Background-The pathogenesis of myocarditis that occurs in Trypanosoma cruzi-infected mice is still poorly understood.Therefore, it is important to know the mediators that trigger leukocyte migration to the heart as well as the cellular source of these possible mediators. In this study, we investigated (1) NO synthase (NOS) induction, (2) NO synthesis, (3) trypanocidal activity, and (4) chemokine and cytokine mRNA expression by isolated cardiomyocytes infected with T cruzi. Methods and Results-Mouse cardiomyocytes were isolated, infected with T cruzi, and evaluated for induction of inducible NOS (iNOS), nitrite production, trypanocidal activity, and cytokine and chemokine mRNA expression. We found that T cruzi-infected murine embryonic cardiomyocytes produced nitrite and expressed mRNAs for the chemokines chemokine growth-related oncogene, monokine induced by interferon-␥, macrophage inflammatory protein-2, interferon-␥-inducible protein, RANTES, and monocyte chemotactic protein, for iNOS, and for the cytokines tumor necrosis factor (TNF)-␣ and interleukin (IL)-1␤. Separate addition of IL-1␤, interferon-␥, TNF-␣ or monocyte chemotactic protein, macrophage inflammatory protein-2, and interferon-␥-inducible protein, to cultured cardiomyocytes resulted in NO production but low trypanocidal activity. However, simultaneous addition of IL-1␤, interferon-␥, and TNF-␣ or the chemokines to cultures resulted in the induction of iNOS, high levels of nitrite, and a marked trypanocidal activity. The iNOS/L-arginine pathway mediated the latter activity, inasmuch as it was inhibited by treatment with N G -monomethyl-L-arginine. Conclusions-These results indicate that iNOS activation and the proinflammatory cytokines and chemokines produced by cardiomyocytes are likely to control parasite growth and cell influx, thus contributing to the pathogenesis of chagasic cardiomyopathy seen in T cruzi-infected mice. (Circulation. 2000;102:3003-3008.)

This study was carried out to determine the role of reactive nitrogen intermediates in Trypanosoma cruzi infection. In vitro, splenocytes obtained during the acute phase of infection produced elevated amounts of nitric oxide (NO) that were correlated with the resistance or susceptibility of the animals. In vivo, the levels of N02 plus N03 in plasma during the later phase of infection were higher in C57BV6 mice than in BALBLJc mice. The treatment of infected C57BL/6 mice with inhibitors of NO synthase increased parasitemia and mortality. Finally, we found that the NO donor drug S-nitroso-acetyl-penicillamine is able to kill trypomastigotes in vitro in the absence of any other cells, suggesting a direct NO-mediated killing of T. cruzi.

1 We investigated the mediators responsible for neutrophil migration induced by ovalbumin (OVA) in immunized mice and the mechanisms involved in their release. 2 OVA administration promoted dose-and time-dependent neutrophil migration in immunized, but not in non-immunized mice, which was mediated by leukotriene B 4 (LTB 4 ) and tumour necrosis factor (TNF)a, since it was inhibited by LTB 4 synthesis inhibitor (MK 886) ). 3 OVA-stimulated peritoneal cells from immunized mice released a neutrophil chemotactic factor which mimicked, in naive mice, neutrophil migration induced by OVA. 4 Supernatant chemotactic activity is due to TNFa and LTB 4 , since its release was inhibited by MK 886 (93%) and dexamethasone (90%), and signi®cant amounts of these mediators were detected. 5 TNFa and LTB 4 released by OVA challenge seem to act through a sequential mechanism, since MK 886 inhibited (88%) neutrophil migration induced by TNFa. Moreover, peritoneal cells stimulated with TNFa released LTB 4 . 6 CD 4 + T cells are responsible for TNFa release, because the depletion of this subset prevented the release of TNFa (control: 400+25; immunized: 670+40; CD 4 + depleted: 435+18 pg ml 71). 7 In conclusion, neutrophil migration induced by OVA depends on TNFa released by CD 4 + cells, which acts through an LTB 4 -dependent mechanism.

Articular inflammation is a major clinical burden in multiple inflammatory diseases, especially in rheumatoid arthritis. Biological anti-rheumatic drug therapies are expensive and increase the risk of systemic immunosuppression, infections, and malignancies. Here, we report that vagus nerve stimulation controls arthritic joint inflammation by inducing local regulation of innate immune response. Most of the previous studies of neuromodulation focused on vagal regulation of inflammation via the efferent peripheral pathway toward the viscera. Here, we report that vagal stimulation modulates arthritic joint inflammation through a novel "afferent" pathway mediated by the locus coeruleus (LC) of the central nervous system. Afferent vagal stimulation activates two sympatho-excitatory brain areas: the paraventricular hypothalamic nucleus (PVN) and the LC. The integrity of the LC, but not that of the PVN, is critical for vagal control of arthritic joint inflammation. Afferent vagal stimulation suppresses articular inflammation in the ipsilateral, but not in the contralateral knee to the hemispheric LC lesion. Central stimulation is followed by subsequent activation of joint sympathetic nerve terminals inducing articular norepinephrine release. Selective adrenergic beta-blockers prevent the effects of articular norepinephrine and thereby abrogate vagal control of arthritic joint inflammation. These results reveals a novel neuro-immune brain map with afferent vagal signals controlling side-specific articular inflammation through specific inflammatory-processing brain centers and joint sympathetic innervations.

In this study, the role of nitric oxide (NO) in neuronal destruction during acute-phase Trypanosoma cruzi infection was evaluated in male C57BL/6 (WT, wild-type) mice and knockout mice [inducible nitric oxide synthase (iNOS)(-/-) and interferon (IFN)(-/-)]. Selected animals were infected by intraperitoneal injection of 100 trypomastigote forms of the Y strain of T. cruzi. Others were injected intraperitoneally with an equal volume of saline solution and served as controls. Our findings support those of previous studies regarding myenteric denervation in acute-phase T. cruzi infection. In addition, we clearly demonstrate that, despite the fact that parasite nests and similar inflammatory infiltrate in the intestinal wall were more pronounced in infected iNOS(-/-) mice than in infected WT mice, the former presented no reduction in myenteric plexus neuron numbers. Neuronal nerve profile expression, as revealed by the general nerve marker PGP 9.5, was preserved in all knockout animals. Infected IFN(-/-) mice suffered no significant neuronal loss and there was no inflammatory infiltrate in the intestinal wall. On days 5 and 10 after infection, iNOS activity was greater in infected WT mice than in controls, whereas iNOS activity in infected knockout mice remained unchanged. These findings clearly demonstrate that neuronal damage does not occur in NO-impaired infected knockout mice, regardless of whether inflammatory infiltrate is present (iNOS(-/-)) or absent (IFN(-/-)). In conclusion, our observations strongly indicate that myenteric denervation in acute-phase T. cruzi infection is because of IFN-gamma-elicited NO production resulting from iNOS activation in the inflammatory foci along the intestinal wall.