Renal dysfunction is a common complication of sepsis. Early diagnosis and prompt treatment of sepsis with renal insufficiency are crucial for improving patient outcomes. Diagnostic markers can help identify patients at risk for sepsis and AKI, allowing for early intervention and potentially preventing the development of severe complications. The aim of the present study was to investigate the expression difference of urinary microRNAs (miRNAs/miRs) in elderly patients with sepsis and secondary renal insufficiency, and to evaluate their diagnostic value in these patients. In the present study, RNA was extracted from urine samples of elderly sepsis-related acute renal damage patients and the expression profiles of several miRNAs were analyzed. In order to evaluate the expression profile of several miRNAs, urine samples from elderly patients with acute renal damage brought on by sepsis were obtained. RNA extraction and sequencing were then performed on the samples. Furthermore, multiple bioinformatics methods were used to analyze miRNA profiles, including differential expression analysis, and Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analysis of different miRNA target genes, to further explore miRNAs that are suitable for utilization as biomarkers. A total of four miRNAs, including hsa-miR-31-5p, hsa-miR-151a-3p, hsa-miR-142-5p and hsa-miR-16-5p, were identified as potential biological markers and were further confirmed in sepsis using reverse transcription-quantitative PCR. The results of the present study demonstrated that the four urinary miRNAs were differentially expressed and may serve as specific markers for prediction of secondary acute kidney injury in elderly patients with sepsis.