Ex vivo pretreatment of donor organ with siRNA nanoparticles attenuates cold ischemia-reperfusion injury in cardiac transplantation

Heart transplantation is an optimal therapeutic regimen for terminal-stage cardiac failure. However, cold ischemia-reperfusion injury (CIRI) remains an unavoidable and outstanding challenge, which is a significant obstacle to early graft dysfunction and long-term survival. Blockage of complement, apoptosis, and inflammation by small interfering RNA is considered a strategy for attenuating CIRI and protecting cardiac function. However, their delivery to the donor organ is still a serious challenge due to the polyanionic nature and high molecular weight properties. Here, we have designed a novel functionalized gene delivery system of direct delivery and sustained release of siRNAs targeting complement C3 (C3), Caspase-3, and nuclear factor κB (NF-κB) to treat the donor organ prior to transplantation. The functionalized gene delivery system (siRNA-TNPs), composed of CaCO3/CaP/TAT embellished carboxymethyl chitosan (CaCO3/CaP/TCMC) and synthesized through the co-precipitation method, efficiently encapsulates siRNAs during self-assembly. The siRNA-TNPs safeguards siRNAs from biological degradation, facilitates intracellular siRNA transfection, promotes lysosomal escape, and enhances the delivery efficiency of siRNA to the donor hearts. Perfusion of donor hearts with siRNA-TNPs prior to transplantation attenuated C3, Caspase-3, and NF-κB genes expression of donor heart for at least 5 days after transplantation. Furthermore, silencing of C3, Caspase-3, and NF-κB genes expression alleviated cell apoptosis, myocardial damage, tissue inflammation, and rejection and improved cardiac function. These data suggest that the multiple-target siRNA-TNPs solution can extend the preservation time for donor grafts, attenuate IRI, and protect cardiac function in murine models of heart transplantation, which provides a principal of concept for potential clinical translation.