Plasma exosomal hsa-miR-339-5p is associated with NOD-like receptor family CARD domain-containing 5 in heart failure with reduced ejection fraction

Heart failure with reduced ejection fraction (HFrEF) is characterized by impaired cardiac function, with myocardial remodeling as a key pathological process. Plasma exosomal microRNAs (miRNAs) are promising non-invasive biomarkers for early diagnosis and treatment of HFrEF. From March to December 2024, plasma samples from 45 HFrEF patients and 45 healthy controls were collected at the First Affiliated Hospital of Anhui University of Traditional Chinese Medicine. Exosomes were isolated and subjected to high-throughput small RNA sequencing in a pilot cohort (n = 5). Differentially expressed miRNAs were identified and analyzed via Gene Ontology (GO) and KEGG pathway enrichment. Candidate miRNAs were validated by qRT-PCR in an expanded cohort (n = 40). In vitro, exosome uptake was evaluated in AC16 cardiomyocytes, and the regulatory role of hsa-miR-339-5p on its predicted target NLRC5 and the downstream PI3K/Akt pathway was investigated using fluorescence in situ hybridization (FISH), immunofluorescence, Western blotting, and functional assays. A total of 27 differentially expressed exosomal miRNAs were identified,, including 10 upregulated and 17 downregulated. qRT-PCR validated the upregulation of hsa-miR-22-5p, hsa-miR-181b-5p, and hsa-miR-339-5p, as well as the downregulation of hsa-miR-192-5p and hsa-miR-1469 (P 0.05). Bioinformatics analysis suggests that hsa-miR-339-5p targets NOD-like receptor family CARD domain-containing 5 (NLRC5) to regulate the PI3K/Akt signaling pathway, thereby influencing myocardial remodeling. qRT-PCR confirmed significantly higher hsa-miR-339-5p expression in HFrEF plasma exosomes compared to controls (P < 0.01). In vitro, overexpression of hsa-miR-339-5p in AC16 cells significantly increased COL1A1 and α-SMA levels (P < 0.01) and activated the PI3K/Akt pathway by suppressing NLRC5 (P < 0.05). Conversely, inhibition of hsa-miR-339-5p reduced these effects (P < 0.05). In vitro, overexpression of hsa-miR-339-5p in AC16 cells significantly increased COL1A1 and α-SMA levels (P < 0.01) and activated the PI3K/Akt pathway by suppressing NLRC5 (P < 0.05). Conversely, inhibition of hsa-miR-339-5p reduced these effects (P < 0.05). Plasma exosomal miRNAs show abnormal expression in HFrEF patients. hsa-miR-339-5p is a circulating miRNA associated with HFrEF and may serve as a biomarker for ventricular remodeling.