A Novel tsRNA, 5’tiRNA-GluCTC, Mediates Cardiomyocyte-Fibroblast Crosstalk to Promote Cardiac Fibrosis in Restrictive Cardiomyopathy

Restrictive cardiomyopathy (RCM) is characterized by pronounced cardiac fibrosis, leading to ventricular stiffening and diastolic dysfunction. While cardiomyocyte mutations are known triggers, the mechanisms initiating pro-fibrotic signaling remain elusive. This study investigates the role of cardiomyocyte-derived exosomes and specific tRNA-derived small RNAs (tsRNAs) in this pathogenic intercellular communication. Using a cTnIR193H knock-in mouse model, we observed pronounced cardiac fibrosis prior to the onset of heart failure, without significant cardiomyocyte apoptosis. This primary fibrotic response was mediated by a paracrine mechanism, as conditioned medium from mutant cardiomyocytes was sufficient to activate fibroblasts. Subsequent small RNA sequencing of cardiomyocyte-derived exosomes identified the tRNA-derived fragment 5'tiRNA-GluCTC as a significantly enriched species. Functional studies established this RNA as a critical mediator, demonstrating that its overexpression exacerbated fibrotic responses in vitro and induced fibrotic remodeling in wild-type mice, while its inhibition via a cardiac-targeted AAV sponge attenuated fibroblast activation and alleviated fibrosis in RCM mice. Mechanistically, 5'tiRNA-GluCTC is transferred to fibroblasts, directly targets the 3'UTR of Foxq1 to repress its expression, and consequently activates the Smad3/p-Smad3 signaling cascade. Collectively, these results delineate a 5'tiRNA-GluCTC/Foxq1/Smad3 signaling axis that drives fibrosis in RCM, uncovering a previously uncharacterized signaling cascade with therapeutic potential.