title: [Construction of an adeno-associated virus vector expressing CTLA-4Ig and its expression in the transplanted liver allografts].
abstract: OBJECTIVE
  To construct a recombinant adeno-associated virus vector pSNAV expressing CTLA-4Ig and to demonstrate its expression in transplanted liver allografts and to see if a long term inhibitive effect of CTLA-4Ig could be obtained though its use.
  
  
  METHODS
  After AAVCTLA-4Ig and PUC18 were cut with BamHI, CTLA-4Ig cDNA was inserted into the plasmid PUC18 by T4DNA ligase and PUC18-CTLA-4Ig was constructed. The obtained PUC18-CTLA-4Ig and pSNAV cut with Kpn I and EcoR I, CTLA-4Ig cDNA was inserted into plasmid pSNAV to construct the recombinant vector pSNAV-CTLA-4Ig, which was transfected into BHK-21 packaging cells by lipofectine-mediated transfection. Then the BHK-21 cell line was infected with HSV1-rc to produce a large amount of pSNAV- CTLA-4Ig. The specificity of the expressed product was identified by digestion with BamHI, PCR and sequence determination. The titer of the virus was detected. The product was infused into rats liver allografts via portal vein and its expression in the transplanted livers was detected immunohistochemically.
  
  
  RESULTS
  Recombinant adeno-associated virus vector pSNAV-CTLA-4Ig was generated and purified into 8.5 x 10(11)/ml. Agarose gel analysis of PCR products verified the presence of CTLA-4Ig. Digestion with BamHI and sequence determination confirmed that pSNAV-CTLA-4Ig was constructed. Expression of CTLA-4Ig in the transplanted livers was detected successfully.
  
  
  CONCLUSIONS
  Prepared pSNAV-CTLA-4Ig was constructed correctly and can express CTLA-4Ig effectively. Besides this, it can express CTLA-4Ig in rat liver allografts. It may be used in the study of transplant tolerance.
authors.affiliation: Center of Liver Transplantation, The First Affiliated Hospital of Nanjing Medical University, Nanjing 210029, China.
authors.forename: Shen
authors.initials: S
authors.lastname: Lu
authors.forename: Xue-hao
authors.initials: XH
authors.lastname: Wang
authors.forename: Guo-qiang
authors.initials: GQ
authors.lastname: Li
authors.forename: Yun
authors.initials: Y
authors.lastname: Gao
authors.forename: Yue
authors.initials: Y
authors.lastname: Yu
mesh_terms: D000069594:Abatacept; D000818:Animals; D000229:Dependovirus / Q000235:genetics* / Q000378:metabolism; D005822:Genetic Vectors; D018796:Immunoconjugates / Q000235:genetics / Q000378:metabolism*; D016031:Liver Transplantation; D011897:Random Allocation; D051381:Rats; D017207:Rats, Sprague-Dawley; D017208:Rats, Wistar; D014184:Transplantation, Homologous
chemical_list: D018796:Immunoconjugates; D000069594:Abatacept