Interferon-inducible macrophages drive fulminant myocarditis via a TLR3-IRF1-ACOD1 mitochondrial fission axis

Background/Introduction Fulminant myocarditis (FM) is a rapidly progressive and often fatal inflammatory heart disease most commonly triggered by viral infection. The cellular drivers and molecular mechanisms underlying FM remain poorly defined, limiting effective therapeutic strategies. Purpose To identify pathogenic immune cell populations and molecular mechanisms driving systemic inflammation and cardiac injury in FM, with a focus on interferon-regulated mitochondrial signaling pathways in macrophage subsets. Methods We performed single-cell and single-nucleus RNA sequencing across nine organs in a coxsackievirus B3 (CVB3)-induced murine FM model, complemented by spatial transcriptomics and plasma proteomics in human FM samples. Genetic, pharmacological, and adoptive cell transfer approaches were used to interrogate the functional role of identified immune cell populations and signaling pathways. Results CVB3 infection induced profound systemic immune activation characterized by a cytokine storm originating from the heart, the primary viral target. Among cardiac immune populations, a distinct subset of interferon-inducible macrophages (IFNICs) expanded markedly and exhibited the strongest proinflammatory transcriptional profile in both mouse and human FM hearts. Transplantation of IFNICs was sufficient to exacerbate myocardial inflammation, impair cardiac function, and increase mortality, even in the absence of viral infection. Mechanistically, IFNIC pathogenicity was driven by a TLR3-IRF1-ACOD1 signaling axis. ACOD1 promoted inflammation independently of itaconate by acting as a mitochondrial scaffold that facilitated CMPK2-mediated phosphorylation of PGAM5, leading to DRP1-dependent mitochondrial fission and amplified inflammatory responses. Genetic or pharmacological inhibition of ACOD1, TLR3, or mitochondrial fission significantly improved survival and cardiac function in FM mice. Conclusion(s) Interferon-inducible macrophages are key pathogenic drivers of fulminant myocarditis. Targeting the TLR3-IRF1-ACOD1/PGAM5/CMPK2–mitochondrial fission axis represents a promising therapeutic strategy for FM.