Antigen Challenge Research Articles

The Impact of Advanced Age on Vaccine-Induced Tissue Immune Recall Responses

Abstract Infectious diseases account for one-third of deaths in individuals over the age of 65. Vaccines are a primary tool to combat infection, yet they are less effective in the aged population. Many have focused on vaccine-induced peripheral blood responses in old age; however, work from our lab demonstrates that immune responses to vaccination and infection in the periphery and local infection site (tissue) differ. To improve health outcomes in our aged population, we must study both systemic and tissue vaccine responses. We do so using a model of Mycobacterium bovis BCG vaccination and subsequent tuberculin skin test (TST) eight weeks after vaccination in adult and aged baboons. Vaccination generates BCG-specific immune cells that are recruited to the skin upon TST challenge, which we can study in skin biopsies reflecting tissue-specific responses. Three days or one week after TST we examine BCG-specific responses in skin biopsies. We also determine BCG-induced responses in the blood to compare tissue and systemic responses. We find increased oxidation and decreased production of immune proteins in aged baboon skin at the site of TST challenge, despite no alterations in antigen-specific peripheral blood mononuclear cell (PBMC) responses between age groups. Interestingly, cell migration assays show increased migration of PBMCs from aged BCG-vaccinated baboons, reinforcing the notion that systemic responses are not altered and that study of the tissue environment is important to understand responses to antigenic challenge with older age.

Signature of Enhanced Effector Function Defines Early Life Regulatory T Cells

Abstract The induction of self-tolerance and regulation of inflammatory immune responses is critical during early life when a myriad of primary antigenic challenges are encountered. Regulatory T Cells (Tregs) promote self-tolerance and function to modulate immune responses utilizing a variety of immunosuppressive mechanisms. Tregs are known to be disproportionately more abundant during early life, suggesting an essential role during immune development. We performed bulk RNA sequencing and multi-parameter flow cytometry on Tregs isolated from adult and pediatric (1 – 3 years of age) tissue (tonsils) and blood to discern the defining characteristics of early life Tregs. Early life Tregs were found to have an increased expression of genes associated with cell cycle (Ki67), effector and suppressive function (ICOS/CTLA4/IL10/GZMB/GZMA) and chemokine signaling (CCR5/CCR9) compared to adults. Importantly, distinctions in canonical effector function genes were apparent across all early life Tregs and not restricted to tissue site. Interestingly, CCR9, essential in gut-specific homing, was upregulated in both tissue and blood early life Tregs, suggesting a biasing towards intestinal immune homeostasis during childhood. We further explored the upregulation of genes related to cell cycle and found that early life Tregs exhibit higher proliferative capacity compared to both adult Tregs and early life CD4 conventional T cells, revealing a potential effector mechanism unique to early life. Early life Tregs are defined by a signature of augmented effector functionality enhancing their ability to respond to the abundant immunological challenges of infancy and childhood.

TCDD exposure differentially alters antigen-specific IgA levels when given before or after immunization

Abstract In mice, activation of the aryl hydrocarbon receptor (AhR) prior to primary antigen challenge can suppress antigen-specific serum IgM and IgG responses. Secondary antibody responses have been reported to be refractory to AhR-mediated suppression. Using ELISAs and female C57Bl/6 mice pretreated with the prototypic AhR agonist 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, 40 μg/Kg in peanut oil) one day prior to oral cholera toxin (CT) immunization (10 μg, repeated weekly), we demonstrated significant suppression of CT-specific IgA levels in the feces after two weeks (by 64.4%); however, fecal IgA levels recovered to control levels by week four. Unlike fecal IgA levels, CT-specific serum IgA levels remained significantly suppressed (by 37.1%) through week four. At that time-point, when compared to control mice, the spleens of TCDD-treated mice contained 40.4% fewer leukocytes, and contained 66.4% fewer IgA+ plasma cells (analyzed by ELISpot) (all significantly different). When TCDD exposure was delayed for three weeks after beginning CT immunization, in comparison to control mice, serum CT-specific IgA levels were significantly suppressed by 33.2%, but fecal CT-specific IgA levels were not different, as in TCDD pre-treated mice. Because enhanced migration of some T cells into the gut has been reported after AhR activation, one explanation for the differential effect of TCDD exposure on CT-specific IgA levels when given either before and after immunization could be enhanced migration of plasma cells away from reticuloendothelial tissues and into mucosal tissues.

Methotrexate reduces circulating Th17 cells and impairs plasmablast and memory B cell expansions following pneumococcal conjugate immunization in RA patients

Methotrexate (MTX) impairs antibody response after pneumococcal vaccination. We aimed to investigate differences in phenotypes of circulating B and T cells after pneumococcal conjugate vaccine (PCV) in rheumatoid arthritis (RA) patients on MTX (MTX group), RA without disease-modifying drugs (0DMARD), and controls (HC). MTX group (n = 11), 0DMARD (n = 12) and HC (n = 13) were studied. Blood samples were collected: before MTX, ≥ 4 weeks on stable MTX dose (prevaccination), and 7 days postvaccination (MTX group), and pre- and 7 days postvaccination (0DMARD and HC). Phenotypes of B- and T cell subsets were determined using flow cytometry. Serotype-specific IgG were quantified using multiplex bead assay, pre- and 4–6 weeks postvaccination. Concentrations of plasmablasts and switched memory B cells increased after PCV in HC (both p = 0.03) and the 0DMARD group (p = 0.01 and p = 0.02), but not in the MTX group. Postimmunization plasmablasts were lower in MTX group, compared to the 0DMARD group and HC (p = 0.002 and p < 0.001). Th17 cells decreased after MTX start (p = 0.02), and increased in HC after immunization (p = 0.01). Postimmunization plasmablasts correlated with mean antibody response ratio in all RA patients (R = 0.57, p = 0.035). Methotrexate reduced Th17 cells and blocked activation of plasmablasts and switched memory B cells following polysaccharide-protein conjugate antigen challenge in RA.

Tofacitinib inhibits CD4 T cell polarisation to Th1 during priming thereby leading to clinical impact in a model of experimental arthritis.

Janus kinases (JAK) are key cell membrane orientated tyrosine kinases that regulate inflammatory responses by transducing signals received by cytokine receptors that directly influence the polarisation and function of Th cells. Tofacitinib is a pan-JAK inhibitor approved for the treatment of RA. In this study, we explored the effects of tofacitinib in the outcomes of CD4+ T cell-dendritic cell (DC) interactions and their impact in autoimmune arthritis. The impact of tofacitinib in CD4+ T cell outcomes during priming or re-activation were analysed using antigen-specific in vitro and/or in vivo systems. A breach of self-tolerance model of arthritis was used to investigate the effects of tofacitinib in the outcomes of newly primed and antigen experienced CD4+ T cells. Tofacitinib inhibited Th1 polarisation during priming both in vitro and in vivo. In vitro, impaired T-bet expression and IFN-y production persisted upon secondary antigen challenge. Tofacitinib treatment during re-activation in vitro did not impact differentiation of antigen experienced CD4+ T cell towards Th1 phenotype. Moreover, JAK inhibition limited adaptive immune responses mediated by recently activated T cells and subsequent breach of self-tolerance in experimental arthritis. Our findings provide a novel mode of action for tofacitinib, demonstrating a potential therapeutic utility via homeostatic immune restoration in very early autoimmune arthritis.

Methamphetamine facilitates pulmonary and splenic tissue injury and reduces T cell infiltration in C57BL/6 mice after antigenic challenge

Methamphetamine (METH) is a strong addictive central nervous system stimulant. METH abuse can alter biological processes and immune functions necessary for host defense. The acquisition and transmission of HIV, hepatitis, and other communicable diseases are possible serious infectious consequences of METH use. METH also accumulates extensively in major organs. Despite METH being a major public health and safety problem globally, there are limited studies addressing the impact of this popular recreational psychostimulant on tissue adaptive immune responses after exposure to T cell dependent [ovalbumin (OVA)] and independent [lipopolysaccharide (LPS)] antigens. We hypothesized that METH administration causes pulmonary and splenic tissue alterations and reduces T cell responses to OVA and LPS in vivo, suggesting the increased susceptibility of users to infection. Using a murine model of METH administration, we showed that METH causes tissue injury, apoptosis, and alters helper and cytotoxic T cell recruitment in antigen challenged mice. METH also reduces the expression and distribution of CD3 and CD28 molecules on the surface of human Jurkat T cells. In addition, METH decreases the production of IL-2 in these T-like cells, suggesting a negative impact on T lymphocyte activation and proliferation. Our findings demonstrate the pleotropic effects of METH on cell-mediated immunity. These alterations have notable implications on tissue homeostasis and the capacity of the host to respond to infection.

Positive selection of IgG+ over IgM+ B cells in the germinal center reaction

Positive selection of high-affinity B cells within germinal centers (GCs) drives affinity maturation of antibody responses. Here, we examined the mechanism underlying the parallel transition from immunoglobulin M (IgM) to IgG. Early GCs contained mostly unswitched IgM+ B cells; IgG+ B cells subsequently increased in frequency, dominating GC responses 14-21days after antigen challenge. Somatic hypermutation and generation of high-affinity clones occurred with equal efficiency among IgM+ and IgG+ GC B cells, and inactivation of Ig class-switch recombination did not prevent depletion of IgM+ GC B cells. Instead, high-affinity IgG+ GC B cells outcompeted high-affinity IgM+ GC B cells via a selective advantage associated with IgG antigen receptor structure but independent of the extended cytoplasmic tail. Thus, two parallel forms of GC B-cell-positive selection, based on antigen receptor variable and constant regions, respectively, operate in tandem to ensure high-affinity IgG antibodies predominate in mature serum antibody responses.

Local immune responses to tuberculin skin challenge in Mycobacterium bovis BCG-vaccinated baboons: a pilot study of younger and older animals

Individuals over the age of 65 are highly susceptible to infectious diseases, which account for one-third of deaths in this age group. Vaccines are a primary tool to combat infection, yet they are less effective in the elderly population. While many groups have aimed to address this problem by studying vaccine-induced peripheral blood responses in the elderly, work from our lab and others demonstrate that immune responses to vaccination and infectious challenge may differ between tissue sites and the periphery. In this pilot study, we established an in vivo delayed-type hypersensitivity model of Mycobacterium bovis BCG vaccination and tuberculin skin test in two adult and two aged baboons. Vaccination generates BCG-specific immune cells that are recruited to the skin upon tuberculin challenge. We tested short term recall responses (8 weeks post-vaccination) and long term recall responses (25 weeks post-vaccination) by performing skin punch biopsies around the site of tuberculin injection. In short term recall responses, we found increased oxidation and decreased production of immune proteins in aged baboon skin at the site of TST challenge, in comparison to adult skin. Differences between adult and aged animals normalized in the long term response to tuberculin. In vitro, aged peripheral blood mononuclear cells had increased migration and functional responses to antigen-specific stimulation, suggesting that age-related changes in the tissue in vivo impairs aged immune recall responses to antigenic challenge. These findings highlight the impact of age-associated changes in the local tissue environment in memory recall responses, which may be more broadly applied to the study of other tissues. Moreover, these findings should be considered in future studies aimed at understanding and improving aging immune responses to vaccination and tissue challenge.

Increased Gene expression of CCL2/CCR2 axis in bronchial smooth muscles of allergen-challenged mice

PurposeAugmented bronchial smooth muscle (BSM) contraction is a cause of airway hyperresponsiveness (AHR) in asthma. Increasing evidence suggest that C-C motif chemokine 2 (CCL2) modulates smooth muscle contractility by activating its binding partner C-C chemokine receptor type 2 (CCR2). In the present study, changes in the gene expression of CCL2/CCR2 axis were determined in the BSMs of a murine model of allergic asthma. Materials and methodsThe ovalbumin (OA)-sensitized mice were repeatedly challenged with aerosolized OA to induce asthmatic reaction. Twenty-four hours after the last antigen challenge, total RNAs of the main BSM tissues and bronchoalveolar lavage fluids (BALFs) were obtained. ResultsOur published microarray data (GEO accession No. GSE116504) detected changes in gene expression associated with the chemokine signaling pathway (KEGG Map ID: 04062) in BSMs of mice with AHR induced by antigen exposure. Among them, quantitative RT-PCR analyses showed significant increase in mRNA expression of Ccl2 and Ccr2. Analysis of BALFs also revealed a significant increase in Ccl2 protein in the airways of the diseased animals. ConclusionIt is thus possible that, in association with the AHR, the CCL2/CCR2 axis is enhanced in the airways of allergic bronchial asthma.

Methamphetamine Compromises the Adaptive B Cell-Mediated Immunity to Antigenic Challenge in C57BL/6 Mice.

Methamphetamine (METH) is a substance of abuse that causes dysregulation of the innate and adaptive immunity in users. B cells are involved in the humoral component of the adaptive immunity by producing and secreting antibodies (Abs). METH modifies Ab production, although limited information on the impact of this psychostimulant on antigen (Ag)-specific humoral immune responses is available. Since T cell-dependent and T cell-independent Ags are involved in the activation of B lymphocytes, we explored the role of METH on humoral immunity to ovalbumin (OVA; T cell-dependent) and bacterial lipopolysaccharide (LPS; T cell-independent) in C57BL/6 mice. We demonstrated that METH extends the infiltration of B cells into pulmonary and splenic tissues 7 days post-Ag challenge. METH impairs Ab responses in the blood of animals challenged with OVA and LPS. Furthermore, METH diminishes the expression and distribution of IgM on B cell surface, suggesting a possible detrimental impact on users' humoral immunity to infection or autoimmunity.

PD-1 Blockade Modulates Functional Activities of Exhausted-Like T Cell in Patients With Cutaneous Leishmaniasis.

Patients infected by Leishmania braziliensis develop debilitating skin lesions. The role of inhibitory checkpoint receptors (ICRs) that induce T cell exhaustion during this disease is not known. Transcriptional profiling identified increased expression of ICRs including PD-1, PDL-1, PDL-2, TIM-3, and CTLA-4 in skin lesions of patients that was confirmed by immunohistology where there was increased expression of PD-1, TIM-3, and CTLA-4 in both CD4+ and CD8+ T cell subsets. Moreover, PDL-1/PDL-2 ligands were increased on skin macrophages compared to healthy controls. The proportions PD1+, but not TIM-3 or CTLA-4 expressing T cells in the circulation were positively correlated with those in the lesions of the same patients, suggesting that PD-1 may regulate T cell function equally in both compartments. Blocking PD-1 signaling in circulating T cells enhanced their proliferative capacity and IFN-γ production, but not TNF-α secretion in response to L. braziliensis recall antigen challenge in vitro. While we previously showed a significant correlation between the accumulation of senescent CD8+CD45RA+CD27- T cells in the circulation and skin lesion size in the patients, there was no such correlation between the extent of PD-1 expression by circulating on T cells and the magnitude of skin lesions suggesting that exhausted-like T cells may not contribute to the cutaneous immunopathology. Nevertheless, we identified exhausted-like T cells in both skin lesions and in the blood. Targeting this population by PD-1 blockade may improve T cell function and thus accelerate parasite clearance that would reduce the cutaneous pathology in cutaneous leishmaniasis.

Fetomaternal immune cross talk modifies T-cell priming through sustained changes to DC function

Prenatal exposure to infections can modify immune development. These environmental disturbances during early life potentially alter the incidence of inflammatory disorders as well as priming of immune responses. Infection with the helminth Schistosoma mansoni is widely studied for its ability to alter immune responsiveness and is associated with variations in coinfection, allergy, and vaccine efficacy in endemic populations. Exposure to maternal schistosomiasis during early life, even without transmission of infection, can result in priming effects on offspring immune responses to bystander antigenic challenges as related to allergic responsiveness and vaccination, with this article seeking to further clarify the effects and underlying immunologic imprinting. Here, we have combined a model of chronic maternal schistosomiasis infection with a thorough analysis of subsequent offspring immune responses to allergy and vaccination models, including viral challenge and steady-state changes to immune cell compartments. We have demonstrated that maternal schistosomiasis alters CD4+ responses during allergic sensitization and challenge in a skewed IL-4/B-cell-dominant response to antigenic challenge associated with limited inflammatory response. Beyond that, we have uncovered previously unidentified alterations to CD8+ T-cell responses during immunization that are dependent on vaccine formulation and have functional impact on the efficacy of vaccination against viral infection in a murine hepatitis B virus model. In addition to steady-state modifications to CD4+ T-cell polarization and B-cell priming, we have traced these modified CD8+ responses to an altered dendritic cell phenotype sustained into adulthood, providing evidence for complex priming effects imparted by infection via fetomaternal cross talk.

Exploiting Rational Assembly to Map Distinct Roles of Regulatory Cues during Autoimmune Therapy.

Autoimmune diseases like multiple sclerosis (MS), type 1 diabetes, and lupus occur when the immune system attacks host tissue. Immunotherapies that promote selective tolerance without suppressing normal immune function are of tremendous interest. Here, nanotechnology was used for rational assembly of peptides and modulatory immune cues into immune complexes. Complexes containing self-peptides and regulatory nucleic acids reverse established paralysis in a preclinical MS model. Importantly, mice responding to immunotherapy maintain healthy, antigen-specific B and T cell responses during a foreign antigen challenge. A therapeutic library isolating specific components reveals that regulatory nucleic acids suppress inflammatory genes in innate immune cells, while disease-matched peptide sequences control specificity of tolerance. Distinct gene expression profiles in cells and animals are associated with the immune signals administered in particulate and soluble forms, highlighting the impact of biophysical presentation of signals. This work provides insight into the rational manipulation of immune signaling to drive tolerance.

Targeted delivery of mycophenolic acid to the mesenteric lymph node using a triglyceride mimetic prodrug approach enhances gut-specific immunomodulation in mice

The mesenteric lymph nodes (MLN) are a key site for the generation of adaptive immune responses to gut-derived antigenic material and immune cells within the MLN contribute to the pathophysiology of a range of conditions including inflammatory and autoimmune diseases, viral infections, graft versus host disease and cancer. Targeting immunomodulating drugs to the MLN may thus be beneficial in a range of conditions. This paper investigates the potential benefit of targeting a model immunosuppressant drug, mycophenolic acid (MPA), to T cells in the MLN, using a triglyceride (TG) mimetic prodrug approach. We confirmed that administration of MPA in the TG prodrug form (MPA-TG), increased lymphatic transport of MPA-related species 83-fold and increased MLN concentrations of MPA >20 fold, when compared to MPA alone, for up to 4 h in mice. At the same time, the plasma exposure of MPA and MPA-TG was similar, limiting the opportunity for systemic side effects. Confocal microscopy and flow cytometry studies with a fluorescent model prodrug (Bodipy-TG) revealed that the prodrug accumulated in the MLN cortex and paracortex at 5 and 10 h following administration and was highly associated with B cells and T cells that are found in these regions of the MLN. Finally, we demonstrated that MPA-TG was significantly more effective than MPA at inhibiting CD4+ and CD8+ T cell proliferation in the MLN of mice in response to an oral ovalbumin antigen challenge. In contrast, MPA-TG was no more effective than MPA at inhibiting T cell proliferation in peripheral LN when mice were challenged via SC administration of ovalbumin. This paper provides the first evidence of an in vivo pharmacodynamic benefit of targeting the MLN using a TG mimetic prodrug approach. The TG mimetic prodrug technology has the potential to benefit the treatment of a range of conditions where aberrant immune responses are initiated in gut-associated lymphoid tissues.

Suppressive Effect of CYM50358 S1P4 Antagonist on Mast Cell Degranulation and Allergic Asthma in Mice.

Levels of sphingosine 1-phosphate (S1P), an intercellular signaling molecule, reportedly increase in the bronchoalveolar lavage fluids of patients with asthma. Although the type 4 S1P receptor, S1P4 has been detected in mast cells, its functions have been poorly investigated in an allergic asthma model in vivo. S1P4 functions were evaluated following treatment of CYM50358, a selective antagonist of S1P4, in an ovalbumin-induced allergic asthma model, and antigen-induced degranulation of mast cells. CYM50358 inhibited antigen-induced degranulation in RBL-2H3 mast cells. Eosinophil accumulation and an increase of Th2 cytokine levels were measured in the bronchoalveolar lavage fluid and via the inflammation of the lungs in ovalbumin-induced allergic asthma mice. CYM50358 administration before ovalbumin sensitization and before the antigen challenge strongly inhibited the increase of eosinophils and lymphocytes in the bronchoalveolar lavage fluid. CYM50358 administration inhibited the increase of IL-4 cytokines and serum IgE levels. Histological studies revealed that CYM50358 reduced inflammatory scores and PAS (periodic acid–Schiff)-stained cells in the lungs. The pro-allergic functions of S1P4 were elucidated using in vitro mast cells and in vivo ovalbumin-induced allergic asthma model experiments. These results suggest that S1P4 antagonist CYM50358 may have therapeutic potential in the treatment of allergic asthma.

Recruitment of inflammatory monocytes by senescent fibroblasts inhibits antigen-specific tissue immunity during human aging.

We have previously shown that healthy older adults exhibit reduced cutaneous immune responses during a varicella zoster virus (VZV) antigen challenge that correlated with a nonspecific inflammatory response to the injection itself. Here we found that needle damage during intradermal injections in older adults led to an increase in the number of cutaneous senescent fibroblasts expressing CCL2, resulting in the local recruitment of inflammatory monocytes. These infiltrating monocytes secreted prostaglandin E2, which inhibited resident memory T cell activation and proliferation. Pretreatment of older participants with a p38 mitogen-activated protein kinase inhibitor in vivo decreased CCL2 expression and inhibited monocyte recruitment and secretion of prostaglandin E2. This coincided with an increased response to VZV antigen challenge in the skin. Our results point to a series of molecular and cellular mechanisms that link cellular senescence, tissue damage, excessive inflammation and reduced immune responsiveness in human skin and demonstrate that tissue-specific immunity can be restored in older adults by short-term inhibition of inflammatory responses.

FOXO1 constrains activation and regulates senescence in CD8 T cells

Naive and memory T cells are maintained in a quiescent state, yet capable of rapid response and differentiation to antigen challenge via molecular mechanisms that are not fully understood. In naive cells, the deletion of Foxo1 following thymic development results in the increased expression of multiple AP-1 family members, rendering T cells less able to respond to antigenic challenge. Similarly, in the absence of FOXO1, post-infection memory T cells exhibit the characteristics of extended activation and senescence. Age-based analysis of human peripheral T cells reveals that levels of FOXO1 and its downstream target, TCF7, are inversely related to host age, whereas the opposite is found for AP-1 factors. These characteristics of aging also correlate with the formation of T cells manifesting features of cellular senescence. Our work illustrates a role for FOXO1 in the active maintenance of stem-like properties in T cells at the timescales of acute infection and organismal life span.

Bisphenol A, an endocrine-disruptor compund, that modulates the immune response to infections.

Bisphenol A (BPA) is an endocrine-disruptor compound that exhibits estrogenic activity. BPA is used in the production of materials such as polycarbonate plastics, epoxy resins and dental sealants. Whereas, the endocrine modulating activity of BPA and its effects on reproductive health have been widely studied, its effects on the function of the immune system are poorly characterized. This might be attributable to the different BPA doses used in a diversity of animal models. Moreover, most studies of the effect of BPA on the immune response are limited to in vitro and in vivo studies that have focused primarily on the impact of BPA on the number and proportion of immune cell populations, without evaluating its effects on immune function in response to an antigenic challenge or infectious pathogens. In this review, we discuss the current literature on the effects of BPA on the function of immune system that potentially increases the susceptibility to infections by the virtue of acting as a pro-inflammatory molecule. Thus, it appears that BPA, while by such an impact might be useful in the control of certain disease states that are helped by an inflmmatory response, it can worsen the prognosis of diseases that are adversely affected by inflammation.

NRF2 activator A-1396076 ameliorates inflammation in autoimmune disease models by inhibiting antigen dependent T cell activation

Nuclear factor (erythroid-derived 2) like 2 (NRF2) is a nuclear transcription factor activated in response to oxidative stress that induces a gene program that dampens inflammation and can limit cell damage that perpetuates the inflammatory response. We have identified A-1396076, a potent and selective NRF2 activator with demonstrated KEAP1 binding and modulation of cellular NRF2 mediated effects. In vivo administration of A-1396076 inhibits inflammation across several rodent models of autoimmunity when administered at or before the time of antigen challenge while also inducing NRF2 modulated gene transcription in the liver of the animals. It was not effective when administered after the time of antigen challenge or in a T cell independent model of arthritis induced by passive transfer of anti-collagen antibodies. A-1396076 inhibited antigen dependent T cell activation as measured by IFN-γ production in an ex vivo re-stimulation assay and following anti-CD3 challenge of MOG-sensitized mice. A-1396076 reduced costimulatory molecule expression on dendritic cells in the lungs of OVA LPS challenged mice suggesting that the mechanism of T cell inhibition was mediated at least partially by interfering with antigen presentation. These data suggest that NRF2 activation may be an effective strategy to dampen inflammation for treatment of autoimmune disease.

IFN-γ+ CD4+T cell-driven prophylactic potential of recombinant LDBPK_252400 hypothetical protein of Leishmania donovani against visceral leishmaniasis

Visceral leishmaniasis (VL) is a potentially fatal parasitic disease causing high morbidity and mortality in developing countries. Vaccination is considered the most effective and powerful tool for blocking transmission and control of diseases. However, no vaccine is available so far in the market for humans. In the present study, we characterized the hypothetical protein LDBPK_252400 of Leishmania donovani (LdHyP) and explored its prophylactic behavior as a potential vaccine candidate against VL. We found reduced hepato-splenomegaly along with more than 50% parasite reduction in spleen and liver after vaccination in mice. Protection in vaccinated mice after the antigen challenge correlated with the stimulation of antigen specific IFN-γ expressing CD4+T cell (~4.6 fold) and CD8+T cells (~2.1 fold) in vaccinated mice in compared to infected mice, even after 2–3 months of immunization. Importantly, antigen-mediated humoral immunity correlated with high antigen specific IgG2/IgG1 responses in vaccinated mice. In vitro re-stimulation of splenocytes with LdHyP enhances the expression of TNF-α, IFN-γ, IL-12 and IL-10 cytokines along with lower IL-4 cytokine and IL-10/IFN-γ ratio in vaccinated mice. Importantly, we observed ~3.5 fold high NO production through activated macrophages validates antigen mediated cellular immunity induction, which is critical in controlling infection progression. These findings suggest that immunization with LdHyP mount a very robust immunity (from IL-10 towards TFN-γ mediated responses) against L. donovani infection and could be explored further as a putative vaccine candidate against VL.