Intranasal Vaccination Promotes Detrimental Th17-Mediated Immunity against Influenza Infection

Influenza disease is a global health issue that causes significant morbidity and mortality through seasonal epidemics. Currently, inactivated influenza virus vaccines given intramuscularly or live attenuated influenza virus vaccines administered intranasally are the only approved options for vaccination against influenza virus in humans. We evaluated the efficacy of a synthetic toll-like receptor 4 agonist CRX-601 as an adjuvant for enhancing vaccine-induced protection against influenza infection. Intranasal administration of CRX-601 adjuvant combined with detergent split-influenza antigen (A/Uruguay/716/2007 (H3N2)) generated strong local and systemic immunity against co-administered influenza antigens while exhibiting high efficacy against two heterotypic influenza challenges. Intranasal vaccination with CRX-601 adjuvanted vaccines promoted antigen-specific IgG and IgA antibody responses and the generation of polyfunctional antigen-specific Th17 cells (CD4⁺IL-17A⁺TNFα⁺). Following challenge with influenza virus, vaccinated mice transiently exhibited increased weight loss and morbidity during early stages of disease but eventually controlled infection. This disease exacerbation following influenza infection in vaccinated mice was dependent on both the route of vaccination and the addition of the adjuvant. Neutralization of IL-17A confirmed a detrimental role for this cytokine during influenza infection. The expansion of vaccine-primed Th17 cells during influenza infection was also accompanied by an augmented lung neutrophilic response, which was partially responsible for mediating the increased morbidity. This discovery is of significance in the field of vaccinology, as it highlights the importance of both route of vaccination and adjuvant selection in vaccine development     

Th1-Mediated Immunity against Helicobacter pylori Can Compensate for Lack of Th17 Cells and Can Protect Mice in the Absence of Immunization

Helicobacter pylori (H. pylori) infection can be significantly reduced by immunization in mice. Th17 cells play an essential role in the protective immune response. Th1 immunity has also been demonstrated to play a role in the protective immune response and can compensate in the absence of IL-17. To further address the potential of Th1 immunity, we investigated the efficacy of immunization in mice deficient in IL-23p19, a cytokine that promotes Th17 cell development. We also examined the course of Helicobacter infection in unimmunized mice treated with Th1 promoting cytokine IL-12. C57BL/6, IL-12 p35 KO, and IL-23 p19 KO mice were immunized and challenged with H. pylori. Protective immunity was evaluated by CFU determination and QPCR on gastric biopsies. Gastric and splenic IL-17 and IFNγ levels were determined by PCR or by ELISA. Balb/c mice were infected with H. felis and treated with IL-12 therapy and the resulting gastric bacterial load and inflammatory response were assessed by histologic evaluation. Vaccine induced reductions in bacterial load that were comparable to wild type mice were observed in both IL-12 p35 and IL-23 p19 KO mice. In the absence of IL-23 p19, IL-17 levels remained low but IFNγ levels increased significantly in both immunized challenged and unimmunized/challenged mice. Additionally, treatment of H. felis-infected Balb/c mice with IL-12 resulted in increased gastric inflammation and the eradication of bacteria in most mice. These data suggest that Th1 immunity can compensate for the lack of IL-23 mediated Th17 responses, and that protective Th1 immunity can be induced in the absence of immunization through cytokine therapy of the infected host.     

Th17/IL-17A Might Play a Protective Role in Chronic Lymphocytic Leukemia Immunity

Th17 cells, a recently discovered subset of T helper cells that secrete IL-17A, can affect the inflammation process autoimmune and cancer diseases development. The purpose of this study was to evaluate the role of Th17 cells and IL17A in biology of CLL. The study group included 294 untreated CLL patients in different clinical stages. Here, we show that higher Th17 and IL-17A values were associated with less advanced clinical stage of CLL. Th17 cells’ percentages in PB were lower in patients who died due to CLL during follow-up due to CLL (as compared to surviving patients) and in patients responding to first-line therapy with fludarabine-based regimens (as compared to non-responders). IL-17A inversely correlated with the time from CLL diagnosis to the start of therapy and was lower in patients who required treatment during follow-up. Th-17 and IL-17A values were lower in patients with adverse prognostic factors (17p and 11q deletion, CD38 and ZAP-70 expression). CLL patients with detectable IL-17A mRNA in T cells were in Rai Stage 0 and negative for both ZAP-70 and CD38 expression. Th17 percentages positively correlated with iNKT and adversely with Treg cells. The results of this study suggest that Th17 may play a beneficial role in CLL immunity.     

Th17细胞在肺部感染免疫中的作用

Th17细胞是近年来发现的一种新的效应T细胞亚群,在自身免疫性疾病和感染中发挥重要的作用,其分泌产生几种致炎细胞因子,包括新发现的细胞因子白细胞介素17。Th17产生的细胞因子与Th1、Th2不同并且与其相互对抗。Th17细胞很可能对防御胞外病原菌的感染及自身免疫性疾病产生影响。综述了Th17细胞产生的细胞因子及其在肺部感染免疫中的作用相关方面的进展。     

Th17 Immunity in Children with Allergic Asthma and Rhinitis: A Pharmacological Approach

Th17 cells and IL-17A play a role in the development and progression of allergic diseases. We analyzed the IL-17A levels in sputum supernatants (Ss), nasal wash (NW) and plasma (P) from Healthy Controls (HC) and children with Asthma/Rhinitis. We tested the expression of IL-17A, RORγ(t) and FOXP3 in peripheral blood T-lymphocytes from intermittent and mild-moderate asthma. The effect of Budesonide and Formoterol was tested “in vitro” on IL-17A, RORγ(t) and FOXP3 expression in cultured T-lymphocytes from mild-moderate asthma/persistent rhinitis patients, and on nasal and bronchial epithelial cells stimulated with NW and Ss from mild-moderate asthma/persistent rhinitis. Further, the effect of 12 weeks of treatment with Budesonide and Formoterol was tested “in vivo” in T-lymphocytes from mild-moderate asthma/persistent rhinitis patients. IL-17A was increased in Ss, NW and P from children with mild-moderate asthma compared with intermittent and HC. In cultured T-lymphocytes IL-17A and RORγ(t) expression were higher in mild-moderate asthma/persistent rhinitis than in mild-moderate asthma/intermittent rhinitis, while FOXP3 was reduced. Budesonide with Formoterol reduced IL-17A and RORγ(t), while increased FOXP3 in cultured T-lymphocytes from mild-moderate asthma/persistent rhinitis, and reduced the IL-8 release mediated by IL-17A present in NW and Ss from mild-moderate asthma/persistent rhinitis in nasal and bronchial epithelial cells. Finally, Budesonide with Formoterol reduced IL-17A levels in P and Ss, CD4⁺IL-17A⁺T-cells, in naïve children with mild-moderate asthma/persistent rhinitis after 12 weeks of treatment. Th17 mediated immunity may be involved in the airway disease of children with allergic asthma and allergic rhinitis. Budesonide with Formoterol might be a useful tool for its therapeutic control.     

High-Virulence Clone of Group B Streptococci Unable to Grow at High Temperatures Is Present in Serotypes Other Than Type III

Highly virulent clonotypes of serotype III seem to cause much of the perinatal morbidity and mortality attributed to Streptococcus agalactiae (group B streptococci, GBS), One of these clonal types, designated the “high-virulence clone” (HVC), was identified by its inability to grow at 40°C in a chemically defined medium. In the present study, this inability to grow at high temperatures was used as a marker to identify HVC in a sample of 286 Mexican GBS isolates. Forty-three isolates (15%) were identified as belonging to this clone: 15 were invasive isolates, 33 were serotype III (77%), and 10 were of serotypes other than type III (23%). These results demonstrate that HVC is more prevalent in Mexico than previously reported and that this clone is not restricted to serotype III isolates.     

A New Recombinant BCG Vaccine Induces Specific Th17 and Th1 Effector Cells with Higher Protective Efficacy against Tuberculosis

Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (Mtb) that is a major public health problem. The vaccine used for TB prevention is Mycobacterium bovis bacillus Calmette-Guérin (BCG), which provides variable efficacy in protecting against pulmonary TB among adults. Consequently, several groups have pursued the development of a new vaccine with a superior protective capacity to that of BCG. Here we constructed a new recombinant BCG (rBCG) vaccine expressing a fusion protein (CMX) composed of immune dominant epitopes from Ag85C, MPT51, and HspX and evaluated its immunogenicity and protection in a murine model of infection. The stability of the vaccine in vivo was maintained for up to 20 days post-vaccination. rBCG-CMX was efficiently phagocytized by peritoneal macrophages and induced nitric oxide (NO) production. Following mouse immunization, this vaccine induced a specific immune response in cells from lungs and spleen to the fusion protein and to each of the component recombinant proteins by themselves. Vaccinated mice presented higher amounts of Th1, Th17, and polyfunctional specific T cells. rBCG-CMX vaccination reduced the extension of lung lesions caused by challenge with Mtb as well as the lung bacterial load. In addition, when this vaccine was used in a prime-boost strategy together with rCMX, the lung bacterial load was lower than the result observed by BCG vaccination. This study describes the creation of a new promising vaccine for TB that we hope will be used in further studies to address its safety before proceeding to clinical trials.     

Simple Procedure for Production by Group C Streptococci of Phage-Associated Lysin Active Against Group A Streptococci

Phage-associated lysin of high potency was prepared by growing the host group C streptococcal strain 26RP66 in a semisynthetic medium. The lysin was stabilized by adding dithiothreitol and neutralized ethylenediaminetetraacetic acid (EDTA) to facilitate further concentration and partial purification. The lysin remained active when stored at -65 C for 1 year. Lysin was active against all strains of group A streptococci tested and was more active against living cells than heat-killed cells. The procedure outlined is practicable for most bacteriological research laboratories and does not require column purification or other complex biochemical procedures. It should be useful to any laboratory which requires small amounts of lysin to produce L-forms and protoplasts or to release streptococcal antigens.     

Reciprocity between Regulatory T Cells and Th17 Cells: Relevance to Polarized Immunity in Leprosy

T cell defect is a common feature in lepromatous or borderline lepromatous leprosy (LL/BL) patients in contrast to tuberculoid or borderline tuberculoid type (TT/BT) patients. Tuberculoid leprosy is characterized by strong Th1-type cell response with localized lesions whereas lepromatous leprosy is hallmarked by its selective Mycobacterium leprae specific T cell anergy leading to disseminated and progressive disease. FoxP3+ Regulatory T cells (Treg) which are essential for maintaining peripheral tolerance, preventing autoimmune diseases and limiting chronic inflammatory diseases also dampen proinflammatory T cells that include T helper 17 (Th17) cells. This study is aimed at evaluating the role of Treg cells in influencing other effector T cells and its relationship with the cytokine polarized state in leprosy patients. Peripheral blood mononuclear cells from of BT/TT (n = 15) and BL/LL (n = 15) patients were stimulated with M. leprae antigen (WCL) in presence of golgi transport inhibitor monensin for FACS based intracellular cytokine estimation. The frequency of Treg cells showed >5-fold increase in BL/LL in comparison to BT/TT and healthy contacts. These cells produced suppressive cytokine, IL-10 in BL/LL as opposed to BT/TT (p = 0.0200) indicating their suppressive function. The frequency of Th17 cells (CD4, CD45RO, IL-17) was, however, higher in BT/TT. Significant negative correlation (r = -0.68, P = 0.03) was also found between IL-10 of Treg cells and IL-17+ T cells in BL/LL. Blocking IL-10/TGF-β restored the IL-17+ T cells in BL/LL patients. Simultaneously, presence of Th17 related cytokines (TGF-β, IL-6, IL-17 and IL-23) decreased the number of FoxP3+ Treg cells concomitantly increasing IL-17 producing CD4+ cells in lepromatous leprosy. Higher frequency of Programmed Death-1/PD-1+ Treg cells and its ligand, PDL-1 in antigen presenting cells (APCs) was found in BL/LL patients. Inhibition of this pathway led to rescue of IFN-γ and IL-17 producing T cells. Results indicate that Treg cells are largely responsible for the kind of immunosuppression observed in BL/LL patients. This study also proves that Treg cells are profoundly affected by the cytokine milieu and this property may be utilized for benefit of the host.     

Human Anti-fungal Th17 Immunity and Pathology Rely on Cross-Reactivity against Candida albicans

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Immunofluorescent Identification of Type 12 Group A Streptococci

The fluorescent antibody (FA) conjugate prepared by labeling streptococcal M type 12 antibody with fluorescein isothiocyanate was found to exhibit considerable nonspecific FA staining with other group A M-serotypes. The cross-reactions could be reduced sufficiently or eliminated by the addition of adsorbed homologous blocking serum (AHB) but not by preimmune serum. The AHB was prepared by adsorbing type 12 antiserum with untreated homologous cells. Comparative staining with unblocked and AHB-blocked FA conjugates enabled type 12 streptococci from clinical specimens to be rapidly and accurately identified.     

Identification of Group A Streptococci by Direct Fluorometry

A simple direct fluorometric method for rapid identification of group A streptococci is described. The method permits the detection of the organism in mixed cultures without the aid of a microscope and is amenable to automated processing of specimens. Experience with the indirect fluorometric method revealed that nontrypsinized cells from a 10-fold dilution of overnight broth cultures could be stained with uniform brilliance with fluorescent antibody (1:15 dilution) and that fluorescent antibody dissociated from such cells at 55 C for 20 min gave serologically specific fluorometric values. With this information, it was possible to develop a simpler fluorometric test which gave results comparable to those obtained by conventional cultural-precipitin grouping techniques. In the direct test described, cultures from throat swabs were incubated overnight, and cells from a 10-fold dilution were stained with specific fluorescent antibody (1:50 dilution) and then rinsed. The stained specimens were transferred to a continuous-filter paper strip (Whatman 3 MM) and read serially in a Turner 110 fluorometer with Corning 5840 and Wratten 2A filters in place. The reagents used required careful standardization and testing to assure that fluorometric readings above a specified value would be indicative of the presence of group A streptococci.     

IL-17A and IL-2-Expanded Regulatory T Cells Cooperate to Inhibit Th1-Mediated Rejection of MHC II Disparate Skin Grafts

Several evidences suggest that regulatory T cells (Treg) promote Th17 differentiation. Based on this hypothesis, we tested the effect of IL-17A neutralization in a model of skin transplantation in which long-term graft survival depends on a strong in vivo Treg expansion induced by transient exogenous IL-2 administration. As expected, IL-2 supplementation prevented rejection of MHC class II disparate skin allografts but, surprisingly, not in IL-17A-deficient recipients. We attested that IL-17A was not required for IL-2-mediated Treg expansion, intragraft recruitment or suppressive capacities. Instead, IL-17A prevented allograft rejection by inhibiting Th1 alloreactivity independently of Tregs. Indeed, T-bet expression of naive alloreactive CD4+ T cells and the subsequent Th1 immune response was significantly enhanced in IL-17A deficient mice. Our results illustrate for the first time a protective role of IL-17A in CD4+-mediated allograft rejection process.     

Transduction of Rifampin Resistance in Group A Streptococci

Rifampin-resistant strains of group A streptococci were isolated as spontaneous mutants. Transduction analyses employing phage A25 showed the rifampin marker to be transferred with high frequency. The mutations conferring resistance to rifampin and streptomycin are not co-transducible.     

Effect of Resistance to Chloramphenicol on Bacteriophage Sensitivity of Group A Streptococci

Several phage hosts of group A streptococci became resistant to lysis by bacteriophage as a consequence of having acquired the ability to grow in the presence of chloramphenicol. The phage was adsorbed to the streptococcal cell, and P(32)-labeling of the phage showed that the phage genome penetrated the chloramphenicol (CM)- resistant cells as it did the parent cells. However, artificial lysis of the infected CM-resistant cells with chloroform or enzymes revealed no intracellular mature phage particles. Lysates of infected CM-resistant cells contained no phage-related antigenic materials which possessed serum-blocking power, although they were readily detected in lysates of infected parent cells. The CM-resistant cells were not lysogenized by the phage. Only cells resistant to more than 10 mug/ml of chloramphenicol were resistant to phage, and this threshold effect was taken as an indication of at least two different loci of chloramphenicol resistance on the streptococcal genome. Strains resistant to high levels of other antibiotics, such as streptomycin and erythromycin, showed no resistance to lysis by phage. Evidence indicated that the mutant cells were deficient in an essential function associated with the phage genome.