The Role Of Lubricin And Galectins In Osteoarthritis And Cartilage Lubrication
The aim of this thesis was to investigate how lubricin and galectins function within the synovial environment to maintain joint integrity and enhance articular cartilage lubrication. The specific aims of the research were to 1) evaluate how lubricin gene and glycoprotein expression were altered in equine carpal osteoarthritis (OA); 2) investigate the interactions between synovial lubricin and galectins-1 and -3 to determine if lubricin and galectins could synergistically enhance articular cartilage boundary lubrication and; 3) evaluate equine mesenchymal stem cell (MSC) galectin expression and the role of galectins-1 and -3 on MSC properties. Lubricin gene expression was quantified in joint tissues obtained from horses with experimental and naturally occurring carpal OA. Experimental OA was induced by creating a 6mm osteochondral chip fracture within the middle carpal joint (MCJ) of horses followed by high-speed treadmill exercise, and joints were assessed 70 days post-fragmentation using gross, histological and immunohistochemical parameters. Synovial fluid lubricin concentrations were serially assessed in experimental horses and at arthroscopy in clinical cases. Lubricin gene expression decreased in OA cartilage but increased in synovial membrane. In both experimental and naturally occurring OA, synovial fluid lubricin levels increased, peaking at 21 days postfragmentation in experimental horses. Increased lubricin immunolocalization was observed at sites of cartilage damage, including fibrillation, clefts, and at sites of fibrocartilaginous repair tissue. Glycan phenotyping of equine lubricin and fluorescent lectin staining of cartilage revealed the presence of core-1 O-linked oligosaccharides on lubricin capable of binding to galectins. Biochemical assays demonstrated strong binding between lubricin and galectin-3, suggesting a potential role for galectin-3 in stabilization of the articular cartilage lubricin boundary layer. Boundary lubrication of cartilage was measured using a custom friction testing apparatus in the presence of recombinant galectins. Galectin-3 enhanced boundary lubrication, but only in the presence of lubricin. Galectin-1 and Galectin-3C, an oligomerization-incompetent mutant, did not enhance lubrication. Equine galectins-1 and -3 were cloned and recombinantly produced, and gene expression was evaluated in equine MSCs. MSCs endogenously expressed high levels of galectins and elevated galectin-1: galectin-3 ratios as compared to other synovial cell types. Galectin-1 and galectin-3 mRNA expression decreased in the presence of the inflammatory cytokines interleukin-1[beta] (IL-1[beta]) and tumor necrosis factor [alpha] (TNF[alpha]). Galectins facilitated MSC adhesion, and mature focal adhesion complexes were abrogated in the presence of the pan-galectin inhibitor [beta]-lactose. Galectins promoted cell spreading and motility in 2D culture, resulting in cells with greater surface areas and more protrusions.
Lubricin; Galectins; Osteoarthritis
Mohammed,Hussni Omar; Bonassar,Lawrence; Sondermann,Holger
Ph. D., Veterinary Medicine
Doctor of Philosophy
dissertation or thesis