Therapeutically targetable Th17-derived miR-721 drives autoimmune myocarditis through PPARγ repression

Abstract BACKGROUND Myocarditis is an inflammatory cardiac disease in which Th17-driven immune responses contribute to progression toward dilated cardiomyopathy and heart failure. Current therapies mainly rely on corticosteroids but lack specificity, while the role of miR-721, synthesized by Th17 cells, remains largely unexplored in disease pathogenesis. METHODS We characterized the presence of mmu-miR-721 and its human homolog hsa-RNA-Chr8:96 in extracellular vesicles (EVs) secreted by Th17 cells from IL-17eGFP mice with experimental autoimmune myocarditis (EAM) and myocarditis patients. MxCre-Pparg fl/fl mice and luciferase reporter assays were used to validate the target genes of miR-721 and hsa-RNA-Chr8:96, respectively. The functional role of miR-721 in EAM was investigated by lentiviral vectors overexpression and inhibition using miRNA sponge molecules. Th17 responses and heart inflammation were assessed and echocardiography was performed after in vivo blockade of mmu-miR-721 in EAM mice. RESULTS Both mmu-miR-721 and hsa-RNA-Chr8:96 were encapsulated in EVs and secreted by Th17 cells of mice and patients with myocarditis. Overexpression of mmu-miR-721 in draining-lymph node cells from EAM mice inhibited Pparg transcription, leading to increased RORγt and IL-17 expression and promoting Th17 differentiation. In contrast, in the absence of Pparg , a target of miR-721, no differences in RORγt expression were observed, indicating that miR-721 promotes Th17 responses through repression of Pparg . Human PPARG was validated as a target gene of hsa-RNA-Chr8:96 and its overexpression in peripheral blood leukocytes downregulated PPARG mRNA levels, suggesting similar pathways involved in human pathology. In vivo blockade of mmu-miR-721 increased Pparg expression, reducing RORγt and IL-17 activation in T cells and leading to decreased leukocyte infiltration in the heart and improved cardiac function. CONCLUSIONS miR-721 is released by Th17 cells in EVs and promotes Th17 responses during myocarditis through repression of PPARγ, identifying this miRNA as both a mechanistic driver of disease and a potential therapeutic target. Graphical abstract Novelty and significance What is known? miR-721 and its human homolog are upregulated in the plasma of mice and humans with myocarditis Th17 cells synthesize miR-721 Mmu-miR-721 targets Pparg mRNA What new information does this article contribute? miR-721 is sorted into extracellular vesicles in the context of acute myocarditis miR-721 enhances Th17 differentiation via the Pparg / Rorc double inhibitory axis. Hsa-RNA-Chr8:96 targets human PPARG mRNA for degradation, inhibiting its expression Blockade of miR-721 dampens acute myocarditis development in vivo This study reveals a novel miRNA-based therapeutic strategy to inhibit Th17 responses and treat myocarditis. Using the experimental autoimmune myocarditis model, the authors unravel the mechanisms by which mmu-miR-721 can enhance Th17 responses and show how targeting this regulatory molecule could ameliorate the progression of the disease. Remarkably, this regulatory axis is suggested to be present in humans as well, since PPARG gene is validated as a target gene for hsa-RNA-Chr8:96. These findings highlights the potential of miR-721 not only as a diagnostic tool but also as a cell-specific therapeutic target to control Th17 responses in the clinical setting.