Background: AMPK activation by metformin protects against cisplatin mediated injury to kidney cells, and this effect of metformin is suppressed by blocking AMPK signaling to acetyl-CoA carboxylase (ACC). ATX-304 is a direct AMPK activator that improves glucose homeostasis and microvascular perfusion in type 2 diabetes.
Methods: Primary cultures of kidney tubular epithelial cells (TECs) were pre-treated with ATX-304 before exposure to cisplatin (20 µM, 23-hrs). Cellular injury was studied in TECs from wild type (WT) C57Bl/6 control mice and ACC1/2 double knock-in (ACC1/2DKI) mice.
Results: In WT TECs, ATX-304 activated AMPK, with 20 µM for 4-hrs increasing AMPK-αThr172 phosphorylation 1.8-fold (P<0.05) and phosphorylation of ACC-Ser79 4.5-fold (P<0.01). By RT-PCR, ATX-304 (20 µM, 4-hrs) increased expression of peroxisome proliferator-activated receptor-γ coactivator (3.5+0.2-fold increase, P<0.0001) and sirtuin-3 (3.4+0.2-fold increase, P<0.0001). Pre-treatment with ATX-304 protected WT TECs from cisplatin mediated cell death, measured by lactate dehydrogenase release assay (control 27.1+4.2% lysis vs ATX304 -0.6+2.5% lysis, p<0.0001), maintained cell viability (MTS assay), and reduced expression of the apoptosis marker cleaved caspase 3. In ACC1/2DKI TECs, greater reduction is cell viability (MTS assay) was observed (FC 0.57+0.03 vs 0.63+0.04 p=0.038). However, ATX-304 remained protective against cisplatin's effect on cell viability in the ACC1/2DKI TECs.
Conclusion: ATX-304 is protective against cisplatin mediated injury in TECs. Whilst blocking AMPK signaling to ACC increases the effect of cisplatin on cell viability, ATX-304 remained protective against injury in ACC1/2DKI TECs, indicating other AMPK actions contribute to this protective effect.