Protein Kinase C-δ Mediates Kidney Tubular Injury in Cold Storage–Associated Kidney Transplantation

Significance Statement Use of cold storage for organ preservation in kidney transplantation is associated with cold ischemia-reperfusion injury that contributes to delayed graft function and affects the long-term outcome of transplanted kidneys. Using rat proximal tubule cells and a mouse model, the authors demonstrated that protein kinase C- δ (PKC δ ), which is implicated in ischemia-reperfusion injury in other organs, is activated in tubular cells during cold storage–associated transplantation and accumulates in mitochondria. There, it mediates phosphorylation of a mitochondrial fission protein, dynamin-related protein 1 (Drp1), at serine 616. Drp1 activation leads to mitochondrial fragmentation, accompanied by mitochondrial damage and kidney tubular cell death. Genetic ablation (in PKC δ -knockout mice) or use of a peptide inhibitor of PKC δ reduced kidney injury in cold storage–associated transplantation, pointing to PKC δ as a promising therapeutic target for kidney transplant. Background Kidney injury associated with cold storage is a determinant of delayed graft function and the long-term outcome of transplanted kidneys, but the underlying mechanism remains elusive. We previously reported a role of protein kinase C- δ (PKC δ ) in renal tubular injury during cisplatin nephrotoxicity and albumin-associated kidney injury, but whether PKC δ is involved in ischemic or transplantation-associated kidney injury is unknown. Methods To investigate PKC δ ’s potential role in injury during cold storage–associated transplantation, we incubated rat kidney proximal tubule cells in University of Wisconsin (UW) solution at 4°C for cold storage, returning them to normal culture medium at 37°C for rewarming. We also stored kidneys from donor mice in cold UW solution for various durations, followed by transplantation into syngeneic recipient mice. Results We observed PKC δ activation in both in vitro and in vivo models of cold-storage rewarming or transplantation. In the mouse model, PKC δ was activated and accumulated in mitochondria, where it mediated phosphorylation of a mitochondrial fission protein, dynamin-related protein 1 (Drp1), at serine 616. Drp1 activation resulted in mitochondrial fission or fragmentation, accompanied by mitochondrial damage and tubular cell death. Deficiency of PKC δ in donor kidney ameliorated Drp1 phosphorylation, mitochondrial damage, tubular cell death, and kidney injury during cold storage–associated transplantation. PKC δ deficiency also improved the repair and function of the renal graft as a life-supporting kidney. An inhibitor of PKC δ , δ V1-1, protected kidneys against cold storage–associated transplantation injury. Conclusions These results indicate that PKC δ is a key mediator of mitochondrial damage and renal tubular injury in cold storage–associated transplantation and may be an effective therapeutic target for improving renal transplant outcomes.