5′tiRNA-32-LysCTT-11 Alleviates Sepsis-Induced Myocardial Injury by stabilizing Mfn2 and inhibiting mitochondrial dysfunction-mediated necroptosis

BACKGROUND: Transfer RNA-derived small RNAs (tsRNAs) represent a novel class of non-coding RNAs increasingly implicated in cardiovascular regulation. However, their roles in sepsis-induced cardiomyopathy (SICM) remain largely undefined. This study aimed to investigate the function and underlying mechanism of 5'tiRNA-32-LysCTT-11-a highly upregulated tsRNA in SICM-in modulating myocardial injury. METHODS: A murine model of sepsis was established via cecal ligation and puncture (CLP), and myocardial injury was assessed by serum CK-MB/LDH levels, histology, and cardiac function via echocardiography. In vitro, H9C2 cardiomyocytes were exposed to conditioned media (CM) from lipopolysaccharide (LPS)-stimulated macrophages. The expression of 5'tiRNA-32-LysCTT-11 was measured by qRT-PCR. Functional assays including CCK-8, LDH release, PI staining, JC-1, ATP, ROS detection, and MitoTracker staining were performed. Necroptosis was evaluated via MLKL phosphorylation; Mitochondria-associated endoplasmic reticulum membranes (MAMs) formation was assessed by dual-label immunofluorescence and Pacs2 expression. Bioinformatics analysis identified Mitofusin 2 (Mfn2) as a putative target, validated by Western blot, mRNA stability assay (Actinomycin D), and rescue experiments. FINDINGS: 5'tiRNA-32-LysCTT-11 was significantly upregulated in SICM. In vivo, its overexpression improved cardiac function and reduced injury biomarkers. In vitro, 5'tiRNA-32-LysCTT-11 mimics preserved mitochondrial integrity, reduced ROS and ATP depletion, suppressed MAM formation and necroptosis. Inhibitor transfection produced opposite effects. Mechanistically, 5'tiRNA-32-LysCTT-11 enhanced Mfn2 mRNA stability and protein expression. Silencing Mfn2 abrogated the protective effects, confirming its central role in the tsRNA's action. INTERPRETATION: 5'tiRNA-32-LysCTT-11 exerts cardioprotective effects during sepsis by stabilizing Mfn2 mRNA, preserving mitochondrial function, limiting MAMs formation, and suppressing necroptosis. These findings uncover a novel regulatory mechanism and suggest 5'tiRNA-32-LysCTT-11 as a promising therapeutic target in SICM.