An Organ Culture System To Study The Effect Of N-acetyl-L-cysteine (NAC) On The Degeneration Of Intervertebral Disc

[Objective]:To develop a model of disc degeneration using an organ culture,and observe the tissue morphology,protein expression and gene transcription so as to access the culture results.We also explore the effect of N-Acetyl-L-cysteine(NAC)on the degeneration of intervertebral disc(IVD).[Methods]:Lumbar intervertebral discs of 8-weeks-old ICR mice were taken out completely by asepsis.And they were randomly divided into three groups: Control group,degeneration group and NAC group.Interleukin-1beta(IL-1?)and tumor necrosis factor-alpha(TNF-?)were taken into the medium of degeneration group.In addition to the above two kinds of cytokines,we added NAC into the NAC groups.The discs were cultured in medium for different periods.We analyzed the tissue properties at 3,7and 10 days by histology and immunohistochemistry.Proteins and RNA were isolated from the intervertebral disc.Degeneration related protein and gene expression levels were evaluated by Western blot and RT-PCR.[Results]:Compared with the control group,sections stained with Alcian blue and hematoxylin-and-eosin counter staining showed confused structure and decreased content of proteoglycan.Collagen ?(Col-?),Collagen X(Col-X)and matrix metalloproteinases(MMP)3 positive area,the expression of Caspase-3,the transcription factor of nox A and Perp were obviously increased in degeneration group.While,the expression of Cyclin D1 and SOD-1 and the transcription factor of GSR and Gpx4 were significantly decreased in degeneration group.NAC group was similar to control group in the tissue morphology,protein expression and gene transcription.[Conclusion]:This study show a vitro model of IVD degeneration by adding IL-1?and TNF-? into the medium.In addition,drip of NAC into the medium could retardate the degeneration of IVD.However,the related mechanism needs further analysis.