Doxorubicin (DOX) is a widely used anticancer drug, but its cardiotoxicity largely limits its clinical utilization. Circular RNA spindle and kinetochore-associated protein 3 (circ-SKA3) were found to be differentially expressed in heart failure patients. In this study, we investigated the role and mechanism of circ-SKA3 in DOX-induced cardiotoxicity.

The quantitative real-time polymerase chain reaction and western blot assays were applied to measure the expression of circ-SKA3, microRNA (miR) -1303, and toll-like receptor 4 (TLR4). The viability and apoptosis of AC16 cells were analyzed using cell counting kit-8, flow cytometry, and western blot assays. The interaction between miR-1303 and circ-SKA3 or TLR4 was verified using dual-luciferase reporter and RNA immunoprecipitation assays. Exosomes were collected from culture media by the use of commercial kits and then qualified by transmission electron microscopy.

The expression of circ-SKA3 and TLR4 was increased, whereas miR-1303 expression was decreased in DOX-treated AC16 cells. DOX treatment promoted cell apoptosis and inhibited cell viability in AC16 cells in vitro, which was partially reversed by circ-SKA3 knockdown, TLR4 silencing, or miR-1303 overexpression. Mechanistically, circ-SKA3 served as a sponge for miR-1303 to upregulate TLR4, which was confirmed to be a target of miR-1303. Additionally, circ-SKA3 contributed to DOX-induced cardiotoxicity through the miR-1303/TLR4 axis. Further studies suggested that circ-SKA3 was overexpressed in exosomes extracted from DOX-mediated AC16 cells, which could be internalized by surrounding untreated AC16 cells.

Circ-SKA3 enhanced DOX-induced toxicity in AC16 cells through the miR-1303/TLR4 axis. Extracellular circ-SKA3 was packaged into exosomes, and exosomal circ-SKA3 could function as a mediator in intercellular communication between AC16 cells.