Reduced myocarditis following Coxsackievirus infection in cellular FLICE inhibitory protein--long form-transgenic mice.

Cellular FLICE inhibitory protein--long form (c-FLIP(L)) is a caspase-defective homologue of caspase-8 that blocks apoptosis by death receptors. c-FLIP(L) expression in T cells can also augment activation of the mitogen-activated protein kinase, extracellular signal-related kinase, as well as nuclear factor-kappaB. This contributes to increased production of interleukin-2 and CD25, resulting in hyperproliferation of T cells from c-FLIP(L)-transgenic mice. c-FLIP also heterodimerizes with and activates caspase-8, resulting in increased death of T cells and a selection of a T helper 2 cytokine profile. The effects of c-FLIP on cytolytic function of CD8(+) T cells have not been examined previously. We studied the cytolytic capacity of T cells from c-FLIP(L)-transgenic mice using an antigen-specific system, as well as the consequences during a viral immune response to Coxsackievirus B3 (CVB3). The increased T-cell receptor (TCR) signalling due to c-FLIP did not alter the cytolytic machinery but did reduce cytotoxicity because of decreased surface expression of TCR and CD8. It also produced a Tc2 cytokine profile. These effects of c-FLIP collectively served to diminish the severity of CVB3-induced myocarditis.

Coxsackievirus B3 induces T regulatory cells, which inhibit cardiomyopathy in tumor necrosis factor-alpha transgenic mice.

Innate immunity promotes both the generation of autoimmunity and immunoregulation of adaptive immunity. Transgenic mice expressing the tumor necrosis factor-alpha (TNF-alpha) gene under the cardiac myosin promoter (TNF1.6 mice) develop dilated cardiomyopathy. Transgenic mice show extensive cardiac inflammation, suggesting that immunopathogenic mechanisms may promote cardiomyopathy. Two coxsackievirus B3 (CVB3) variants infect and replicate in the heart. H3 variant is highly myocarditic, but H310A1 variant activates CD4(+) T regulatory cells, which protect against viral myocarditis. T-cell depletion of TNF1.6 mice using monoclonal anti-CD3 or anti-CD4 antibody significantly reduced heart size and plasma troponin I concentrations compared with control TNF1.6 mice. Cardiomyopathy in TNF1.6 mice correlates to a CD4(+)Th1 response and autoimmune IgG2a antibodies. TNF1.6 mice infected with H310A1 virus reduced heart size and cardiac inflammation corresponding to the activation of CD4(+)CD25(+)FoxP3(+) (T regulatory cells). Immunosuppression is dependent on IL-10 but not TGFbeta. Adoptive transfer of the CD4(+)CD25(+) cells from H310A1-infected mice into uninfected TNF1.6 recipients abrogated cardiomyopathy. Exogenous administration of recombinant TNF-alpha to H310A1-infected mice for 4 days abrogated immunosuppression. Cardiac enlargement in TNF1.6 mice is partly attributable to T-cell activation and humoral autoimmunity caused by cytokine expression. T regulatory cells induced by H310A1 virus abrogate autoimmunity caused by TNF-alpha overexpression. H3 virus infection induces high levels of systemic TNF-alpha, whereas H310A1 virus does not. The low TNF-alpha response during H310A1 infections is likely responsible for the T regulatory cell response in these animals.

Decay-accelerating factor (CD55) promotes CD1d expression and Vgamma4+ T-cell activation in coxsackievirus B3-induced myocarditis.

BALB/c mice infected with the H3 variant of Coxsackievirus B3 (CVB3) develop severe myocarditis which is initiated by up-regulation of CD1d during infection and CD1d-dependent activation of T cells expressing the Vgamma4 T cell receptor. Previous studies have shown that a mutant variant of the H3 virus which shows reduced binding avidity to one of the known CVB3 virus receptors, decay accelerating factor (DAF), fails to up-regulate CD1d or activate Vgamma4+ cells. To determine if DAF has a role in CD1d expression during infection or Vgamma4+ cell activation, BALB/c and BALB/c DAF-/- mice were infected with CVB3. Infected DAF-/- mice show modest increases in CD1d expression compared to infected wild-type BALB/c mice; and although total numbers of Vgamma4+ cells in the spleen are the same as in BALB/c mice, few Vgamma4+ IFNgamma+ cells are detected in infected DAF-/- animals. Vgamma4+ cell depletion protects infected BALB/c mice from myocarditis but does not protect infected DAF-/- animals, indicating that Vgamma4+ cells are not important to disease in these animals. Anti-CD8 depletion of CD8+ T cells protects infected BALB/c mice but aggravates disease in infected DAF-/- animals, indicating that the immunopathogenicity of viral myocarditis differs in the absence of the DAF virus receptor.

T cells expressing the Vgamma1 T-cell receptor enhance virus-neutralizing antibody response during coxsackievirus B3 infection of BALB/c mice: differences in male and female mice.

Coxsackievirus B3 infection causes severe cardiac inflammation in male but not female mice. CD3+ T cells and T cells expressing the Vgamma4 T cell receptor (TCR) predominate in the cardiac inflammatory cell infiltrate in infected male BALB/c mice. Infected females have mostly CD19+ (B lymphocyte) and Vgamma1+ cells. No significant differences in CD11b+ (monocyte) cells were observed between the sexes. Infected males showed a predominant CD4+Th1 (IFNgamma+) response, whereas females showed a predominant CD4+Th2 response. The importance of IFNgamma for myocarditis susceptibility and IL-4 for protection was confirmed using IFN-gamma-/- and IL-4-/- mice. Antibody depletion of Vgamma1+ cells augmented myocarditis susceptibility, whereas antibody depletion of Vgamma4+ cells was protective. Cardiac virus titers inversely correlated with virus neutralizing antibodies and showed that Vgamma1+ cells are important for virus neutralizing antibody response. IFNgamma affected the Vgamma4+ cell response in the heart, as IFNgamma-/- mice had few Vgamma4+ cells; but exogenous administration of recombinant IFNgamma to IFNgamma-/- mice restored myocarditis susceptibility, Th1 bias, and Vgamma4+ cell infiltration of the myocardium. These results demonstrate that two gammadelta+ T cell populations, Vgamma1+ and Vgamma4+, have different functions during myocarditis, in that Vgamma1+ cells promote humoral immunity and protection whereas Vgamma4+ cells are pathogenic.

Correlation between serum estradiol in the follicular phase of the ovarian cycle and decay acceleration factor (DAF) expression on red blood cells and coxsackiervirus B-3 induced hemagglutination in young cycling women.

Decay accelerating factor (DAF) is a widely distributed glycoprotein which aids in the inactivation of complement. DAF is also a cellular receptor for certain group B coxsackieviruses (CVB) and is responsible for the viral hemagglutinating activity for human red blood cells (RBC). Healthy, young female volunteers donated blood on days 11 and 22 of the ovarian cycle. Samples were categorized into luteal and follicular phases based on serum progesterone level (P4 either < 2.0 ng/mL, follicular; P4 > or = 2.0 ng/mL, luteal) and analyzed by flow cytomtery for DAF expression on RBC and CD21 + B lymphocytes. Cycling females showed significant variation in CVB-induced hemagglutination and % RBC or CD19 + cells which were DAF +. There was a strong correlation between serum estradiol levels and % RBC expressing DAF (P < 0.01) in the follicular, but not in the luteal ovarian phase. Infection of white blood cells with green-fluorescent protein CVB (GFP-CVB) showed a correlation between infectivity of CD19+ cells and DAF expression. This indicates that women may show differential susceptibility to CVB infection in the luteal and follicular phases of the ovarian cycle.

Role of CD1d in coxsackievirus B3-induced myocarditis.

The myocarditic (H3) variant of Coxsackievirus B3 (CVB3) causes severe myocarditis in BALB/c mice and BALB/c mice lacking the invariant J alpha 281 gene, but minimal disease in BALB/c CD1d(-/-) animals. This indicates that CD1d expression is important in this disease but does not involve the invariant NKT cell often associated with CD1d-restricted immunity. The H3 variant of the virus increases CD1d expression in vitro in neonatal cardiac myocytes whereas a nonmyocarditic (H310A1) variant does not. V gamma 4(+) T cells show increased activation in both H3-infected BALB/c and J alpha 281(-/-) mice compared with CD1d(-/-) animals. The activated BALB/c V gamma 4(+) T cells from H3-infected mice kill H3-infected BALB/c myocytes and cytotoxicity is blocked with anti-CD1d but not with anti-MHC class I (K(d)/D(d)) or class II (IA/IE) mAbs. In contrast, H3 virus-infected CD1d(-/-) myocytes are not killed. These studies demonstrate that CD1d expression is essential for pathogenicity of CVB3-induced myocarditis, that CD1d expression is increased early after infection in vivo in CD1d(+) mice infected with the myocarditic but not with the nonmyocarditic CVB3 variant, and that V gamma 4(+) T cells, which are known to promote myocarditis susceptibility, appear to recognize CD1d expressed by CVB3-infected myocytes.