noc-18 and Osteoarthritis

Osteoarthritis (OA) is associated with increased levels of reactive oxygen species. This study investigated if increased oxidative DNA damage accumulates in OA articular cartilage compared with non-OA articular cartilage from pigs with spontaneous OA. Additionally, the ability of nitric oxide (NO) or peroxynitrite (ONOO(-)) induced DNA damage in non-OA chondrocytes to undergo endogenous repair was investigated. Porcine femoral condyles were graded for the stage of OA, macroscopically by the Collins Scale, and histologically by the modified Mankin Grade. Levels of DNA damage were determined in non-OA and OA cartilage, using the comet assay. For calibration, DNA damage was measured by exposing non-OA chondrocytes to 0-12 Gray (Gy) of X-ray irradiation. Non-OA articular chondrocytes were treated with 0-500 microM of NO donors (NOC-18 or SIN-1), and DNA damage assessed after treatment and 5 days recovery. A significant increase (P < 0.01) in oxidative DNA damage occurred in OA chondrocytes in joints with Mankin Grades 3 or greater, compared to non-OA chondrocytes. The percentage of nuclei containing DNA damage increased significantly (P < 0.001) from early to late grades of OA. An increase of approximately 0.65-1.7 breaks/1,000 kb of DNA occurred in OA, compared to non-OA nuclei. NOC-18 or SIN-1 caused significant DNA damage (P < 0.001) in non-OA chondrocytes that did not undergo full endogenous repair after 5 days (P < 0.05). Our data suggest significant levels of oxidative DNA damage occur in OA chondrocytes that accumulates with OA progression. Additionally, DNA damage induced by NO and ONOO(-) in non-OA chondrocytes does not undergo full endogenous repair.

Topics: Acetonitriles; Animals; Calibration; Cartilage, Articular; Cell Nucleus; Chondrocytes; DNA; DNA Damage; DNA Repair; Morpholines; Nitric Oxide; Nitroso Compounds; Osteoarthritis; Oxidative Stress; Peroxynitrous Acid; Swine; X-Rays

Osteoarthritis (OA) is associated with increased levels of reactive nitrogen and oxygen species and pro-inflammatory cytokines, such as interleukin-1 (IL-1). Nitric oxide (NO) can mediate a number of the catabolic effects of IL-1 in articular cartilage. The aims of this study were to determine if OA cartilage shows evidence of DNA damage, and if IL-1 could induce DNA damage in non-OA cartilage by increasing NO or superoxide.. Articular chondrocytes were isolated from porcine femoral condyles and embedded in 1.2% alginate. The effects of 24h incubation with IL-1, the nitric oxide synthase 2 (NOS2)-selective inhibitor, the free radical scavenger superoxide dismutase (SOD), the NO donor NOC18, or the combined NO and peroxynitrite donor SIN-1 on DNA damage were tested, using the "comet" assay. NO production was measured using the Griess assay. The type of oxidative damage present was assessed using a modified comet assay.. OA cartilage had significantly more DNA damage than non-OA cartilage (P<0.001). IL-1 caused an increase in DNA damage (P<0.01), which was associated with increased NO production (P<0.01). Both oxidative DNA strand breaks and base modifications of purines and pyrimidines were observed. IL-1-induced DNA damage was inhibited by an NOS2 inhibitor or by SOD (P<0.01). Furthermore, NOC18 or SIN-1 caused DNA damage (P<0.001).. Our work shows chondrocytes in osteoarthritic cartilage exhibit DNA damage, and that IL-1 induces DNA damage and reactive oxygen and nitrogen species in non-OA chondrocytes in alginate.

Topics: Animals; Cartilage, Articular; Cells, Cultured; Chondrocytes; DNA Damage; Female; Interleukin-1alpha; Molsidomine; Nitric Oxide Donors; Nitroso Compounds; Osteoarthritis; Reactive Nitrogen Species; Reactive Oxygen Species; Sus scrofa