Integrated transcriptomic and single-cell analysis reveals key immune-related genes and cellular crosstalk in the myocarditis microenvironment

Background: Myocarditis, an autoimmune disease characterized by inflammation of the heart muscle, can lead to impaired cardiac function. Understanding the cellular subpopulations within the myocarditis microenvironment is crucial for improving diagnosis and treatment strategies. Methods: We first performed differential expression analysis using bulk RNA sequencing data from myocarditis mouse models, to identify key genes and pathways. Subsequently, we integrated single-cell RNA sequencing data to explore different cell subpopulations through clustering analysis, utilizing GSEA, GSVA, and CellChat to examine their functions and interactions. Results: Differential expression analysis revealed 690 upregulated genes associated with immune regulation and inflammation, as well as pathways related to cardiovascular diseases, such as viral myocarditis and lipid metabolism. Analysis of fibroblasts indicated significant heterogeneity, including normal fibroblasts, inflammatory fibroblasts, myofibroblasts, and antigen-presenting fibroblasts. We observed a marked increase in Nfkbia + M1 macrophages during myocarditis, along with pre-exhausted Ccl5+ CD8+ cytotoxic T cells within T/NK cell subgroups, which receive antigen presentation signals from fibroblasts and macrophages. Conclusion: Our study reveals novel insights into the cellular complexity of the myocarditis microenvironment, particularly the identification of distinct fibroblast subpopulations, the role of Nfkbia + M1 macrophages in inflammation, and the pre-exhausted state of Ccl5+ CD8+ T cells. These findings not only provide a deeper understanding of the immune mechanisms underlying myocarditis but also highlight potential therapeutic targets and avenues for future research aimed at improving the diagnosis and treatment of myocarditis.