Immunosuppressive role of fibrinogen-like protein 2 (FGL2) in CD8+regulatory T cells-mediated long-term graft survival
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KeywordsAdenovirus Recombinant Cardiac Allograft Transcriptomic Comparison Immunosuppressive Function Immunoregulatory Property
We have previously reported that, in a model of cardiac allograft in rat, blockade of CD40-CD40L interaction induces long-term graft survival mediated by CD8+CD45RClow regulatory T cells (Tregs) . Transcriptomic comparison of Tregs from AdCD40Ig-treated vs naïve rats highlighted the overexpression of FGL2 whose immunoregulatory properties are little known .
Material and methods
A lewis 1W rat heart is grafted in a MHC-mismatched Lewis 1A rat and infected with 2.1010pi of adenovirus recombinant for CD40Ig molecule (AdCD40Ig) the day of the graft. Tregs, effector CD4+CD25- T lymphocytes (TL), and plasmacytoïde dendritic cells (pDC) from spleen are sorted by FACS Aria for in vitro tests. For in vivo studies, 4,5.1011vg of FGL2-recombinant adenovirus associated virus (AAVFGL2) are intramuscularly or intravenously injected in recipients 30 days before the graft. Splenocytes are transferred to sublethaly irradiated rats by i.v injection the day before the graft.
We confirmed FGL2-overexpression in splenic Tregs and in the graft of AdCD40Ig-treated vs non-treated and naïve rats, at mRNA and protein level. FGL2 involvement in Tregs immunosuppressive function was proved by in vitro and in vivo experiments. Indeed, Tregs from AdCD40Ig-treated rats inhibit TL proliferation in response to allogeneic pDC. This inhibition is abrogated by FGL2-blocking antibodies  and can be mimicked by FGL2 protein alone. Moreover, AAV-mediated FGL2 overexpression in rat prolongs graft survival with a median of 18.5 days vs 11 days for controls by i.m injection and survival is improved when i.v injected. Furthermore, adoptive transfer of splenocytes from an AAVFGL2-treated tolerant rat, to irradiated rats, transmits long-term graft survival iteratively.
This is the first demonstration that the immunosuppressive molecule FGL2 is able to induce a long-term graft survival and that this tolerance is active and transferable by splenocytes. Work is under progress to identify the population responsible for this infectious tolerance.
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