Suppression of MHC expression on corneal cells to prevent immune rejection after allogeneic transplantation: Experimental investigation in a mouse model

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The high variability of the major histocompatibility complex (MHC), in humans histocompatibility leukocyte antigen (HLA) complex, and of the minor histocompatibility antigens (mHA) significantly contributes to elicit an immune response after allogeneic cornea transplantation, and may lead to the development of acute or chronic rejection. In addition, an often inevitable second cornea transplantation is always associated with an increased risk of graft rejection. This project aims to permanently reduce HLA class I and class II expression on cornea transplants to decrease the risk of graft rejection after keratoplasty. This method represents a new therapeutic concept and may contribute to prevent graft rejection. RNA-interference technology will be used to induce the downregulation of MHC expression. This method not only silences the expression of MHC molecules on the cell surface but also contributes to abrogate the presentation of minor histocompatibility antigens derived from other polymorphic molecules. Previously, we demonstrated that silencing HLA expression in a permanent manner can efficiently abrogate a deleterious immune response against allogeneic cells and significantly prolongs the survival of cells after allogeneic transplantation. In addition we have shown the feasibility of silencing HLA class I expression in human cornea endothelium in its original 3D-structure. In the proposed study, the transplantation and monitoring of rejection of manipulated and non-manipulated corneas will be performed in a mouse keratoplasty model for graft rejection (BALB/c and C3H). A lentiviral system will be used for the delivery of short hairpin RNA (shRNA) targeting mouse beta2-microglobulin or targeting the alpha chain of H2-E (H2-Ea). The presence of antigen presenting cells (APCs) and rejection markers will be assessed at defined time points. In addition, morphological changes after silencing MHC expression and cornea transplantation will be analysed by microscopy. Also, apoptosis and necrosis in the corneas will be evaluated. This study presents a new strategy to overcome the limitations and drawbacks caused by HLA mismatches in keratoplasty.