title: Synergic silencing of costimulatory molecules prevents cardiac allograft rejection.
abstract: BACKGROUND
  While substantial progress has been made in blocking acute transplant rejection with the advent of immune suppressive drugs, chronic rejection, mediated primarily by recipient antigen presentation, remains a formidable problem in clinical transplantation. We hypothesized that blocking co-stimulatory pathways in the recipient by induction of RNA interference using small interference RNA (siRNA) expression vectors can prolong allogeneic heart graft survival.
  
  
  METHODS
  Vectors expressing siRNA specifically targeting CD40 and CD80 were prepared. Recipients (BALB/c mice) were treated with CD40 and/or CD80 siRNA expression vectors via hydrodynamic injection. Control groups were injected with a scrambled siRNA vector and sham treatment (PBS). After treatment, a fully MHC-mismatched (BALB/c to C57/BL6) heart transplantation was performed.
  
  
  RESULTS
  Allogeneic heart graft survival (>100 days) was approximately 70% in the mice treated simultaneously with CD40 and CD80 siRNA expression vectors with overall reduction in lymphocyte interstitium infiltration, vascular obstruction, and edema. Hearts transplanted into CD40 or CD80 siRNA vector-treated recipients had an increased graft survival time compared to negative control groups, but did not survive longer than 40 days. In contrast, allogenic hearts transplanted into recipients treated with scrambled siRNA vector and PBS stopped beating within 10-16 days. Real-time PCR (RT-PCR) and flow cytometric analysis showed an upregulation of FoxP3 expression in spleen lymphocytes and a concurrent downregulation of CD40 and CD80 expression in splenic dendritic cells of siRNA-treated mice. Functional suppressive activity of splenic dendritic cells (DCs) isolated from tolerant recipients was demonstrated in a mixed lymphocyte reaction (MLR). Furthermore, DCs isolated from CD40- and CD80-treated recipients promoted CD4+CD25+FoxP3+ regulatory T cell differentiation in vitro.
  
  
  CONCLUSIONS
  This study demonstrates that the simultaneous silencing of CD40 and CD80 genes has synergistic effects in preventing allograft rejection, and may therefore have therapeutic potential in clinical transplantation.
authors.forename: Xusheng
authors.initials: X
authors.lastname: Zhang
authors.forename: Yanling
authors.initials: Y
authors.lastname: Liu
authors.forename: Guangfeng
authors.initials: G
authors.lastname: Zhang
authors.forename: Jun
authors.initials: J
authors.lastname: Shi
authors.forename: Xiao
authors.initials: X
authors.lastname: Zhang
authors.forename: Xiufen
authors.initials: X
authors.lastname: Zheng
authors.forename: Alex T
authors.initials: AT
authors.lastname: Jiang
authors.forename: Zhu-Xu
authors.initials: ZX
authors.lastname: Zhang
authors.forename: Nathan
authors.initials: N
authors.lastname: Johnston
authors.forename: King Sun
authors.initials: KS
authors.lastname: Siu
authors.forename: Ruiqi
authors.initials: R
authors.lastname: Chen
authors.forename: Dameng
authors.initials: D
authors.lastname: Lian
authors.forename: David
authors.initials: D
authors.lastname: Koos
authors.forename: Douglas
authors.initials: D
authors.lastname: Quan
authors.affiliation: Department of Surgery, Pathology, and Ocology, University of Western Ontario, London, Canada. weiping.min@uwo.ca.
authors.forename: Wei-Ping
authors.initials: WP
authors.lastname: Min
mesh_terms: D000818:Animals; D018122:B7-1 Antigen / Q000235:genetics*; D001483:Base Sequence; D019013:CD40 Antigens / Q000235:genetics*; D017931:DNA Primers; D005434:Flow Cytometry; D020868:Gene Silencing; D006084:Graft Rejection / Q000235:genetics / Q000517:prevention & control*; D016027:Heart Transplantation; D007959:Lymphocyte Culture Test, Mixed; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D034741:RNA, Small Interfering / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D018122:B7-1 Antigen; D019013:CD40 Antigens; D017931:DNA Primers; D034741:RNA, Small Interfering