Neddylation blockade protects endothelial cells against inflammation and ischemia-reperfusion injuries.

BACKGROUND

Long-term allograft survival is limited by the occurrence of chronic allograft dysfunction, mainly due to antibody mediated rejection and aspecific fibrosis. The latter is favored by ischemia-reperfusion injury (IRI), particularly in organs from extended criteria donors and from donors after cardiocirculatory arrest. The graft endothelium is highly sensitive to IRI that causes chronic damages and increases its immunogenicity. Thus, improvement of preservation methods and the treatment of organs prior to implantation (so called "graft preconditioning") constitute a major clinical challenge to improve graft survival. Among the mechanisms implicated in the reactivity of endothelial cells, neddylation has emerged as a potential candidate of interest.

METHODS

To study the potential of neddylation blockade as a preconditioning strategy, we used an in vitro model of microvascular endothelial cells (ECs) treated in conditions close to those of organ transplant preservation. ECs were subjected to hypothermic preservation (4 °C) in the presence of the neddylation inhibitor MLN4924, and following rewarming (37 °C), stimulated with TNAα and IFNγ to mimic sterile inflammation at reperfusion.

RESULTS

We found that preconditioning of ECs with MLN4924 had potent anti-inflammatory effects by blocking the induction of adhesion molecules (ICAM1, VCAM1, E-Selectin), of HLA class I and II, and of the pro-inflammatory cytokines and chemokines IL6, RANTES and Fractalkine. Functionally, this translated into inhibition of EC-mediated T lymphocyte activation and of T cell adhesion to ECs. MLN4924 preconditioning also blocked the induction of prothrombotic molecules (Tissue Factor and PAI-1), and platelet adhesion to ECs. Finally, MLN4924 enhanced the expression of the complement inhibitor CD55 and of the antioxidant molecule HO-1, and protected ECs from hypothermia and hypoxia induced cell death.

CONCLUSIONS

Our results identify the neddylation inhibitor MLN4924 as a powerful protective molecule for endothelial cells and a promising candidate for graft preconditioning and protection of organs against IRI.