RNA sequencing reveals target genes of temporomandibular joint osteoarthritis in rats after the treatment of low-intensity pulsed ultrasound.

PURPOSE: To explore the potential molecular mechanism of low-intensity pulsed ultrasound (LIPUS) in the treatment of temporomandibular joint osteoarthritis (TMJ-OA), and identify the target genes for therapy of TMJ-OA.

METHODS: Rat TMJ-OA was induced by unilateral occlusal trauma (UOT). At 8 weeks, the experimental group rats were treated by LIPUS for 4 weeks (5 days every week). The cartilage was examined by histological techniques. Gene expression profile in control, placebo and LIPUS-treated group were measured by RNA sequencing (RNA-Seq). Gene oncology (GO) and kyoto encyclopedia of genes and genomes (KEGG) annotated were performed and ten differentially expressed genes (DEGs) were further validated in another individual by quantitative real-time polymerase chain reaction (qRT-PCR). Per-2, a circadian rhythm gene, was further confirmed by western blot.

RESULTS: TMJ-OA model was successfully established in rats through UOT. LIPUS played a positive role in attenuating the retrogression of cartilage. The cartilage lesion was determined by HE and Safranin-O staining. A significant and bran-new gene profile of 58 mRNAs was obtained from the RNA-Seq (LIPUS-treated/placebo) and generated approximately 30GB data. Annotation, functional classification and pathway of the data were analyzed based on GO and KEGG database and ten candidate DEGs were identified. Some of these genes were proved to be related to OA, such as matrix-degrading enzyme (ADAMTS-8), complement (C1qa, C3, C5aR1). Some were reported for the first time in TMJ-OA, such as circadian gene (Per-2, Dbp, Npas2 and Arntl). According to the results of qRT-PCR validation, the sequencing data was with a high degree of credibility. The circadian gene Per-2 was up-regulated by LIPUS in TMJ-OA on the mRNA and protein level.

CONCLUSION: This study reveals the potential therapeutic genes related to TMJ-OA. Especially the circadian Per-2 gene was detected up-regulated by the treatment of LIPUS. It provides us a precious, new target OA-related gene and further investigation of gene-function will provide us new insights in understanding the potential mechanical underling TMJ-OA.