IL-12 is required for differentiation of pathogenic CD8+ T cell effectors that cause myocarditis

Abstract

Cardiac antigen-specific CD8(+) T cells are involved in the autoimmune component of human myocarditis. Here, we studied the differentiation and migration of pathogenic CD8(+) T cell effector cells in a new mouse model of autoimmune myocarditis. A transgenic mouse line was derived that expresses cardiac myocyte restricted membrane-bound ovalbumin (CMy-mOva). The endogenous adaptive immune system of CMy-mOva mice displays tolerance to ovalbumin. Adoptive transfer of naive CD8(+) T cells from the ovalbumin-specific T cell receptor-transgenic (TCR-transgenic) OT-I strain induces myocarditis in CMy-mOva mice only after subsequent inoculation with ovalbumin-expressing vesicular stomatitis virus (VSV-Ova). OT-I effector T cells derived in vitro in the presence or absence of IL-12 were adoptively transferred into CMy-mOva mice, and the consequences were compared. Although IL-12 was not required for the generation of cytolytic and IFN-gamma-producing effector T cells, only effectors primed in the presence of IL-12 infiltrated CMy-mOva hearts in significant numbers, causing lethal myocarditis. Furthermore, analysis of OT-I effectors collected from a mediastinal draining lymph node indicated that only effectors primed in vitro in the presence of IL-12 proliferated in vivo. These data demonstrate the importance of IL-12 in the differentiation of pathogenic CD8(+) T cells that can cause myocarditis.

T-bet Controls Pathogenicity of CTLs in the Heart by Separable Effects on Migration and Effector Activity

“…To explore the possible role of T-bet in the differentiation of pathogenic CD8 ϩ effectors in cardiac disease, we used a transgenic mouse model of CD8 ϩ T cell-mediated myocarditis (cMy-mOVA) (17,18). IL-12 plays a critical role in the differentiation of effector CD8 ϩ T cells that cause disease in cMy-mOVA mice (17).…”

“…IL-12 plays a critical role in the differentiation of effector CD8 ϩ T cells that cause disease in cMy-mOVA mice (17). This model facilitates the analysis of migration and effector function of a monospecific population of CD8 ϩ T cells that recognize an Ag expressed exclusively in the heart.…”

“…Cell suspensions were prepared from spleen and lymph node of OT-I T-bet ϩ/ϩ or OT-I T-bet Ϫ/Ϫ TCRtransgenic mice, and naive OT-I cells were isolated from the suspensions by CD8 magnetic beads (Miltenyi Biotec), as previously described (17,18). The naive T cells were stimulated in vitro with mitomycin-C (SigmaAldrich) treated C57BL/6 spleen cell suspensions (APCs) at a T cell:APC ratio of 1:10, and OVA peptide Ag (SIINFEKL) was added at a final concentration of 1 M/L.…”

“…into cMy-mOVA mice, as previously described (17,18). Cell doses transferred in this study ranged from 0.25 to 1.0 ϫ 10 6 cells per mouse as indicated.…”

“…Cytotoxic activity of effector OT-I cells was measured by coculturing the T cells with [ 51 Cr]sodium chromate loaded H-2K b -expressing EL4 target cells (American Type Culture Collection) at various E:T cell ratios. Target cells were pulsed with SIIN FEKL, and released 51 Cr in supernatants was detected by gamma counting, as described (17). Cytokine secretion by effector T cells was stimulated by culturing 2 ϫ 10 4 effector T cells with 2 ϫ 10 5 mitomycin C (SigmaAldrich) treated C57BL/6 spleen cells and SIINFEKL (1 g/ml) in 200-l microtiter wells, or by culturing 2 ϫ 10 4 T cells in microwells precoated with anti-CD3 (145:2C11; BD Pharmingen).…”