Experimental Study On The Treatment Of Sj(?)gren's Syndrome Of Mouse With Adenovirus-mediated CTLA-4Ig

OBJECTIVE Sj?gren's syndrome is a system autoimmune disorder ,whose key feature is involving salivary gland,lacrimal gland etc,and cause gland secrete functional disturbance,appear Dryness of eye and Dry mouth.Its major mechanism involves T-cell mediated autoimmune response.Its pathologic characteristics are infiltration of lymphocyte of anacrine gland,submaxillary dut ectasia and destruction of gland alveolus.At present,because the exact pathogenesis of SS remains unclear,effective therapeutic measures are unavailable.In this study,the animal model of CTLA-4 was used to block the initial stage of immune response,i.e.,experimental autoimmune encephalomyelitis(SS) was established using BALB/c mouse.AdCTLA-4Ig was reconstructed by combining the gene of a fusion protein of CTLA-4 and adenoviral vectors.AdCTLA-4Ig was injected into animal body to observe its effects in blocking immune response and, thus,providing an experimental basis for the treatment of SS wihe CTLA-4Ig.METHODS CTLA4Ig gene in recombinant adenvius vector was Constructed, SS animal model was established.Adenovirus-mediated CTLA-4Ig was administered via intra-peritoneally.The therapeutic effect of AdCTLA-4Ig on SS was observed in contrast with that of thymic peptide therapy.The experiment was composed of three steps:1.construction and amplification of CTLA4Ig gene in recombinant adenvius vector: Recombinant adenovirus vector AQP1 was constructed by using a new simplified bacterial homologous recomblnant system.The pCTLA4Ig and shuttle plasmid pAdTrack-CMV were enzymetic digestion to generate recombinant shuttle plasmid.After cotransformation BJ5183 E.coli.bacteria by lineared recombinant shuttle plasmid and adenovirus backbone plasmid pAdEasy-1,recombinant adenovirus vector plasmid was generated by homologous recombination Follow transfection 293 cells by liPofectamine, recombinant adenovirus was generated and amplified. Tite was determined by expression of report gene GFP in infected 293 cell under fluorescence microscope.After repeated infection-collection-freeze/thaw,the lyses...