Research On The Expression Of Survivin In Synovial Fibroblasts Of Rheumatoid Athritis

Purpose:Search fast and effective methods for primary culture of RA Fibroblast-like synoviocytes(FLS) in vitro.We studied whether or not survivin has a potential as a protective agent against apoptosis of RA FLS in vitro.Methods:1.Synovial tissue was obtained during surgery from patients undergoing total knee replacement or arthroscope owing to RA or external injury.Primary synovial cells were cultured by means of four methods.We calculate main operational time for each method,account number of cells after seven days cultured with each method. Cultured cells were defined by light microscope and immunochemistry detections with Vimentin monoclonal antibody.2.Synovial cells in passages 3～8 were used for the experiment.20 RA FLS and 8 surgical trauma FLS were devided into 4 subgroups:methotrexate(MTX) -treated group,AS₂O₃-treated group,blank control group and surgical trauma FLS group.In each group after these treatment,the expression of Survivin and Caspase-3 mRNA were examined semiquantitatively by RT-PCR at 0h,24h and 48h respectively.Results:1.The sequence of four methods' effectivity was explant＞double enzyme＞collagenase＞trypsin method;in time consuming,trypsin was significantly faster than other methods;cell numbers was accounted at the 7th day of primary culture,there was significantly difference between each method(Chi-Square test,P＜0.05);immunohistochemistry demonstrated that there was no significantly difference between each methods(P＞0.1) and cells were most FLS(＞95%) In passage3,the positive cells of vimentin were more than 95%.2.Survivin and caspase-3 expressed in untreated RA FLS,but not in normal FLS.3.Treatment of RA FLS with MTX and AS₂O₃ resulted in an inhibition of cell proliferation,accompanied by down-regulation of survivin and concomitant up-regulation of Caspase-3.Conclusion:We propose that tissue block method is more effective than others while trypsin method is faster than others,survivin expressed in FLS of RA,and it may be a pathogenesis agent to protect the RA FLS against apoptosis,MTX and AS₂O₃ maybe control the diease by induce the apoptosis of RA FLS through down-regulation of survivin directly or indirectly.