Single-cell transcriptomic analysis deciphers the inflammatory microenvironment characterized by CXCL9+ fibroblasts...

Immune-related myocarditis induced by immune checkpoint inhibitors (ICIs) is a rare immune-related adverse event (irAE) but is characterized by a high mortality rate. However, the specific pathological mechanisms underlying immune-related myocarditis remain largely unclear. In this study, we aimed to elucidate the inflammatory microenvironment within cardiac tissues affected by immune-related myocarditis at the single-cell level to identify potential therapeutic targets.

We performed single-cell RNA sequencing (scRNA-seq) on an endomyocardial biopsy specimen obtained from a patient with pancreatic neuroendocrine carcinoma who developed immune-related myocarditis following treatment with ICIs. Additionally, the scRNA-seq data of heart specimens from deceased donors without cardiovascular diseases were collected and applied as normal control. To validate our findings and assess their specificity to ICI-related pathology, we analyzed mouse scRNA-seq data, including controls, ICI-related myocarditis, viral myocarditis, and autoimmune myocarditis.

We found elevated proportions of lymphocytes, myeloid cells, and fibroblasts in the irAE group, suggesting an intensified inflammatory microenvironment in human immune-related myocarditis. Within the lymphocyte compartment, increased proportions of CD8 + T exhausted cells and CD8 + T proliferative cells were observed in the irAE group. The upregulated differentially expressed genes in myeloid cells in the irAE group were enriched in pro-inflammatory pathways, consistent with the observed metabolic shift from oxidative phosphorylation to glycolysis. CXCL9 + fibroblasts, characterized by the production of multiple pro-inflammatory cytokines and enriched in the JAK-STAT and TNFα signaling pathways, were predominantly found in the irAE group. Venous endothelial cells specifically expressing atypical chemokine receptor-1 (ACKR1) interacted with myeloid cells and CXCL9 + fibroblasts through the CXCL signaling pathway, facilitating chemokine transcytosis and leukocyte recruitment. Analysis of murine scRNA-seq data further supported these findings, revealing that exhausted CD8 + T cells and pro-inflammatory fibroblasts were uniquely enriched in ICI-related myocarditis, reflecting its distinct inflammatory microenvironment.

We elucidated the unique inflammatory microenvironment of immune-related myocarditis at the single-cell level. Our work revealed key cell subpopulations that were significantly implicated in inflammation, thus offering potential therapeutic targets.