Cold Storage Followed by Transplantation Induces Immunoproteasome in Rat Kidney Allografts: Inhibition of Immunoproteasome Does Not Improve Function

Abstract

Key Points Cold storage (CS) increases the severity of graft dysfunction in a time-dependent manner, and prolonged CS decreases animal survival.CS plus transplant increases iproeasome levels/assembly in renal allografts; IFN-γ is a potential inducer of the iproteasome.Inhibiting iproteasome ex vivo during renal CS did not confer graft protection after transplantation. Background It is a major clinical challenge to ensure the long-term function of transplanted kidneys. Specifically, the injury associated with cold storage (CS) of kidneys compromises the long-term function of the grafts after transplantation. Therefore, the molecular mechanisms underlying CS-related kidney injury are attractive therapeutic targets to prevent injury and improve long-term graft function. Previously, we found that constitutive proteasome function was compromised in rat kidneys after CS followed by transplantation. Here, we evaluated the role of the immunoproteasome (iproteasome), a proteasome variant, during CS followed by transplantation. Methods Established in vivo rat kidney transplant model with or without CS containing vehicle or iproteasome inhibitor (ONX 0914) was used in this study. The iproteasome function was performed using rat kidney homogenates and fluorescent-based peptide substrate specific to β5i subunit. Western blotting and quantitative RT-PCR were used to assess the subunit expression/level of the iproteasome (β5i) subunit. Results We demonstrated a decrease in the abundance of the β5i subunit of the iproteasome in kidneys during CS, but β5i levels increased in kidneys after CS and transplant. Despite the increase in β5i levels and its peptidase activity within kidneys, inhibiting β5i during CS did not improve graft function after transplantation. Summary These results suggest that the pharmacologic inhibition of immunoproteasome function during CS does not improve graft function or outcome. In light of these findings, future studies targeting immunoproteasomes during both CS and transplantation may define the role of immunoproteasomes on short-term and long-term kidney transplant outcomes.