Abstract: TH-PO119
Necroptosis and Ferroptosis Inhibitors Have Significant Benefit in Acute Ischemic Kidney Injury In-Vivo and In-Vitro
Session Information
- AKI: Inflammation, New Technologies, Omics
October 25, 2018 | Location: Exhibit Hall, San Diego Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Acute Kidney Injury
- 103 AKI: Mechanisms
Authors
- Gallagher, Kevin Michael, University of Edinburgh, Edinburgh, United Kingdom
- Beal, Allison M., GlaxoSmithKline, Collegeville, Pennsylvania, United States
- Tam, Yuen bun, University of Edinburgh, Edinburgh, United Kingdom
- Ross, James A., University of Edinburgh, Edinburgh, United Kingdom
- Marson, Lorna, The Queens Medical Research Institute, Edinburgh, United Kingdom
- Bertin, John, GlaxoSmithKline, Collegeville, Pennsylvania, United States
- Wigmore, Stephen J., University of Edinburgh, Edinburgh, United Kingdom
- Hughes, Jeremy, The Queens Medical Research Institute, Edinburgh, United Kingdom
- Harrison, Ewen M., University of Edinburgh, Edinburgh, United Kingdom
Background
We aimed to determine if necroptosis or ferroptosis inhibitors are beneficial in murine ischemia reperfusion injury, and if necroptosis or ferroptosis occur in human tubular cells.
Methods
Mice were subjected to 18m of bilateral renal ischemia with 24 or 48h reperfusion. Highly specific RIPK1 inhibitor, GSK547a, or ferroptosis inhibitor Liproxstatin-1 were given from 4h after injury. Outputs included creatinine, acute tubular necrosis (ATN) score, phosphorylated MLKL (pMLKL, end effector of necroptosis) stain, TUNEL stain and qPCR. Human renal tubular cells (HK-2) underwent 3 models of ischemia in-vitro +/- necroptosis/ferroptosis inhibitors.
Results
With drug given 4h after injury, both RIPK1 inhibitor and liproxstatin-1 significantly reduced serum creatinine (mmol/L)(Vehicle:239.8 (+/- 51.8), RIPK1 inhibitor:103.9 (+/- 48.2) p<0.001, Liproxstatin-1:43.1 (+/-14.4) p<0.001) and ATN score (Vehicle: 3/4 (3-4), RIPK1 inhibitor: 2/4 (1-2) p=0.008, Liproxstatin 1: 2/4 (1-2) p<0.001)(9/group) 48h after injury. 24h after injury there was extensive pMLKL in vehicle treated injured kidneys. pMLKL, ATN score and TUNEL stain were significantly reduced by RIPK1 inhibition(N=6). There was median 5-fold increase in renal RIPK3 expression 48h after injury compared to sham animals, this was significantly reduced by RIPK1 inhibition (p=0.004) but not by liproxstatin-1 (p=0.67) (N=9). A20 (necroptosis inhibitor) expression was significantly higher vs vehicle in RIPK1 inhibitor (p=0.02) and liproxstatin-1 (p=0.01) treated kidneys. Despite less ATN, Liproxstatin-1, but not RIPK1 inhibitor treated kidneys demonstrated higher expression of MCP-1 (p=0.002) and TNF-α (p<0.001) compared to vehicle. In in-vitro ischemic HK2 cell injury, there was evidence of both necroptosis (MLKL phosphorylation) and ferroptosis (iron dependent lipid peroxidation) with associated benefit for cell survival from necroptosis and ferroptosis inhibitors (N=3-6), although to different extents depending on the type and severity of injury.
Conclusion
Both necroptosis and ferroptosis contribute to ischemic kidney injury. There may be differential effects of related inhibitors on inflammatory genes. Further study is required to determine if this impacts long term renal function after an ischemic injury.
Funding
- Other NIH Support –