Effects Of Nitric Oxide Synthase Inhibitor On Repair Tissue In Articular Cartilage

BACKGROUNDArticular cartilage damage and degradation are the most common diseases in clinic, but the self-repaired ability of articular cartilage is limited. How could restore cartilage defects is still the problem of orthopedic doctors. No matter what applying to autograft and allograft, or chondrocytes graft and tissue engineer culture, the defect could be filled with para-hyaline cartilage or hyaline cartilage in the early stage that subsequently could be fibrous degeneration ineluctability. It is important to promote repair tissues quality and retard its degeneration; because these repair tissues have different structure, biochemical and biomechanics characteristic. A serial of studies showed NOS inhibitors were benefit for arthritis. At present, Studies emphasize to the skill of cartilage defect repair, so that it is inadequate to promote the quality of repair tissues and retard its degeneration. Now that nitric oxide could increase cartilage matrix synthesis, NOS inhibitors were applied for cartilage repair would mediator repair tissue degradation and ameliorate its quality.OBJECTIVE( 1 ) To explore the effects of NOS inhibitors for chondrocytes and cartilage entities, and offer bases of animal model study. (2) To discuss the effects of NOS inhibitors for the cartilage repair tissue. (3) To study theclinical potential effects of NOS inhibitors in OA.METHODS( 1) Articular entities and chondrocytes taken from mature New Zealand white rabbits, the toxicity of IL-lp and LPS and NOS inhibitor on chondrocytes were measured through MTT. Then the experiment were performed in five groups: control group; IL-lp+LPS group; IL-lp+LPS + SMT group; IL-lp+LPS + L-NIL group; IL-lp + LPS + L-NMA group. The release of NO and the activity of NOS in rabbit chondrocytes and cartilage entities were measured by Griess reaction and spectrophotometric methods. The proteoglycan synthesis was assessed by incorporation of radiolabelled sodium sulphate Na235S04. iNOS and MMP9mRNA expression were measured by in situ hybridization and RT-PCR.(2) Twenty-one New Zealand white rabbits were divided into normal group, the control and SMT group. The normal group did not use any drug interfere, the control and SMT groups were given normal saline and SMT respectively by a single hypodermic injection after IL-lp and LPS were administered by interarticular injection. The NO release and the activity of NOS were measured by Griess reaction and spectrophotometric methods in 8h. The proteoglycan synthesis and iNOSmRNA expression was assessed by incorporation of radiolabelled sodium sulphate Na235SO4 and in situ hybridization respectively after 48h. Through polyacrylamide and mouse muscle bag model, the bioactivity of rhBMP-2 protein was identified.(3) Full-thickness defects of cartilage were created in the trochlearn groove of sixty-six adult New Zealand White rabbits. Forty-four defects were empty, Forty-four were filled with a fibrin glue impregnated with rhBMP, and Forty-four were filled with a fibrin glue impregnated with rhBMP and hypodermic injection with Smg-kg'Mltf1 SMT. The animals were killed at eight, sixteen weeks and one-year postoperatively, and the gross appearance of the healed defect was assessed. The repair tissue was examined histologically and was evaluated according to a grading scale. The tissuesections were immunostained with antibodies against type-II collagen and type- I collagen, RT-PCR examines the expreesion of iNOSniRNA, MMPsmRNA and type- II collagen.(4) 17 specimens of articular cartilage were taken from osteoarthrosis patients. The experiments were performed in three groups: control group and L-NIL.and SMT intervenient group. The release of NO and the activity of NOS on OA cartilage were measured by Griess reaction and spectrophotometric methods after 72h culture. iNOSmRNA and MMPg expression was measured by in situ hybridization. Proteoglycan and hydroxyproline in OA cartilage was measured after lOd culture.RESULTS( 1 ) chondrocytes incubated in vitro for 72h in presen...