Department of Cardiology, Jiangxi Provincial People's Hospital Affiliated to Nanchang University.
Department of Neurology, Jiangxi Provincial People's Hospital Affiliated to Nanchang University.
Int Heart J. 2021 Sep 30;62(5):1112-1123. doi: 10.1536/ihj.20-809. Epub 2021 Sep 17.
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.
阿霉素（DOX）是一种广泛应用的抗癌药物，但它的心脏毒性在很大程度上限制了其临床应用。研究发现，心力衰竭患者中环状 RNA 纺锤体和动粒相关蛋白 3（circ-SKA3）的表达存在差异。在本研究中，我们研究了 circ-SKA3 在 DOX 诱导的心脏毒性中的作用和机制。实时定量聚合酶链反应和 Western blot 检测用于测量 circ-SKA3、微小 RNA（miR）-1303 和 Toll 样受体 4（TLR4）的表达。使用细胞计数试剂盒-8、流式细胞术和 Western blot 检测分析 AC16 细胞的活力和凋亡。使用双荧光素酶报告和 RNA 免疫沉淀检测验证 miR-1303 与 circ-SKA3 或 TLR4 的相互作用。使用商业试剂盒从培养基中收集外泌体，并通过透射电子显微镜对其进行鉴定。在 DOX 处理的 AC16 细胞中，circ-SKA3 和 TLR4 的表达增加，而 miR-1303 的表达减少。DOX 处理在体外促进 AC16 细胞凋亡并抑制细胞活力，circ-SKA3 敲低、TLR4 沉默或 miR-1303 过表达部分逆转了这一作用。机制上，circ-SKA3 作为 miR-1303 的海绵，上调 TLR4，而 TLR4 被证实是 miR-1303 的靶标。此外，circ-SKA3 通过 miR-1303/TLR4 轴促进 DOX 诱导的心脏毒性。进一步的研究表明，circ-SKA3 在 DOX 介导的 AC16 细胞中提取的外泌体中过度表达，可被周围未处理的 AC16 细胞内化。Circ-SKA3 通过 miR-1303/TLR4 轴增强 DOX 诱导的 AC16 细胞毒性。细胞外 circ-SKA3 被包装到外泌体中，外泌体 circ-SKA3 可以作为 AC16 细胞之间细胞间通讯的介质。
