L. IERINO1, D. CHRISTIANSEN2, E. MOUHTOURIS2, M. SANDRIN2
1Department of Nephrology and University of Melbourne, St Vincent’s Health, Fitzroy, Victoria; 2Department of Surgery and University of Melbourne, Austin Health, Heidelberg, Victoria.
Background: Our research has focused on the therapeutic strategy of locally secreted immunomodulatory molecules by cellular xenografts and previously demonstrated that survival of the porcine endothelial cell line (PIEC) engineered to secret soluble ICOS-Ig is specifically prolonged compared to wild-type PIEC. Xenograft prolongation was associated with CD4+CD25+Foxp3+ graft infiltrating T cells consistent with regulatory T cells. The effect of locally secreted ICOS-Ig on allograft survival was also examined.
Aim: (a) Study the mechanism of xenograft prolongation in more detail and (b) Determine whether allografts are prolonged using locally secreted ICOS-Ig.
Method and Results: CD4+25+ T cells from ICOS-Ig secreting xenografts were purified and the phenotype shown to express a range of markers consistent with regulatory T cells including Foxp3, Helios, CD127low, CD275 (ICOSL), CD278 (ICOS), CD39 and other regulatory T cell markers. Gene analysis of CD4+25+ T cells demonstrated increased Foxp3, IL-10, ICOS, granzyme B (GzmB) and perforin (pfp). The functional role of IL-10, pfp, GzmB in mediating xenograft prolongation was confirmed by transplanting ICOS-Ig PIEC and wild type PIEC into IL-10-/-, pfp-/- and GzmB-/- mice which demonstrated that ICOS-Ig PIEC and wild type PIEC were rejected in a similar time. The outcome of allografts was examined using locally expressed ICOS-Ig. A stable clone of NS-1 cell line (BALB/c origin) secreting ICOS-Ig was generated and effect on allograft survival examined by grafting onto C57BL/6 mice. Graft survival was significantly prolonged from 8 days to 15 days (p=0.008).
Conclusion: Prolonged survival of xenografts expressing ICOS-Ig was mediated by CD4+CD25+Foxp3+ Regulatory T cells. Allograft survival was also prolonged using locally secreted ICOS-Ig. The data indicates this therapeutic strategy may have broad clinical applications in transplantation.