Mounting evidence implicates extracellular vesicles (EVs) factors as mediators of cell therapy. Cardiosphere‐derived cells are cardiac‐derived cells with tissue reparative capacity. Activation of a downstream target of wnt/β‐catenin signalling, tryptophan 2,3 dioxygenase (TDO2) renders therapeutically inert skin fibroblasts cardioprotective. Here, we investigate the mechanism by which concentrated conditioned media from TDO2‐augmented fibroblasts (TDO2‐CCM) exert cardioprotective effects. TDO2‐CCM is cardioprotective in a mouse model of MI compared to CCM from regular fibroblasts (HDF‐CCM). Transcriptomic analysis of cardiac tissue at 24 h demonstrates broad suppression of inflammatory and cell stress markers in animals given TDO2‐CCM compared to HDF‐CCM or vehicle. Sequencing analysis of TDO2‐EV RNA demonstrated abundance of a small Y‐derived small RNA dubbed ‘NT4’. Purification of TDO2‐EVs by size‐exclusion chromatography and RNAse protection assays demonstrated that NT4 is encapsulated inside EVs. Consistently with TDO2‐CCM, macrophages exposed to NT4 showed suppression of the inflammatory and cell stress mediators, particularly p21/cdkn1a. NT4‐depleted TDO2‐CCM resulted in diminished immunomodulatory capacity. Finally, administration of NT4 alone was cardioprotective in an acute model of myocardial infarction. Taken together, these findings elucidate the mechanism by which TDO2 augmentation mediates potency in secreted EVs through enrichment of NT4 which suppresses upstream cell stress mediators including p21/cdkn1a.

Effective treatment approaches for patients with COVID-19 remain limited and are neither curative nor widely applicable. Activated specialized tissue effector extracellular vesicles (ASTEX) derived from genetically-enhanced skin fibroblasts, exert disease-modifying bioactivity in vivo in models of heart disease. Here we report that ASTEX antagonizes SARS-CoV-2 infection and its pathogenic sequelae: In human lung epithelial cells exposed to SARS-CoV-2, ASTEX is cytoprotective and antiviral biologics. Transcriptomic analysis implicated the mammalian target of rapamycin (mTOR) pathway, as infected cells upregulated mTOR signaling and pre-exposure to ASTEX attenuated it. The implication of mTOR signaling was further confirmed using mTOR inhibition and activation, which increased and decreased viral load, respectively. Dissection of ASTEX cargo identifies miR-10a and miR-16 as potential EV inhibitors of mTOR signaling. The findings reveal a novel, dual mechanism of action for ASTEX as a therapeutic candidate for COVID-19, with synergistic antiviral and cytoprotective benefits.