Mincle-Mediated Coxsackievirus-B3 Infection Induced Fulminant Myocarditis Via Trained Immunity

Fulminant myocarditis (FM) is a lethal form of acute myocardial inflammation. To investigate this mechanism, we examined whether innate immunity trained by sequential exposure to lipopolysaccharide (LPS) and Coxsackievirus B3 (CVB3) could induce FM in CVB3-resistant C57BL/6J mice. Male 7–8-week-old C57BL/6J mice were used, and LPS + CVB3 double-hit treatment significantly increased mortality to 60%, accompanied by severe cardiac dysfunction (LVEF: 59.3% ± 7.8% vs. 69.7% ± 4.1%, p < 0.01) and markedly enhanced inflammatory cell infiltration compared to the CVB3-only group. In contrast, no significant inflammation was observed three weeks after LPS injection alone. Immunofluorescence analysis revealed that M1 macrophages dominated the inflammatory cell infiltration in the heart following CVB3 infection, with a more pronounced effect in the double-hit group than in the CVB3-only group. Furthermore, macrophage depletion alleviated inflammatory infiltration and improved cardiac function in the double-hit group. ATAC sequencing analysis demonstrated increased chromatin accessibility in bone marrow-derived macrophages three weeks after LPS injection. Single-cell RNA sequencing further indicated that macrophage-inducible C-type lectin (Mincle) plays a critical role in double-hit-induced FM and that Mincle blockade rescues cardiac dysfunction in mice with CVB3-induced myocarditis. CUT&Tag sequencing data revealed increased enrichment of histone H3 lysine 27 acetylation (H3K27ac) modifications on the Mincle gene in LPS-treated bone marrow macrophages. The p300 inhibitor protected against cardiac dysfunction in FM mice. Collectively, these findings demonstrate that low-dose LPS-induced trained immunity in macrophages exacerbates severe inflammatory infiltration and cardiac dysfunction in CVB3-infected C57BL/6J mice, leading to FM. This study provides a novel mechanistic insight into FM and establishes a mouse model for FM, identifying Mincle as a potential therapeutic target for this condition.