Klotho Suppresses Indoxyl Sulfate-Mediated Apoptosis in Human Kidney Proximal Tubular (HK-2) Cells through Modulating the AKT/Nrf2 Mechanism

Chronic kidney disease (CKD) is a progressive condition with substantial prevalence worldwide. The uremic toxin indoxyl sulfate (IS) is known to induce tubulotoxicity and adverse effects in various organs. It has been shown that the expression of the antiaging klotho protein is downregulated in the IS-stimulated proximal tubule cells and kidneys, but the detailed mechanism underlying the implication of reduced klotho in nephropathy remains largely unclear. In the present study, we demonstrated that the repressed klotho following IS stimulation contributed to the reduced cell viability and increased cytotoxicity of HK-2 cells (a proximal tubular cell line). We showed that recombinant klotho reversed the AKT/Nrf2 axis in the IS-treated HK-2 cells, leading to the restoration of the antioxidant HO-1, NQO1, and SOD as well as diminished ROS production. Most importantly, our results suggested that the IS-induced alteration of mitochondrial membrane potential, mitochondrial COX-III mRNA expression, and mitochondrial complex III activity was all mediated by the klotho/AKT/Nrf2 axis. By examining the expression of Bax, Bcl-2, and cytochrome <i>c</i> using Western blot and caspase-3-positive cells using flow cytometry, we demonstrated that klotho participated in the IS-induced apoptosis of HK-2 cells as well. Taken together, our data revealed that downregulation of klotho in the IS-stimulated HK-2 cells leads to decreased cell viability as a result of lower antioxidant capacity and ROS accumulation following mitochondrial respiratory chain dysfunction and subsequent apoptosis, possibly through inhibition of the AKT/Nrf2 axis.