IL-10 Deficiency Reveals a Role for TLR2-Dependent Bystander Activation of T Cells in Lyme Arthritis.

T cells predominate the immune responses in the synovial fluid of patients with persistent Lyme arthritis; however, their role in Lyme disease remains poorly defined. Using a murine model of persistent Lyme arthritis, we observed that bystander activation of CD4+ and CD8+ T cells leads to arthritis-promoting IFN-γ, similar to the inflammatory environment seen in the synovial tissue of patients with posttreatment Lyme disease. TCR transgenic mice containing monoclonal specificity toward non-Borrelia epitopes confirmed that bystander T cell activation was responsible for disease development. The microbial pattern recognition receptor TLR2 was upregulated on T cells following infection, implicating it as marker of bystander T cell activation. In fact, T cell-intrinsic expression of TLR2 contributed to IFN-γ production and arthritis, providing a mechanism for microbial-induced bystander T cell activation during infection. The IL-10-deficient mouse reveals a novel TLR2-intrinsic role for T cells in Lyme arthritis, with potentially broad application to immune pathogenesis.

The immune response to picornavirus infection and the effect of immune manipulation on acute seizures.

Viral infection of the central nervous system can result in encephalitis. About 20% of individuals who develop viral encephalitis go on to develop epilepsy. We have established an experimental model where virus infection of mice with Theiler's murine encephalomyelitis virus (TMEV) leads to acute seizures, followed by a latent period (no seizures/epileptogenesis phase) and then spontaneous recurrent seizures-epilepsy. Infiltrating macrophages (CD11b+CD45hi) present in the brain at day 3 post-infection are an important source of interleukin-6, which contributes to the development of acute seizures in the TMEV-induced seizure model. Time course analysis of viral infection and inflammatory [CD11b+CD45hiLy-6Chi] and patrolling [CD11b+CD45hiLy-6Clow] monocyte and T cell infiltration into the brains of TMEV-infected C57BL/6J mice over the entire course of the acute viral infection was performed to elucidate the role of virus and the immune response to virus in seizures and viral clearance. The infiltrating inflammatory macrophages were present early following infection but declined over the course of acute viral infection, supporting a role in seizure development, while the lymphocyte infiltration increased rapidly and plateaued, advocating that they play a role in viral clearance. In addition, we showed for the first time that, while TMEV infection of RAG1-/- mice did not alter the number of mice experiencing acute seizures, TMEV infection of C57BL/6J mice depleted of macrophages resulted in a significant decrease in the number of mice experiencing seizures, again supporting a role for infiltrating macrophages in the development of acute seizures in the TMEV-induced seizure model.

Localized production of IL-10 suppresses early inflammatory cell infiltration and subsequent development of IFN-γ-mediated Lyme arthritis.

IL-10 is a nonredundant inflammatory modulator that suppresses arthritis development in Borrelia burgdorferi-infected mice. Infected C57BL/6 (B6) IL-10(-/-) mice were previously found to have a prolonged IFN-inducible response in joint tissue. Infection of B6 IL-10 reporter mice identified macrophages and CD4(+) T cells as the primary sources of IL-10 in the infected joint tissue, suggesting that early local production of IL-10 dampened the proarthritic IFN response. Treatment of B6 IL-10(-/-) mice with anti-IFN-γ reduced the increase in arthritis severity and suppressed IFN-inducible transcripts to wild-type levels, thereby linking dysregulation of IFN-γ to disease in the B6 IL-10(-/-) mouse. Arthritis in B6 IL-10(-/-) mice was associated with elevated numbers of NK cell, NKT cell, α/ß T cell, and macrophage infiltration of the infected joint. FACS lineage sorting revealed NK cells and CD4(+) T cells as sources of IFN-γ in the joint tissue of B6 IL-10(-/-) mice. These findings suggest the presence of a positive-feedback loop in the joint tissue of infected B6 IL-10(-/-) mice, in which production of inflammatory chemokines, infiltration of IFN-γ-producing cells, and additional production of inflammatory cytokines result in arthritis. This mechanism of arthritis is in contrast to that seen in C3H/He mice, in which arthritis development is linked to transient production of type I IFN and develops independently of IFN-γ. Due to the sustained IFN response driven by NK cells and T cells, we propose the B6 IL-10(-/-) mouse as a potential model to study the persistent arthritis observed in some human Lyme disease patients.

Endothelial cells and fibroblasts amplify the arthritogenic type I IFN response in murine Lyme disease and are major sources of chemokines in Borrelia burgdorferi-infected joint tissue.

Localized elevation in type I IFN has been uniquely linked to the severe Lyme arthritis that develops in C3H mice infected with the spirochete Borrelia burgdorferi. In this study, the dynamic interactions that result in generation of these responses were further examined in C3H mice carrying the type I IFN receptor gene ablation, which effectively blocks all autocrine/paracrine signaling crucial to induction of downstream effectors. Reciprocal radiation chimeras between C3H and IFNAR1⁻/⁻ mice implicated both radiation-sensitive and radiation-resistant cells of the joint tissue in the proarthritic induction of type I IFN. Ex vivo analysis of cells from the naive joint revealed CD45⁺ cells residing in the tissue to be uniquely capable of initiating the type I IFN response to B. burgdorferi. Type I IFN responses were analyzed in real time by lineage sorting of cells from infected joint tissue. This demonstrated that myeloid cells, endothelial cells, and fibroblasts were responsible for propagating the robust IFN response, which peaked at day 7 postinfection and rapidly resolved. Endothelial cells and fibroblasts were the dominant sources of IFN signature transcripts in the joint tissue. Fibroblasts were also the major early source of chemokines associated with polymorphonuclear leukocyte and monocyte/macrophage infiltration, thus providing a focal point for arthritis development. These findings suggest joint-localized interactions among related and unrelated stromal, endothelial, and myeloid cell lineages that may be broadly applicable to understanding the pathogeneses of diseases associated with type I IFN signature, including systemic lupus erythematosus and some rheumatoid arthritides.

Autologous cancer cell vaccination, adoptive T-cell transfer, and interleukin-2 administration results in long-term survival for companion dogs with osteosarcoma.

BACKGROUND: Osteosarcoma (OSA) in dogs is an aggressive bone tumor with frequent chemotherapy failure and translational relevance for human health. HYPOTHESIS/OBJECTIVES: We hypothesized that dogs with OSA could be treated safely by ex vivo activated T-cells that were generated by autologous cancer vaccination and supported by interleukin-2 (IL-2) treatment with survival more than twice that reported for amputation alone. ANIMALS: Osteosarcoma-bearing dogs (n = 14) were enrolled in a single-arm prospective trial after complete staging before amputation. Four healthy dogs also were treated in a safety study. METHODS: Autologous cancer cell vaccinations were administered intradermally and dogs underwent leukapheresis. Mononuclear cell products were stimulated ex vivo with a T-cell-activating agent. Activated product was transfused and 5 SC IL-2 injections were administered q48h. Dogs were monitored for metastasis by thoracic radiography every 3 months. RESULTS: Autologous cancer cell vaccine and activated cellular therapy (ACT) products were successfully generated. Toxicity was minimal after premedicants were instituted before ACT. With premedication, all toxicities were grade I/II. Median disease-free interval for all dogs was 213 days. One dog developed cutaneous metastasis but then experienced spontaneous complete remission. Median survival time for all dogs was 415 days. Five dogs survived >730 days. CONCLUSIONS AND CLINICAL IMPORTANCE: This immunotherapy protocol without cytotoxic chemotherapy is safe and tolerable. Compared to historical amputation reports, survival was notably prolonged in this group of patients. Additional prospective studies are warranted to elucidate active immunologic mechanisms and further improve disease response and survival.