Temperature-Sensitive Mutant of Coxsackievirus B3 Establishes Resistance in Neonatal Mice That Protects Them During Adolescence Against Coxsackievirus B3-Induced Myocarditis

摘要

Inoculation of neonatal CD-1 mice by multiple routes with an amyocarditic temperature-sensitive (ts) mutant (ts 1) derived from a myocarditic parent variant of coxsackievirus B3 (CVB3m) resulted in approximately half of the neonates surviving to adolescence. Challenge of the ts 1 survivors with CVB3m did not induce myocarditis, as assessed by histological examination of heart tissues. Virus was not detected in heart tissues of adolescent ts 1 survivors, but inoculation of these mice with CVB3m resulted in virus concentrations similar in titers to those found in CVB3m-inoculated normal adolescent mice. The ts 1 survivors did not contain detectable levels of anti-CVB3m neutralizing antibody, but upon challenge with CVB3m they produced antibody more rapidly and to higher titers than did normal CD-1 adolescents after primary inoculation with CVB3m. Cell-mediated immunity in ts 1 survivors was compared with that of normal mice after challenge with CVB3m. The capacity for production of migration inhibitory factor was assessed by the agarose droplet cell migration inhibition assay, using peritoneal exudate cells and a CVB3m cell lysate or KCl-extracted antigens from heart tissues of CVB3m-inoculated mice. Migration inhibitory factor activity was not detected in cultures of splenic leukocytes from ts 1 survivors of CVB3m-inoculated ts 1 survivors, but it was readily detected in cultures of splenic leukocytes from CVB3m-inoculated normal adolescent mice. The [³H]thymidine stimulation assay, performed with splenic lymphoid cells and purified CVB3m particles, revealed that lymphocytes from normal mice, whether inoculated with CVB3m or not, were not stimulated by CVB3m particle antigens, whereas lymphoid cells from a significantly higher proportion of ts 1 survivors, whether inoculated with CVB3m or not, responded with a stimulation index ≥2.0. The cells responding with positive stimulation were T lymphocytes. A higher proportion of normal mice and ts 1 survivors, both inoculated with CVB3_m, contained splenic cytotoxic T lymphocytes with higher reactivity against CVB3_m-infected neonatal skin fibroblasts than against normal skin fibroblasts, as assessed by a ⁵¹Cr release assay. The group of uninoculated ts 1 survivors present as a high proportion of individuals with cytotoxic T-lymphocyte reactivity against both uninoculated and CVB3_m-inoculated skin fibroblasts. However, ts 1 survivors and normal mice possessed the same proportions of splenic lymphocytes carrying either allele for Lyt 1 and Lyt 2 surface markers. The results suggest two mechanisms by which ts 1 survivors exhibit resistance to CVB3_m induction of myocarditis, namely, the rapid production of high-titered anti-CVB3_m neutralizing antibody in response to CVB3_m inoculation and altered cell-mediated immune responses against CVB3_m-induced viral or novel cellular antigens. The data are compatible with the notion that an immune deviation mechanism, thought to be controlled through a mechanism requiring suppressor cell activity which inhibits macrophage activation in ts 1 survivors, protects these mice from induction of myocarditis.