Persistent expression of cytokines in the chronic stage of CVB3-induced myocarditis in NMRI mice

Coxsackievirus B3 (CVB3)-induced myocarditis in NMRI mice represents a mode l for studying the pathogenesis of this chronic heart disease. Previously, we reported on specific cytokine patterns during the acute stage of myocard itis since cytokines are thought to play the important role in this cardiom yopathy. In this study. the expression of various cytokine mRNAs and CVB3-R NA kinetics was examined with particular emphasis oil the late phase of myo carditis, by using reverse transcriptase-polymerase chain reaction (RT-PCR) , in situ hybridization (ISH) and immunohistochemistry (IHC). In addition, replicating and persisting CVB3-RNAs were semiquantified by PCR-ELISA. Dist inct histopathological changes responsible for ongoing heart disease were f ound and characterized by increased fibrosis, persistent cellular infiltrat ion and degenerated necrotic myocytes. One of the most important findings o f this study was that the mRNA-expression of TNF-alpha. IL-1 alpha, interfe ron-gamma, IL-10, IL-18, macrophage inflammatory protein-1 alpha (MIP-1 alp ha), transforming growth factor-beta (TGF-beta) and inducible nitric oxide synthase (iNOS) persisted as long as 98 days after the virus infection. The induction of IL-10 as well as IFN-gamma, mRNAs was also verified by ISH an d IHC at days 28 and 98 p.i. The clearly apparent persistence of the viral genomes in the myocardium of infected mice was confirmed by seminested PCR, ISH, and PCR-enzyme linked immunoabsorbent assay (ELISA). showing the high est amount of viral RNA in myocardial cells at day 7 after infection. These data indicate that the persistence of viral RNA is associated with persist ently high levels of cytokine mRNAs which, when translated, could severely contribute to pathological changes and injury of connective tissue in the c hronic stage of myocarditis. ((C) 2001 Academic Press.