a 2.6-fold increase in the amount of carbonylated α-actin and creatine kinase. Furthermore, TNF-α led to an increased NAD(P)H oxidase activity in both sedentary and exercised mice whereas xanthine oxidase activity and intramitochondrial ROS production was only enhanced in sedentary animals by TNF-α. Exercise training prevented the TNF-α induced force reduction and led to an enhanced mRNA expression and activity of glutathione peroxidase. Carbonylation of proteins, in particular of α-actin and creatine kinase, was diminished by exercise training.

Conclusion

TNF-α reduces the force development in the diaphragm of mice. This effect is almost abolished by exercise training. This may be a result of reduced carbonylation of proteins due to the antioxidative properties of exercise training.