microRNAs are differentially expressed in equine plasma of horses with osteoarthritis and osteochondritis dissecans versus control horses.

Osteoarthritis (OA) is a leading cause of lameness in horses with no effective disease-modifying treatment and challenging early diagnosis. OA is considered a disease of the joint involving the articular cartilage, subchondral bone, synovial membrane, and ligaments. Osteochondritis dissecans (OCD) is a joint disease consisting of focal defects in the osteochondral unit which may progress to OA later in life. MicroRNAs (miRNAs) have been recognized as small non-coding RNAs that regulate a variety of biological processes and have been detected in biological fluids. MiRNAs are currently investigated for their utility as biomarkers and druggable targets for a variety of diseases. The current study hypothesizes that miRNA profiles can be used to actively monitor joint health and differences in miRNA profiles will be found in healthy vs diseased joints and that differences will be detectable in blood plasma of tested horses. Five horses with OA, OCD, and 4 controls (C) had blood plasma and synovial fluid collected. Total RNA, including miRNA was isolated before generating miRNA libraries from the plasma of the horses. Libraries were sequenced at the Schroeder Arthritis Institute (Toronto). Differential expression analysis was done using DESeq2 and validated using ddPCR. KEGG pathway analysis was done using mirPath v.3 (Diana Tools). 57 differentially expressed miRNAs were identified in OA vs C plasma, 45 differentially expressed miRNAs in OC vs C plasma, and 21 differentially expressed miRNAs in OA vs OCD plasma. Notably, miR-140-5p expression was observed to be elevated in OA synovial fluid suggesting that miR-140-5p may serve as a protective marker early on to attenuate OA progression. KEGG pathway analysis of differentially expressed plasma miRNAs showed relationships with glycan degradation, glycosaminoglycan degradation, and hippo signaling pathway. Interestingly, ddPCR was unable to validate the NGS data suggesting that isomiRs may play an integral role in miRNA expression when assessed using NGS technologies.

Why the hype - What are microRNAs and why do they provide unique investigative, diagnostic, and therapeutic opportunities in veterinary medicine?

MicroRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression by inhibiting translation or inducing transcript degradation. MiRNAs act as fine-tuning factors that affect the expression of up to 60% of all mammalian protein coding genes. In contrast to proteins, there is widespread conservation of miRNA sequences across species. This conservation strongly suggests that miRNAs appeared early in evolution and have retained their functional importance. Cross-species conservation provides advantages when compiling candidate markers for health and disease compared to protein-based discoveries. This broad utility is accompanied by the emergence of inexpensive sequencing protocols for the identification of all RNAs in a sample (including miRNAs). With the use of miRNA mimics and antagonists, unique research questions can be answered in biological systems with 'cause and effect' methodology. MiRNAs are readily detectable in blood making them attractive candidates as biomarkers for disease. Here, we review their utility as biomarkers and their potential as therapeutic agents or targets to combat disease.

Pourquoi la frénésie ­ Que sont les microRNAs et pourquoi fournissent-ils des opportunités uniques pour investiguer, diagnostiquer et traiter en médecine vétérinaire? Les microRNAs (MiRNAs) sont de petits segments non-codants d'ARN qui régulent l'expression des gènes en inhibant la traduction ou en induisant la dégradation du transcript. Les MiRNAs agissent comme des facteurs d'ajustement fin qui affectent l'expression pouvant aller jusqu'à 60 % de tous les gènes mammaliens codant pour des protéines. Contrairement aux protéines, il y a un conservatisme étendu des séquences des miRNA à travers les espèces. Ce conservatisme suggère fortement que les miRNAs sont apparus tôt dans l'évolution et ont conservé leur importance fonctionnelle. La conservation inter-espèces fournie des avantages lors de la compilation de candidats marqueurs pour la santé et la maladie comparativement aux découvertes basées sur les protéines. Cette large utilité est accompagnée par l'émergence de protocoles de séquençage peu dispendieux pour l'identification de tous les ARNs dans un échantillon (incluant miRNAs). Avec l'utilisation d'imitations et d'antagonistes de miRNA, des questionnements rares en recherche peuvent être répondus dans des systèmes biologiques avec des méthodologies « cause et effet ¼. Les miRNAs sont facilement détectables dans le sang ce qui les rend des candidats attirants comme biomarqueurs de maladies. Ici, nous faisons une revue de leur utilité comme biomarqueurs et leur potentiel comme agents thérapeutiques ou cibles pour combattre des maladies.(Traduit par Dr Serge Messier).

Hypoxia enhances chondrogenic differentiation of human adipose tissue-derived stromal cells in scaffold-free and scaffold systems.

Human adipose-derived stromal cells (hASCs) possess the potential for chondrogenic differentiation. Recent studies imply that this differentiation process may be enhanced by culturing the cells in low oxygen tension in combination with three-dimensional (3D) scaffolds. We report the evaluation of the chondrogenic potential of hASC pellets in 5 and 21% O2 and as cell-scaffold constructs using a collagen I/III scaffold with chemical induction using TGF-ß3. hASCs from four human donors were cultured both in a micromass pellet system and in 3D collagen I/III scaffolds in either 5 or 21% O2. Chondrogenesis was evaluated by quantitative gene expression analysis of aggrecan, SOX9, collagen I, II and X and histological evaluation with H&E and toluidine blue staining. Induced pellets cultured in 5% O2 showed increased peripheral cellularity and matrix deposition compared with 21% O2. Induced pellets cultured in 5% O2 had increased control-adjusted gene expression of aggrecan, SOX9 and collagen I and decreased collagen X compared with 21% O2 cultures. Induced pellets had higher gene expression of aggrecan, SOX9, collagen I, II and X and increased ratios of collagen II/I and collagen II/X compared with controls. As for pellets, scaffold cultures showed cellularity and matrix deposition organized in a zonal manner as a function of the oxygen tension, with a cartilage-like morphology and matrix deposition peripherally in the 5% O2 group and a more centrally located matrix in the 21% O2 group. There were no differences in histology and gene expressions between pellet and scaffold cultures. Five percent O2 in combination with chondrogenic culture medium stimulated chondrogenic differentiation of hASCs in vitro. We observed similar patterns of differentiation and matrix disposition in pellet and scaffold cultures.