Abstract: SA-PO098
Role of AMPK and KLF4 in Determining the Survival of Cisplatin-Treated Human Umbilical Vein Endothelial Cells
Session Information
- AKI: Mechanisms - Primary Injury and Repair - II
November 09, 2019 | Location: Exhibit Hall, Walter E. Washington Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Acute Kidney Injury
- 103 AKI: Mechanisms
Authors
- Lieberthal, Wilfred, Stony Brook Medicine, Stony Brook, New York, United States
- Tang, Meiyi, Stony Brook Medicine, Stony Brook, New York, United States
- Liu, Qin, Stony Brook Medicine, Stony Brook, New York, United States
- Boriushkin, Evgenii, Stony Brook Medicine, Stony Brook, New York, United States
- Mallipattu, Sandeep K., Stony Brook Medicine, Stony Brook, New York, United States
Background
We have reported that preconditioning renal tubular cells with A-769662, a specific pharmacologic activator of AMPK, increased the survival of these cells when subjected to hypoxic stress. We have also shown that preconditioning mice A-769662 ameliorates the severity of ischemic AKI in mice in vivo. In these studies we examined the role of AMPK and the Kruppel-like transcription factor-4 (KLF4) in determining the fate of human umbilical epithelial cells ( HUVECs) exposed to cisplatin
Methods
The phosphorylation (activity) of AMPK was determined by immunoblotting and expressed as % of total AMPK. The expression of KLF4 was knocked down (by >85%) using a specific siRNA (“KD cells”). “Control cells” were transfected with a scrambled siRNA. The response of HUVECVs to cisplatin induced-injury was determined by first pretreating the cells with either A-769662 (250uM) or its vehicle for 24 h., followed by incubation with either cisplatin (100uM) or its vehicle) for an additional 18h. Then cell survival was determined by flow cytometry and expressed as a % of vehicle-treated cells.
Results
A-769662 increased the phosphorylation (activity) of AMPK to a comparable extent in control and KD cells (by 6.3±.24 and by 5.9±2.2 fold respectively). In control cells, the activation of AMPK induced by A-76962 (250mM), increased the expression of KLF4 mRNA (by 4±2.3 fold), and of KLF4 protein (by 3.9±2.2 fold). As expected, A-769662 had no effect on the expression of KLF4 in KD cells. In the absence of preconditioning with A-769662, the survival after cisplatin treatment was substantially higher in control cells (55.4%±4.2) than in KD cells (15.5±3.4%). These data show that KLF4 has pro-survival effects that are independent of AMPK. Furthermore, preconditioning HUVECs with A-769662, increased the survival of both control cells to 84.4±6.5%, and of KD cells to 57.9±7.6%. These data demonstrate, that the pro-survival effect of KLF4 is increased by the activation of AMPK.
Conclusion
i) AMPK induces the expression of KLF4; ii) AMPK and KLF4 both promote the survival of cisplatin-treated cells; iii) the pro-survival effects of AMPK and KLF4 are mediated by different pathways; iv) the activation of AMPK augments the pro-survival effect of KLF4 by increasing its expression.
Funding
- Private Foundation Support