3'tRNAAsp(GTC)-derived fragment links inflammation to post-transcriptional reprogramming in chondrocytes during osteoarthritis. [Primary Chondrocytes small RNA-seq]
Background/Aim: Transfer RNA-derived fragments (tRFs) are an emerging class of small non-coding RNAs that dynamically respond to metabolic stressors and drive different pathological processes, yet their role in osteoarthritis (OA) remains poorly explored. We aimed to define the tRF landscape in OA and investigate the function of 3'tRFAsp(GTC) in chondrocyte stress adaptation and translational control. Methods: Ex vivo, cartilage specimens from OA patients and healthy donors were analyzed by small RNA sequencing to define disease-associated tRF signatures. In vitro, primary chondrocytes derived from OA patients were treated with lipopolysaccharide (LPS) to mimic the pro-inflammatory environment of OA. Functional studies included antisense oligonucleotide (ASO)-mediated 3'tRFAsp(GTC) inhibition, AGO2-RNA immunopreicipitation (RIP), polysome profiling, stress granule (SG) immunofluorescence, and differential protein analysis. Computational target prediction and pathway enrichment were used to explore tRF-mediated regulatory networks. Results: Both ex vivo OA cartilage and LPS-treated OA chondrocytes displayed upregulation of 3'tRFAsp(GTC) and 5'tRFGlu(CTC), indicating a shared inflammatory tRF signature. Silencing 3'tRFAsp(GTC) attenuated LPS-induced COX2 and MMP13 expression, prevented ER stress, and blocked SG assembly. RIP confirmed selective recruitment of 3'tRFAsp(GTC) into AGO2 complexes. Polysome profiling revealed association with 40S ribosomal subunits, mediating translational arrest and influencing selective mRNA expression. Predicted targets of upregulated tRFs were enriched in stress-adaptive, proteostasis, and translational control pathways, whereas downregulated tRFs modulated mitochondrial processes. Conclusions: 3'tRFAsp(GTC) emerges as crucial regulator linking inflammatory stress to translational control and SG dynamics in OA. tRFs thus could represent a novel therapeutic targets to counteract chronic inflammatory stress in degenerative joint disease. Overall design: Small RNA-sequencing of primary human articular chondrocytes isolated from osteoarthritic cartilage, comparing lipopolysaccharide (LPS)-treated and untreated cells in a patient-paired experimental design.