Abstract
Hyperacute rejection in xenotransplantation is caused by activation of complement (C) on endothelium. We have previously shown that purified C-regulators of the pig (CD59 and membrane cofactor protein [MCP]) are efficient regulators of human C (HuC). The aim of this study was to clarify the role of endogenously expressed C-regulatory molecules on pig endothelium in the protection against hyperacute rejection. Porcine aortic endothelial cells (PAEC) were harvested and cultured for various passages. PAEC were examined for the expression of endogenous pig CD59 and MCP by flow cytometry. PAEC were assessed for their susceptibility to lysis by HuC. The effect of phorbol 12-myristate 13-acetate and various cytokines on the expression of MCP and CD59 and C-susceptibility was assessed. Primary PAEC showed an initial high level of expression of pig CD59, however, upon culturing, CD59 levels decreased dramatically to about 20% after five passages. In contrast, levels of MCP doubled upon culturing of PAEC to confluency and remained stable during at least five passages. Primary cells and cells in the early passages were more resistant to HuC than cells that were cultured for longer. Blocking the function of CD59 but not of MCP using monoclonal antibody increased the susceptibility to HuC. Purified human CD59 incorporated to a level of expression similar to that of pig CD59 reversed the increased C-susceptibility, suggesting that pig and human CD59 are similarly protective against HuC. Increase of C-resistance and of expression of pig MCP, but not of CD59, was achieved upon incubation with phorbol 12-myristate 13-acetate. Tumor necrosis factor-alpha, interleukin-1beta, interleukin-4, or interferon-gamma had no effect on C-regulator expression or C-susceptibility. These data demonstrate the importance of using primary PAEC or cells in the first passages of culturing in in vitro models of xenotransplantation and show that pig MCP and, in particular, pig CD59 play an important role in protection of PAEC from HuC.
Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
