Abstract: FR-PO1015

Role of Ubiquitin Proteasome System during Renal Cold Storage and Transplantation

Session Information

Category: Transplantation

  • 1701 Transplantation: Basic and Experimental

Authors

  • Parajuli, Nirmala, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States
  • Lo, Sorena B, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States
  • MacMillan-Crow, Lee Ann, University of Arkansas for Medical Sciences, Little Rock, Arkansas, United States
Background

We previously reported that renal cold storage (CS) leads to increased mitochondrial injury and renal damage following transplantation (Tx). However, CS induced molecular pathways responsible for worsening mitochondrial and renal damage following Tx are poorly understood. Alteration of Ubiquitin Proteasome System (UPS) has been reported in numerous diseases. The goal of this study was to evaluate if UPS was altered, and if this contributes to mitochondrial and renal damage after CS plus Tx (CS/Tx).

Methods

Rat kidneys exposed to CS (18hr) followed by Tx (CS/Tx) were used. Sham, autotransplant (ATx, a Tx without CS), and CS alone kidneys were used as control. The proteasome function in the renal extracts was measured using fluorogenic peptide substrates and spectrofluorometer. Mitochondrial function was assessed via high resolution respirometry.

Results

Proteasome function (chymotrypsin-like) was compromised after CS/Tx, but not in sham, ATx, or CS alone kidneys. A selective reduction of β5 (catalytic) subunit of the proteasome was observed only after CS/Tx. None of the groups showed change in expression of the predicted molecular weight of Rpt6 subunits of the proteasome. However, only CS/Tx kidneys showed an intense Rpt6 reactive bands of high and low molecular weights. Co-immunoprecipitation of renal extracts with Rpt6 antibody showed an association of Rpt6 subunit with heat shock proteins, which was significantly altered after CS/Tx. Similarly, compromised mitochondrial function was evident after CS, which was exacerbated after CS/Tx. Proteasome inhibition of NRK cells with Bortezomib showed reduced activity for mitochondrial complexes I, II and III. Similarly, antimycin A, a mitochondrial complex III inhibitor, treatment of NRK cells showed compromised proteasome function.

Conclusion

These data suggest, for the first time, that renal CS/Tx leads to altered expression/function of the UPS as well as its compromised association with heat shock proteins. Similarly, in vivo data suggest that the mitochondrial dysfunction precedes proteasome inactivation during CS/Tx. In vitro studies confirmed that a functional interaction exists between renal mitochondria and the UPS. New studies designed to preserve the UPS/mitochondrial function may have promising therapeutic implications for better outcomes after renal transplantation.

Funding

  • Other NIH Support