BCG Vaccination Induces M. avium and M. abscessus Cross-Protective Immunity

Pulmonary non-tuberculous mycobacterial (NTM) infections particularly caused by Mycobacterium avium complex (MAC) and Mycobacterium abscessus (MAB) are becoming major health problems in the U.S. New therapies or vaccines which will help prevent the disease, shorten treatment duration and/or increase treatment success rates are urgently needed. This study was conducted with the objective of testing the hypothesis that Bacillus Calmette Guerin (BCG), a vaccine used for prevention of serious forms of tuberculosis (TB) in children and adolescents in tuberculosis hyperendemic countries, induces cross-protective T cell immunity against Mycobacterium avium (MAV) and MAB. Human TB and NTM cross-protective T cells were quantified using flow cytometric assays. The ability of BCG expanded T cells to inhibit the intracellular growth of MAV and MAB was assessed in co-cultures with infected autologous macrophages. In both BCG-vaccinated and M. tuberculosis (Mtb)-infected mice, NTM cross-reactive immunity was measured using IFN-γ ELISPOT assays. Our results demonstrate the following key findings: (i) peripheral blood mononuclear cells from TB skin test-positive individuals contain MAV and MAB cross-reactive T cells, (ii) both BCG vaccination and Mtb infection of mice induce MAV and MAB cross-reactive splenic cells, (iii) BCG-expanded T cells inhibit intracellular MAV and MAB, (iv) CD4, CD8, and γδ T cells play important roles in inhibition of intracellular MAV and MAB and (v) BCG vaccination of healthy volunteers induces TB and NTM cross-reactive T cells. In conclusion, BCG-vaccination induces NTM cross-reactive immunity, and has the potential for use as a vaccine or immunotherapy to prevent and/or treat pulmonary NTM disease

Immune responses to gp82 provide protection against mucosal Trypanosoma cruzi infection

The potential use of the Trypanosoma cruzi metacyclic trypomastigote (MT) stage-specific molecule glycoprotein-82 (gp82) as a vaccine target has not been fully explored. We show that the opsonization of T. cruzi MT with gp82-specific antibody prior to mucosal challenge significantly reduces parasite infectivity. In addition, we investigated the immune responses as well as the systemic and mucosal protective immunity induced by intranasal CpG-adjuvanted gp82 vaccination. Spleen cells from mice immunized with CpG-gp82 proliferated and secreted IFN-γ in a dose-dependent manner in response to in vitro stimulation with gp82 and parasite lysate. More importantly, these CpG-gp82-immunized mice were significantly protected from a biologically relevant oral parasite challenge.Saint Louis University Department of Molecular MicrobiologyUniversidade Federal de São Paulo (UNIFESP) Escola Paulista de Medicina Departamento de Microbiologia, Imunologia e ParasitologiaUNIFESP, EPM, Depto. de Microbiologia, Imunologia e ParasitologiaSciEL

Mice with genetic deletion of group VIA phospholipase A2β exhibit impaired macrophage function and increased parasite load in Trypanosoma cruzi-induced myocarditis

Trypanosoma cruzi infection, which is the etiological agent of Chagas disease, is associated with intense inflammation during the acute and chronic phases. The pathological progression of Chagas disease is influenced by the infiltration and transmigration of inflammatory cells across the endothelium to infected tissues, which are carefully regulated processes involving several molecular mediators, including adhesion molecules and platelet-activating factor (PAF). We have shown that PAF production is dependent upon calcium-independent group VIA phospholipase A(2)β (iPLA(2)β) following infection of human coronary artery endothelial cells (HCAECs) with T. cruzi, suggesting that the absence of iPLA(2)β may decrease the recruitment of inflammatory cells to the heart to manage parasite accumulation. Cardiac endothelial cells isolated from iPLA(2)β-knockout (iPLA(2)β-KO) mice infected with T. cruzi demonstrated decreased PAF production compared to that by cells isolated from wild-type (WT) mice but demonstrated increases in adhesion molecule expression similar to those seen in WT mice. Myocardial inflammation in iPLA(2)β-KO mice infected with T. cruzi was similar in severity to that in WT mice, but the iPLA(2)β-KO mouse myocardium contained more parasite pseudocysts. Upon activation, macrophages from iPLA(2)β-KO mice produced significantly less nitric oxide (NO) and caused less T. cruzi inhibition than macrophages from wild-type mice. Thus, the absence of iPLA(2)β activity does not influence myocardial inflammation, but iPLA(2)β is essential for T. cruzi clearance

The absence of myocardial calcium-independent phospholipase a2γ results in impaired prostaglandin e2 production and decreased survival in mice with acute trypanosoma cruzi infection

Cardiomyopathy is a serious complication of Chagas' disease, caused by the protozoan parasite Trypanosoma cruzi. The parasite often infects cardiac myocytes, causing the release of inflammatory mediators, including eicosanoids. A recent study from our laboratory demonstrated that calcium-independent phospholipase A(2)γ (iPLA(2)γ) accounts for the majority of PLA(2) activity in rabbit ventricular myocytes and is responsible for arachidonic acid (AA) and prostaglandin E(2) (PGE(2)) release. Thus, we hypothesized that cardiac iPLA(2)γ contributes to eicosanoid production in T. cruzi infection. Inhibition of the isoform iPLA(2)γ or iPLA(2)β, with the R or S enantiomer of bromoenol lactone (BEL), respectively, demonstrated that iPLA(2)γ is the predominant isoform in immortalized mouse cardiac myocytes (HL-1 cells). Stimulation of HL-1 cells with thrombin, a serine protease associated with microthrombus formation in Chagas' disease and a known activator of iPLA(2), increased AA and PGE(2) release, accompanied by platelet-activating factor (PAF) production. Similarly, T. cruzi infection resulted in increased AA and PGE(2) release over time that was inhibited by pretreatment with (R)-BEL. Further, T. cruzi-infected iPLA(2)γ-knockout (KO) mice had lower survival rates and increased tissue parasitism compared to wild-type (WT) mice, suggesting that iPLA(2)γ-KO mice were more susceptible to infection than WT mice. A significant increase in iPLA(2) activity was observed in WT mice following infection, whereas iPLA(2)γ-KO mice showed no alteration in cardiac iPLA(2) activity and produced less PGE(2). In summary, these studies demonstrate that T. cruzi infection activates cardiac myocyte iPLA(2)γ, resulting in increased AA and PGE(2) release, mediators that may be essential for host survival during acute infection. Thus, these studies suggest that iPLA(2)γ plays a cardioprotective role during the acute stage of Chagas' disease

Absence of calcium‐independent phospholipase A2β impairs platelet‐activating factor production and inflammatory cell recruitment in Trypanosoma cruzi‐infected endothelial cells

Both acute and chronic phases of Trypanosoma cruzi (T. cruzi) infection are characterized by tissue inflammation, mainly in the heart. A key step in the inflammatory process is the transmigration of inflammatory cells across the endothelium to underlying infected tissues. We observed increased arachidonic acid release and platelet‐activating factor (PAF) production in human coronary artery endothelial cells (HCAEC) at up to 96 h of T. cruzi infection. Arachidonic acid release is mediated by activation of the calcium‐independent phospholipase A(2) (iPLA(2)) isoforms iPLA(2)β and iPLA(2)γ, whereas PAF production was dependent upon iPLA(2)β activation alone. Trypanosoma cruzi infection also resulted in increased cell surface expression of adhesion molecules. Increased adherence of inflammatory cells to T. cruzi‐infected endothelium was blocked by inhibition of endothelial cell iPLA(2)β or by blocking the PAF receptor on inflammatory cells. This suggests that PAF, in combination with adhesion molecules, might contribute to parasite clearing in the heart by recruiting inflammatory cells to the endothelium

Use of Leishmania major parasites expressing a recombinant Trypanosoma cruzi antigen as live vaccines against Chagas disease

INTRODUCTION: METHODS: We generated recombinant RESULTS: We demonstrate that mice inoculated with these recombinant TS-expressing DISCUSSION: Altogether, these data indicate tha

An immunoinformatic approach for identification of Trypanosoma cruzi HLA-A2-restricted CD8\u3csup\u3e+\u3c/sup\u3e T cell epitopes

Chagas disease is a major neglected tropical disease caused by persistent chronic infection with the protozoan parasite Trypanosoma cruzi. An estimated 8 million people are infected with T. cruzi, however only 2 drugs are approved for treatment and no vaccines are available. Thus there is an urgent need to develop vaccines and new drugs to prevent and treat Chagas disease. In this work, we identify T cell targets relevant for human infection with T. cruzi. The trans-sialidase (TS) gene family is a large family of homologous genes within the T. cruzi genome encoding over 1,400 members. There are 12 highly conserved TS gene family members which encode enzymatically active TS (functional TS; F-TS), while the remaining TS family genes are less conserved, enzymatically inactive and have been hypothesized to be involved in immune evasion (non-functional TS; NF-TS). We utilized immunoinformatic tools to identify HLA-A2-restricted CD8+ T cell epitopes conserved within F-TS family members and NF-TS gene family members. We also utilized a whole-genome approach to identify T cell epitopes present within genes which have previously been shown to be expressed in life stages relevant for human infection (Non-TS genes). Thirty immunogenic HLA-A2-restricted CD8+ T cell epitopes were identified using IFN-γ ELISPOT assays after vaccination of humanized HLA-A2 transgenic mice with mature dendritic cells pulsed with F-TS, NF-TS, and Non-TS peptide pools. The immunogenic HLA-A2-restricted T cell epitopes identified in this work may serve as potential components of an epitope-based T cell targeted vaccine for Chagas disease

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-gamma ELISPOT, serum IgG ELISAs, intracellular cytokine staining (IFN-gamma, TNF-alpha, 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-gamma 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 (P6 months post immunization. Also, these TS-specific T cells were better able to expand after in vitro restimulation.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.National Institutes of HealthFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Millennium Institute for Gene TherapySt Louis Univ, Dept Internal Med, St Louis, MO 63103 USAUniversidade Federal de São Paulo, Ctr Terapia Celular & Mol, Escola Paulista Med, São Paulo, BrazilSt Louis Univ, Dept Mol Microbiol, St Louis, MO 63103 USAUniv Fed Minas Gerais, Inst Ciencias Biol, Dept Microbiol, Belo Horizonte, MG, BrazilUniversidade Federal de São Paulo, Ctr Terapia Celular & Mol, Escola Paulista Med, São Paulo, BrazilNational Institutes of Health: RO1 AI040196CNPq: 420067/2005-1Web of Scienc

CD4+ and CD8+ T cells have opposing roles in breast cancer progression and outcome

The Cancer Immunoediting concept has provided critical insights suggesting dual functions of immune system during the cancer initiation and development. However, the dynamics and roles of CD4(+) and CD8(+) T cells in the pathogenesis of breast cancer remain unclear. Here we utilized two murine breast cancer models (4T1 and E0771) and demonstrated that both CD4(+) and CD8(+) T cells were increased and involved in immune responses, but with distinct dynamic trends in breast cancer development. In addition to cell number increases, CD4(+) T cells changed their dominant subsets from Th1 in the early stages to Treg and Th17 cells in the late stages of the cancer progression. We also analyzed CD4(+) and CD8(+) T cell infiltration in primary breast cancer tissues from cancer patients. We observed that CD8(+) T cells are the key effector cell population mediating effective anti-tumor immunity resulting in better clinical outcomes. In contrast, intra-tumoral CD4(+) T cells have negative prognostic effects on breast cancer patient outcomes. These studies indicate that CD4(+) and CD8(+) T cells have opposing roles in breast cancer progression and outcomes, which provides new insights relevant for the development of effective cancer immunotherapeutic approaches

Th17 cells are more protective than Th1 cells against the intracellular parasite Trypanosoma cruzi

Th17 cells are a subset of CD4+ T cells known to play a central role in the pathogenesis of many autoimmune diseases, as well as in the defense against some extracellular bacteria and fungi. However, Th17 cells are not believed to have a significant function against intracellular infections. In contrast to this paradigm, we have discovered that Th17 cells provide robust protection against Trypanosoma cruzi, the intracellular protozoan parasite that causes Chagas disease. Th17 cells confer significantly stronger protection against T. cruzi-related mortality than even Th1 cells, traditionally thought to be the CD4+ T cell subset most important for immunity to T. cruzi and other intracellular microorganisms. Mechanistically, Th17 cells can directly protect infected cells through the IL-17A-dependent induction of NADPH oxidase, involved in the phagocyte respiratory burst response, and provide indirect help through IL-21-dependent activation of CD8+ T cells. The discovery of these novel Th17 cell-mediated direct protective and indirect helper effects important for intracellular immunity highlights the diversity of Th17 cell roles, and increases understanding of protective T. cruzi immunity, aiding the development of therapeutics and vaccines for Chagas disease