Assessment of Coxsackievirus B3 ts Mutants for Induction of Myocarditis in a Murine Model

Abstract

Ten temperature-sensitive ( ts ) mutants isolated from a myocarditis-inducing wild-type (WT) coxsackievirus B3 parent did not induce myocarditis in adolescent CD-1 mice. An avirulent prototype ts mutant from one of the three complementation groups adsorbed to murine cardiac tissue, as did WT virus. Heart tissues from mice inoculated with WT virus contained 100- to 1,000-fold more virus than heart tissues from mice inoculated with any of the three prototype ts mutants. WT virus exhibited a greater capsid stability and a higher efficiency of replication at 37°C than any of the three prototype ts mutants. All three prototype ts mutants induced less interferon in vivo than WT virus. Cell-mediated immune responses, assessed by the cell migration inhibition assay, were different in mice inoculated with WT virus when compared to ts 5 mutant virus. Peritoneal exudate cells from mice inoculated with WT but not ts 5 virus reacted specifically against antigens in WT virus HeLa cell lysates and antigens extracted with KCl from cardiac tissues of mice inoculated with WT virus. Cardiac tissues of mice inoculated with WT but not ts 5 virus contained KCl-extractable antigens which were able to specifically inhibit the migration of peritoneal exudate cells taken from mice immunized with WT virus. Therefore, ts 5 neither elicited a measurable cell-mediated immune response nor induced antigens in cardiac tissues which were immunoreactive with sensitized-(WT virus)-peritoneal exudate cells. Of 9 revertant viruses isolated from the 10 ts mutants, 5 showed covariance in ability to replicate at 39.5°C and capacity for induction of myocarditis. Some revertants exhibited a reduced capsid thermostability compared to WT virus but yet retained the capacity for induction of myocarditis. The data suggest that induction of myocarditis by coxsackievirus B3 variants depends on a combination of several variables, including capsid stability, capacity for replication at 37°C, and expression of the three identified genes. All three prototype ts mutants served as vaccine viruses in preventing myocarditis in adolescent mice subsequently challenged with WT virus. However, all three prototype ts mutants and their revertant variants retained partial to complete lethality in CD-1 neonates.

Assessment of cell-mediated immunity against coxsackievirus B3-induced myocarditis in a primate model (Papio papio)

“…1 (top). These lesions were morphologically dissimilar to those previously encountered in heart tissue from CVB3m-inoculated adolescent CD-1 mice (5,11,20,22). Myocarditis lesions in heart tissues of CVB3m-inoculated baboons were of a spider-like appearance, with mononuclear cells arranged in branches extending from a central mass of monocytes.…”

“…Myocarditis lesions in heart tissues of CVB3m-inoculated baboons were of a spider-like appearance, with mononuclear cells arranged in branches extending from a central mass of monocytes. In contrast, most myocarditic lesions in CVB3m-inoculated mice are quasi-spherical or quasi-rectangular in shape with irregular borders (20). Normal baboon heart tissue is shown in Fig.…”

“…The variant was propagated between 8 and 12 times in HeLa cells, and stock virus preparations containing 1 x 109 to 2 x 109 plaque-forming units (PFU) per ml were prepared in HeLa cells and stored at -90°C. Plaque assays were performed on monolayer cultures of HeLa cells as previously described (5,19,20). Preparation of KCl-extracted antigens.…”

Cellular immune responses in mice challenged with an amyocarditic variant of coxsackievirus B3

“…Mice inoculated with myocarditic strains of Coxsackievirus B3 (CVB3) develop myocarditis, ie, focal lesions containing necrotic cells and a mononuclear cell infiltrate, between 6 and 10 days postinoculation (pi) [Gauntt et al, , 1984Paque et al, 1978;Trousdale et al, 1979;Wilson et al, 1969;Woodruff and Woodruff, 19741. During days 10-14 pi, mononuclear cells disaggregate from the lesion area and fibrous scar tissue develops [Hashimoto and Komatsu, 1978;Roesing et al, 1979;Wilson et al, 1969;Woodruff, 19801.…”

“…During days 10-14 pi, mononuclear cells disaggregate from the lesion area and fibrous scar tissue develops [Hashimoto and Komatsu, 1978;Roesing et al, 1979;Wilson et al, 1969;Woodruff, 19801. Pathogenesis likely originates in virus-infected cells, but considerable evidence suggests that delayed hypersensitivity contributes to the disease process [Gauntt et al, , 1983aGudvangen et al, 1983;Huber et al, 1981;Paque and Gauntt, 1983;Paque et al, 1978Paque et al, , 1981Trousdale et al, 1979;Wong et al, 1977 a-c;Woodruff, 1979, 19801. Cytotoxic T lymphocytes (CTL) have been suggested by Woodruff and colleagues [Wong et al, 1977a,b;Gudvangen et al, 19831, by Huber and colleagues [Huber et al, 1980[Huber et al, , 1981[Huber et al, , 1984Huber and Job, 19831, and by Gauntt and colleagues Gudvangen et al, 19831 to have a major role in CVB3-induced myocarditis.…”