Upregulation of the fibronectin transcript FN1–208 was identified in cartilage from patients with osteoarthritis (OA) and found to be unbeneficial for deposition of the cartilage matrix, according to study results published in Rheumatology.
Researchers identified FN1 transcripts associated with the OA process and investigated the effects of modulating FN1 expression and transcript ratio. They used previously assessed RNA sequencing data from the Research Osteoarthritis and Articular Cartilage study. Data were collected from lesioned and preserved OA cartilage samples from patients who underwent joint replacement surgery. The FN1 transcripts were detected using differential transcript expression analysis on all 27 FN1 transcripts annotated in the Ensembl database.
Knockdown experiments were carried out by transducing human primary chondrocytes with lentiviral particles containing small hairpin RNA targeting full-length FN1 transcripts and nontargeting control virus particles. Matrix deposition was then induced in a 3-dimensional in vitro neo-cartilage model. Alcian blue staining and dimethylmethylene blue assay was used to measure the effects of FN1 transcript ratio changes on sulphated glycosaminoglycan deposition. Reverse-transcription quantitative polymerase chain reaction was performed on 17 cartilage-relevant genes to investigate the effect of changes in FN1 transcript ratios on gene expression.
Within the FN1 transcriptome, 16 significantly upregulated FN1 transcripts were identified. The most significantly upregulated protein coding transcript was FN1–208, which encodes migration-stimulating factor and had not previously been associated with OA.
Downregulation of full-length FN1, which increased the FN1–208 ratio, led to a decrease in sulphated glycosaminoglycan deposition. Decreased expression of ACAN and COL2A1 and increased expression of ADAMTS-5, ITGB1, and ITGB5 was also observed.
One limitation of the study was that the analyses included only end-stage OA cartilage.
The study authors concluded, “[W]e identified multiple novel FN1 transcripts associated with OA pathophysiology, while signifying herein the potential role of FN1–208. We show that downregulation of full-length FN1 was unbeneficial for neo-cartilage deposition and resulted in upregulation of integrin β1 and β5 expression levels, likely via decreased availability of the classical [arginine-glycine-aspartic acid] integrin-binding site of fibronectin.”
van Hoolwerff M, Tuerlings M, Wijnen IJL, et al. Identification and functional characterization of imbalanced osteoarthritis associated fibronectin splice variants. Rheumatology (Oxford). Published online May 9, 2022. doi:10.1093/rheumatology/keac272