Dr Marina Katerelos1, Dr Aspasia Pefanis1,2, Mr Kurt Gleich1, Dr Chathri Ratnayake1,2, Dr Mardiana Lee1,2, Dr Geoff Harley1, Prof David Power1,2, Prof. Peter Mount1,2
1Department of Nephrology, Austin Health, Heidelberg, Australia, 2Department of Medicine (Austin), Heidelberg, Australia
Biography:
Dr Peter Mount is Director of Nephrology at Austin Health and a Professor of Renal Medicine (Honorary) with the University of Melbourne. Prof Mount’s research is funded by NHMRC and is focused on the role of disturbance in renal energy metabolism in acute and chronic kidney disease, including the role of the energy-sensor AMP-activated protein kinase. Prof Mount also has longstanding involvements in clinical research, including his contribution to the BEST-FLUIDS trial. Prof Mount is committed to improving the lives of people with kidney disease, and was previously chair of the ANZSN Policy and Quality Committee.
Aim:
To investigate the role of regulated cell death pathways on the impact of obesity on acute kidney injury (AKI) in folic acid nephropathy (FAN).
Background:
Obesity dysregulates renal energy metabolism and increases AKI severity with FAN. The mechanisms are poorly understood, and the role of cell death pathways such as necroptosis, ferroptosis and apoptosis, remains unknown.
Methods:
Obesity was induced in male mice by high fat diet (HFD), versus control diet (CD), for 6-weeks. AKI was induced by FAN (AKI-FAN), through intraperitoneal folic acid administration (240 μg/g), versus vehicle in controls, with endpoint samples at 48-hrs. mRNA was measured by RT-PCR, and protein by western blot (WB) and immunohistochemistry.
Results:
FAN-AKI increased mRNA of necroptosis molecules including receptor-interacting kinase 1, receptor-interacting kinase 3 (RIP3), and mixed lineage kinase domain-like (MLKL) (P <0.0001). Obesity increased RIP3 mRNA expression with FAN-AKI [HFD-FAN 13.3(3.6)-fold vs CD-FAN 9.0(2.6)-fold, P=0.0094], which was confirmed by WB [HFD-FAN 18.8(2.6)-fold vs CD-FAN 14.8(3.4)-fold, P=0.0057]. Increased MLKL phosphorylation, a key step in activating necroptosis, was increased by obesity in FAN-AKI, measured by immunohistochemistry for %phospho-MLKL positive tubules [HFD-FAN 31.9(5.9) vs CD-FAN 21.0(4.3), P=0.0038]. FAN-AKI caused ferroptosis, with reduced mRNA for ACSL4, and increased for GPX4, however these changes were not different with obesity. Regarding apoptosis, BAX and BCL2 mRNA changes were not different between FAN-AKI groups with or without obesity, and cleaved caspase-3 protein expression (WB) was absent in both HFD and CD conditions.
Conclusion:
Obesity increased renal tubular necroptosis but not apoptosis or ferroptosis in this model of nephrotoxic AKI. These results suggest that necroptosis rather than apoptosis, is the dominant mechanism of regulated cell death increased by obesity in nephrotoxic AKI.
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