Abstract: FR-PO843
Proteomics Analysis Reveals Novel Molecular Mechanisms Associated with Normothermic Ex-Vivo Perfusion
Session Information
- Transplantation: Basic
October 26, 2018 | Location: Exhibit Hall, San Diego Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Transplantation
- 1801 Transplantation: Basic
Authors
- McEvoy, Caitriona M., University Health Network, Toronto, Toronto, Ontario, Canada
- Reid, Shelby, Institute of Medical Science, University of Toronto, Toronto, Ontario, Canada
- Urbanellis, Peter, University of Toronto, Toronto, Ontario, Canada
- Kaths, Moritz, University Hospital Essen, Essen, Germany
- Robinson, Lisa, The Hospital for Sick Children, Toronto, Ontario, Canada
- Selzner, Markus, Toronto General Hospital, Toronto, Ontario, Canada
- Konvalinka, Ana, University Health Network, University of Toronto, Toronto, Ontario, Canada
Background
Normothermic ex-vivo perfusion (NEVKP) is associated with significantly improved graft function following transplantation in comparison to static cold storage (SCS); however, the molecular mechanisms underpinning this improvement remain unclear. We hypothesized that NEVKP induces key alterations in the renal proteome compared to SCS, enabling us to characterize the molecular mechanisms that lead to superior graft function in this setting.
Methods
Porcine kidneys were removed following 30 minutes of warm ischemia, before being subjected to either SCS or NEVKP (n=5, each group) for 8 hours prior to autotransplantation. Kidney biopsies were collected at time zero, upon reperfusion, and at POD3. We conducted an unbiased proteomics analysis by LC-MS/MS on Q-Exactive-Plus mass spectrometer. Subsequent analyses were performed using MaxQuant, Perseus, pathDIP, mirDIP, NaVIGaTOR and Cytoscape.
Results
Renal function was significantly improved with NEVKP in comparison to SCS with decreased peak serum creatinine on POD1, and higher creatinine clearance on POD3. We detected 6354 proteins in total (FDR <0.01), with 71 proteins identified as significantly differentially expressed between experimental groups and time points (2-way ANOVA, p<0.05). Pathway enrichment analysis demonstrated that proteins increased in SCS compared to NEVKP kidneys mediated inflammation, antigen presentation, extracellular matrix production and fibrosis (e.g. THBS1, XRCC6, TMCO1, JUN, CD40). We identified microRNAs predicted to regulate these SCS-dominant proteins (e.g. miR-129, miR-206). In contrast, proteins increased in NEVKP compared to SCS were mostly metabolic, mediating energy production through TCA cycle (ACO2, COX4I1, ATP5J2, ETFB, MPC2, NDUFAF7). These proteins were predicted to be regulated by miR-203 and miR-199, and the transcription factor ZNF143.
Conclusion
NEVKP may thus repair grafts by diminishing inflammatory/fibrotic signals and altering metabolism via the TCA cycle. Conversely, proteins enriched in SCS kidneys are associated with fibrogenic and inflammatory pathways. Our findings may yield novel therapeutic targets to attenuate injury associated with warm ischemia.
Funding
- Private Foundation Support