Effects of CD25siRNA gene transfer on high-risk rat corneal graft rejection.

BACKGROUND: Corneal graft rejection is the major cause of corneal failures. Previous studies have shown that the CD25 monoclonal antibody can inhibit corneal graft rejection during the acute phase of rejection in rat models. In the current study, we evaluated the safety and efficacy of the topical Entranster™ vector in rat corneal applications and further investigated the effects of CD25siRNA gene transfer on high-risk rat corneal graft rejection.

METHODS: Fluorescence detection, clinical assessment, hematoxylin and eosin (HE), terminal deoxynucleotidyl transferase biotin-dUTP nick end labelling (TUNEL) and CD11b assays were used to evaluate the safety and efficacy of CD25siRNA gene transfer in normal SD rat corneas. Orthotopic corneal transplants were performed in alkali burned SD rats. Corneal recipients were divided into four groups that received different treatments. Clinical assessment, western blotting, quantitative real-time polymerase chain reaction (qRT-PCR), immunohistochemistry, HE and transmission electron microscopy (TEM) were performed on all grafts.

RESULTS: Low toxicity, no immunogenicity and high transfection efficiency were observed in rat corneas treated with the Entranster™ vector. Reduced endothelial cell apoptosis, inflammatory cell infiltration and graft neovascularisation were observed in the CD25siRNA treatment groups. The graft survival curves showed that CD25siRNA treatment significantly prolonged graft survival time, with better graft transparency and less graft oedema. Lower CD25 and higher IL-10 expression were detected in the CD25siRNA treatment groups during the study period, and a higher FOXP3 level was found in the CD25siRNA group than in the CD25siRNA-twice group on days 14 and 21 post-operation.

CONCLUSIONS: The Entranster™ vector is an effective vector for corneal gene therapy. CD25 siRNA gene transfer inhibits corneal graft rejection via upregulation of anti-inflammatory molecule expression.