Equine synovial fluid small non-coding RNA signatures in early osteoarthritis

Abstract

BackgroundOsteoarthritis remains one of the greatest causes of morbidity and mortality in the equine population. The inability to detect pre-clinical changes in osteoarthritis has been a significant impediment to the development of effective therapies against this disease. Synovial fluid represents a potential source of disease-specific small non-coding RNAs (sncRNAs) that could aid in the understanding of the pathogenesis of osteoarthritis. We hypothesised that early stages of osteoarthritis would alter the expression of sncRNAs, facilitating the understanding of the underlying pathogenesis and potentially provide early biomarkers.MethodsSmall RNA sequencing was performed using synovial fluid from the metacarpophalangeal joints of both control and early osteoarthritic non-Thoroughbred horses. A group of differentially expressed sncRNAs was selected for further validation through qRT-PCR using an independent cohort of synovial fluid samples from control and early osteoarthritic horses. Bioinformatic analysis was performed in order to identify putative targets of the differentially expressed microRNAs and to explore potential associations with specific biological processes.ResultsResults revealed 22 differentially expressed sncRNAs including 13 microRNAs; miR-10a, miR-223, let7a, miR-99a, miR-23b, miR-378, miR-143 (and six novel microRNAs), four small nuclear RNAs; U2, U5, U11, U12, three small nucleolar RNAs; U13, snoR38, snord96, and one small cajal body-specific RNA; scarna3. Five sncRNAs were validated; miR-223 was significantly reduced in early OA and miR-23b, let-7a-2, snord96A and snord13 were significantly upregulated. Significant cellular functions deduced by the differentially expressed microRNAs included apoptosis (P < 0.0003), necrosis (P < 0.0009), autophagy (P < 0.0007) and inflammation (P < 0.00001). A conservatively filtered list of 57 messenger RNA targets was obtained; the top biological processes associated were regulation of cell population proliferation (P < 0.000001), cellular response to chemical stimulus (P < 0.000001) and cell surface receptor signalling pathway (P < 0.000001).ConclusionsSynovial fluid sncRNAs can be used as molecular biomarkers for early disease in equine osteoarthritic joints. The biological processes they regulate may play an important role in understanding early osteoarthritis pathogenesis. Characterising these dynamic molecular changes could provide novel insights on the process and mechanism of early osteoarthritis development and is critical for the development of new therapeutic approaches.

Small non-coding RNA landscape of extracellular vesicles from a post-traumatic model of equine osteoarthritis

“…They were altered at day 28 compared to day 0 in the same model and the same horses (48). MiR-23a was also increased in equine early OA SF (8) and increased in late versus early human OA SF (21). Mir-23a contributes to OA progression by directly targeting SMAD3 (76).…”

“…In our experiment we had access to samples from four horses but were able to examine, for the first-time in any species, temporal OA changes in EV characteristics and sncRNA cargo using SF and plasma and spanning seven time points from day 0-72 (study end). Whilst our groups have examined the sncRNA cargo in equine SF in early OA (8) and the temporal changes in microRNAs in SF from same equine osteochondral fragment model (39) using less time points here we tried to identify the changing sncRNA landscape within SF or plasma-derived EVs.…”

Use of Omics Data in Fracture Prediction; a Scoping and Systematic Review in Horses and Humans

“…Castanheira and their colleagues [ 23 ] revealed 22 differentially expressed synovial fluid small non-coding RNAs from the metacarpophalangeal joints of early osteoarthritis compared to the control, which may provide a potential usage of synovial fluid small non-coding RNAs as molecular biomarkers for early disease in equine osteoarthritic joints. Balaskas et al [ 22 ] demonstrated 83 differentially expressed small non-coding RNAs between the young and old equine chondrocytes that were normal on gross appearance, highlighting the importance of age-matched controls when planning miRNA studies.…”

“…Only a small number of miRNAs and targets were consistently identified as affected by disease; therefore, assessment of the potential of miRNAs as biomarkers for detection of fractures requires further research. The reviewed work was not substantial enough to identify an effect of age on miRNA expression and detection in horses, as many of the studies had too small a sample size to detect the influence of age, as has been reported subsequent to the literature search for this review [ 22 , 23 , 24 ].…”

“…As circulating miRNAs have been identified as potential biomarkers of fractures in osteoporosis [ 14 , 15 , 16 , 17 , 18 ] and are beginning to be evaluated in equine orthopaedic disease [ 19 , 20 , 21 , 22 , 23 , 24 ], the analysis of circulating miRNAs represents an exciting opportunity to study musculoskeletal health, yet currently, there are no standardised pre-analytical, analytical and post-analytical guidelines to allow for comparison between studies. It is therefore timely to conduct a review of the literature.…”