Temporal effects of cytokine treatment on lubricant synthesis and matrix metalloproteinase activity of fibroblast-like synoviocytes.

Fibroblast-like synoviocytes (FLS) are major contributors to the composition and function of synovial fluid (SF). In disease, changes to important SF molecules such as hyaluronic acid (HA), lubricin, and numerous inflammatory markers contribute to a loss of SF functional properties. Previous studies characterized the ability of FLS to produce SF molecules in short-term cultures using continuous cytokine supplementation. This study assessed the HA, lubricin, and matrix metalloproteinase-2 (MMP-2) secretion profile of FLS over 12 days of culture. FLS were subjected to continuous, intermittent, and sequential cytokine treatments of interleukin-1 beta (IL-1ß), tumour necrosis factor-alpha (TNF-α), and transforming growth factor-beta 1 (TGF-ß1). HA was assessed by an enzyme-linked immunosorbent assay (ELISA) for content and agarose gel electrophoresis for molecular weight distribution. Relative lubricin content was determined by western blot. Pro MMP-2 and active MMP-2 were quantified by gelatin zymography. All intermittent and sequential treatments significantly increased secretion of high-molecular-weight (>3 MDa) HA for the duration of the culture. Sequentially treated groups elevated lubricin synthesis, whereas only groups receiving IL-1ß and TNF-α for 2 days followed by TGF-ß1 for 1 day reduced active MMP-2 to unstimulated control levels. These data provide important information on the long-term functional potential of cytokine-stimulated FLS and suggest that temporal regulation of cytokine exposure can be a powerful tool to guide healthy synovial secretions.

Cartilage Metabolism is Modulated by Synovial Fluid Through Metalloproteinase Activity.

Synovial fluid (SF) contains various cytokines that regulate chondrocyte metabolism and is dynamically associated with joint disease. The objective of this study was to investigate the effects of diluted normal SF on catabolic metabolism of articular cartilage under inflammatory conditions. For this purpose, SF was isolated from healthy bovine joints, diluted, and added to cartilage explant cultures stimulated with interleukin-1 (IL-1) for 12 days. The kinetic release of sulfated glycosaminoglycan (sGAG) and collagen, as well as nitric oxide and gelatinase matrix metalloproteinases were analyzed in the supernatant. Chondrocyte survival and matrix integrity in the explants were evaluated with Live/Dead and histological staining. Diluted synovial fluid treatment suppressed sGAG and collagen release, downregulated the production of nitric oxide and matrix metalloproteinases, reduced IL-1-induced chondrocyte death, and rescued matrix depletion. Our results demonstrate that normal SF can counteract inflammation-driven cartilage catabolism. This study reports on the protective function of healthy SF and the therapeutic potential of recapitulation of SF for cartilage repair.

Zonal variation of MRI-measurable parameters classifies cartilage degradation.

Osteoarthritis (OA) is a degenerative joint disease resulting in the deterioration of articular cartilage, a tissue with minimal ability to self-repair. Early diagnosis of OA with non-invasive imaging techniques such as magnetic resonance imaging (MRI) could provide an opportunity to intervene and slow or reverse this degeneration process. This study examines the classification of degradation states using MRI measurements. Enzymatic degradation was used to specifically target proteoglycans alone, collagen alone and both cartilage components sequentially. The resulting degradation was evaluated using MRI imaging techniques (T1, T2, diffusion tensor imaging, and gadolinium enhanced T1) and derived measures of water, glycosaminoglycan and collagen content. We compared the classification ability of full thickness averages of these parameters with zonal averages (superficial, medial, and deep). Finally, we determined minimum variables sets to identify the smallest number of variables that allowed for complete separation of all degradation groups and ranked them by impact on the separation. Zonal analysis was much more sensitive than full thickness averages and allowed perfect separation of all four groups. Superficial zone cartilage was more sensitive to enzymatic degradation than the medial or deep zone, or the full thickness average. Variable ranking consistently identified collagen content and organization as the most impactful variables in the classification algorithm. The aim of this study is to classify cartilage degradation using only non-invasive MRI parameters that could be applied to OA diagnosis. Our results highlight the importance of zonal variation in the diagnosis of cartilage degeneration. Our novel, non-invasive collagen content measurement was crucial for complete separation of degraded groups from control cartilage. These findings have significant implications for clinical cartilage MRI for disease diagnosis.