This work aims to investigate the function and mechanism of long non-coding RNA (lncRNA) cytoskeleton regulator RNA (CYTOR) in myocardial injury induced by sepsis. The sepsis-induced myocardial injury model in mice was established by intraperitoneal injection of LPS (10 mg/kg) in vivo, and cardiomyocyte H9c2 was treated with LPS to mimic sepsis in vitro. CYTOR expression and miR-24 expression were detected by qRT-PCR. After up-regulation or down-regulation of CYTOR and miR-24 expression in the H9c2 cells, and the viability of the cells was detected via MTT assay, and cell apoptosis was detected by TUNEL assay. Western blot was applied to determine the expression level of caspase 3, Bax and X-chromosome-linked inhibitor of apoptosis (XIAP). Interaction between CYTOR and miR-24 was determined by bioinformatics analysis, RT-PCR and dual luciferase reporter assay. Interaction between miR-24 and XIAP was determined through bioinformatics analysis, RT-PCR, western blot and dual luciferase reporter assay. CYTOR was markedly down-regulated. CYTOR interacted with miR-24, and negatively regulated its expression level. Over-expression of CYTOR or transfection of miR-24 inhibitors significantly promoted viability and inhibited apoptosis of H9c2 cells, while the knockdown of CYTOR and transfection of miR-24 mimics had opposite effects. CYTOR suppressed the expression level of apoptosis-related proteins, but miR-24 increased them. miR-24 directly targeted the 3’UTR of XIAP, and suppressed it, and XIAP was modulated indirectly by CYTOR. Down-regulation of CYTOR aggravates sepsis-induced cardiac injury via regulating miR-24 and XIAP.© 2020 John Wiley & Sons Ltd.