Selective inhibition of inducible nitric oxide synthase in experimental osteoarthritis is associated with reduction in tissue levels of catabolic factors

Abstract

Objective: We used the experimental dog model of osteoarthritis (OA) to examine the in vivo effects of N-iminoethyl-L-lysine (L-NIL), a selective inhibitor of the inducible nitric oxide synthase (iNOS), on the tissue level and distribution of interleukin-1beta (IL-1beta), collagenase-1, stromelysin-1, cyclooxygenase-2 (COX-2), iNOS and nitrotyrosine, through immunohistochemical and morphometric analysis.

Methods: Cartilage and synovial membrane specimens were obtained from 3 experimental groups of dogs: Group I--unoperated dogs that received no treatment; Group II--dogs subjected to a sectioning of the anterior cruciate ligament of the right knee and given no treatment; and Group III--operated dogs that received oral treatment with L-NIL (10 mg/kg twice daily/po) for 10 weeks starting immediately after surgery. The operated dogs were killed 10 weeks post-surgery. The tissue distributions of IL-1beta, metalloproteases (MMP), COX-2, iNOS and nitrotyrosine were documented by immunohistochemistry using specific antibodies, and quantified by morphometric analysis.

Results: In cartilage, the cell scores (percentage of chondrocytes staining positive for the antigen) for iNOS and 3-nitrotyrosine were dramatically enhanced in OA specimens compared to normal (p < 0.0001). However, the cartilage of dogs treated with L-NIL showed significantly lower cell scores for iNOS (p<0.0001, condyle; p<0.001, plateau), nitrotyrosine (p<0.0004; p<0.0001) and COX-2 (p<0.0001; p<0.001) compared to that of untreated OA dogs. Similar findings were observed for collagenase-1 and stromelysin-1, where the increased cell scores of these 2 MMP in OA cartilage were reduced after treatment with L-NIL (collagenase: p<0.002, condyle; p<0.0003, plateau; stromelysin: p<0.006; p<0.0001). The increased cell scores for the IL-1beta, COX-2, iNOS and nitrotyrosine found in the synovial lining and mononuclear cell infiltrate of operated animals were also found to be markedly reduced in dogs treated with L-NIL.

Conclusion: Our study demonstrates for the first time in vivo in an experimental model of OA, that a selective inhibition of iNOS by L-NIL and the subsequent decreased production of NO also results in a marked decrease in production of major catabolic factors such as MMP, IL-1beta and peroxynitrite, as well as a reduction in COX-2 expression.