Role of tRNA derived fragments in renal ischemia–reperfusion injury

Ischemia–reperfusion injury (IRI) is one of the major causes of acute kidney injury (AKI). tRNA derived fragments (tRFs/tiRNAs) are groups of small noncoding RNAs derived from tRNAs. To date, the role of tRFs/tiRNAs in renal IRI has not been reported. Herein, we aimed to investigate the involvement of tRFs/tiRNAs in the occurrence and development of ischemia–reperfusion-induced AKI. Moderate/severe renal IRI mouse models were established by bilateral renal pedicle clamping. The tRF/tiRNA profiles of healthy controls and moderate/severe IRI-stressed kidney tissues were sequenced by Illumina NextSeq 500. Candidate differentially expressed tiRNAs were further verified by RT-qPCR. Biological analysis was also performed. Overall, 152 tRFs/tiRNAs were differentially expressed in the moderate ischemic injury group compared with the normal control group (FC &gt; 2, <i>p</i> &lt; 0.05), of which 47 were upregulated and 105 were downregulated; in the severe ischemic injury group, 285 tRFs/tiRNAs were differentially expressed (FC &gt; 2, <i>p</i> &lt; 0.05), of which 157 were upregulated, and 128 were downregulated. RT-qPCR determination of eight abundantly expressed tiRNAs was consistent with the sequencing results. Gene Ontology analysis for target genes of the tRFs/tiRNAs showed that the most enriched cell components, molecular functions and biological processes were Golgi apparatus, cytoplasmic vesicles, protein binding, cellular protein localization and multicellular organism development. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that these target genes were mainly involved in the natural killer cell mediated cytotoxicity pathway, citrate cycle, and regulation of actin cytoskeleton signaling pathway. Our results indicated that tRFs/tiRNAs were involved in renal IRI. These tRFs/tiRNAs may be effective partly <i>via</i> regulation of renal immunity, inflammation and metabolism processes. Candidate genes, including tiRNA-Gly-GCC-003, tiRNA-Lys-CTT-003, and tiRNA-His-GTG-002, might be potential biomarkers and therapeutic targets of ischemia–reperfusion injury-induced acute kidney injury.缺血再灌注损伤（Ischemia–reperfusion injury, IRI）是急性肾损伤（acute kidney injury, AKI）的主要致病因素之一。tRNA来源片段（tRNA derived fragments, tRFs/tiRNAs）是一类由tRNA剪切产生的小型非编码RNA。截至目前，tRFs/tiRNAs在肾脏IRI中的作用尚未见报道。本研究旨在探讨tRFs/tiRNAs在缺血再灌注诱导的AKI发生与发展过程中的参与机制。本研究通过双侧肾蒂夹闭法构建中度/重度肾脏IRI小鼠模型。采用Illumina NextSeq 500测序平台对健康对照组与中度/重度IRI应激的肾脏组织的tRF/tiRNA表达谱进行测序。通过逆转录实时定量聚合酶链反应（RT-qPCR）对筛选出的差异表达tiRNAs进行验证，并开展生物学功能分析。与正常对照组相比，中度缺血损伤组中共鉴定出152个差异表达的tRFs/tiRNAs（FC>2，p < 0.05），其中47个表达上调，105个表达下调；重度缺血损伤组中共鉴定出285个差异表达的tRFs/tiRNAs（FC>2，p < 0.05），其中157个表达上调，128个表达下调。针对8个高表达tiRNAs的RT-qPCR验证结果与测序结果一致。对tRFs/tiRNAs的靶基因进行基因本体论（Gene Ontology, GO）富集分析显示，富集程度最高的细胞组分、分子功能及生物过程分别为高尔基体、细胞质囊泡、蛋白质结合、细胞蛋白质定位以及多细胞生物体发育。京都基因与基因组百科全书（Kyoto Encyclopedia of Genes and Genomes, KEGG）富集分析结果表明，这些靶基因主要参与自然杀伤细胞介导的细胞毒性通路、三羧酸循环以及肌动蛋白细胞骨架信号调控通路。本研究结果证实，tRFs/tiRNAs参与了肾脏IRI的发生发展过程，其可能通过调控肾脏免疫、炎症及代谢过程发挥作用。候选靶基因包括tiRNA-Gly-GCC-003、tiRNA-Lys-CTT-003及tiRNA-His-GTG-002，有望成为缺血再灌注损伤诱导的急性肾损伤的潜在生物标志物与治疗靶点。