Obesity is one of risk factors of anterior cruciate ligament tear in man or cranial cruciate ligament (CCL) tear in dog. Adipokines are biologically active mediators released from adipocytes, and correlate with changes in body mass index. In order to study the possibility that adipocytes play a role in the pathogenesis of CCL disease, we investigated alterations of the matrix degradation biomarker genes (matrix metalloproteinase-13 [MMP-13], aggrecan) in CCL cells after stimulating with adipokines. We collected CCLs from 6 dog cadavers that had been euthanased for reasons other than musculoskeletal disease. CCL cells were isolated and treated with key adipokines including of adiponectin, leptin and visfatin at different concentration (0.1 ng/mL, 1 ng/mL and 10 ng/mL), and at three different time points (1 h, 6 h and 24 h). Real-time PCR was used to determine gene expression for MMP-13 and aggrecan in CCL cells comparing with negative control. In addition, lipopolysaccharide was used as a positive control. The statistical significance of differences between groups was determined using non-parametric Friedman test, followed by the Conover post-hoc test, and data were considered statistically significant at P<0.05.Introduction
Materials and Methods
The two-dimensional (2D) monolayer culture paradigm has limited translational potential to physiological systems; chondrocytes and tenocytes in monolayer lose expression of hallmarks of differentiated status (dedifferentiation). Qualitative assessment of three-dimensional (3D) cultures in musculoskeletal biology relative to native tissues has been limited. An understanding of prevailing gene regulatory networks is required to define whether 3D culture systems faithfully restitute the native tissue phenotype (redifferentiation). Using a systems biology approach to explore the gene networks associated with de- and re-differentiation may define targetable regulators associated with phenotypic plasticity of adult musculoskeletal cells. Global transcriptomic and proteomic profiling of matrix-depleted chondrocytes and tenocytes from the rat was performed for each of three conditions (native tissue, monolayer at passage three, or tissue-appropriate 3D cultures). Differential analysis of mRNA and protein abundance, gene ontology annotation, pathway topology impact analysis, and derivation of common mechanistic networks was undertaken to define consensus expression profiles, signalling pathways, and upstream regulators for de- and re-differentiation in each cell type.Introduction
Materials and Methods
Tendons and ligaments (TLs) play key roles in the musculoskeletal system. However, they are commonly damaged due to age-related wear and tear or torn in traumatic/sport related incidents resulting in pain and immobility. TLs contain cells and extracellular matrix (ECM) comprised of collagen, elastin, glycoproteins and proteoglycans. Although TLs are composed of similar components, their precise composition and arrangement of matrix macromolecules differ to provide specific mechanical properties and functions. To date little is known about how the main ECM proteins are arranged between the two tissue types. This data will provide essential information on fundamental structure of TLs leading to increased understanding of the function relationship between these tissue types. The aim of this study was to compare tendon-ligament differences in their ECM distribution of collagens, proteoglycans and elastic fibres. Anterior cruciate ligament (ACL) and long digital extensor tendon (LDET) were harvested from disease free cadaveric canine knee joints (n=3). Distributions of the main ECM components were assessed on longitudinal sections of ACL and LDET mid-substance. Antibody staining were assessed for collagen type I, III, VI, agreccan, versican, decorin, biglycan, elastin, fibrillin 1 and fibrillin 2.Introduction
Materials and Methods
