Nrf2-Heme Oxygenase-1 Attenuates High-Glucose-Induced Epithelial-to-Mesenchymal Transition of Renal Tubule Cells by Inhibiting ROS-Mediated PI3K/Akt/GSK-3β Signaling

Abstract

Background. Epithelial-to-mesenchymal transition (EMT) is thought to play a significant role in the advancement to chronic kidney disease and contributes to the deposition of extracellular matrix proteins and renal fibrosis relating to diabetic nephropathy. Method. We studied the effect of Nrf2-HO-1 signaling on high-glucose- (HG-) induced EMT in normal human tubular epithelial cells, that is, HK2 cells. In short, we treated HK2 cells with HG and sulforaphane (SFN) as an Nrf2 activator. EMT was evaluated by the expression activity of the epithelial marker E-cadherin and mesenchymal markers such as vimentin and fibronectin. Results. Exposure of HK2 cells to HG (60 mM) activated the expression of vimentin and fibronectin but decreased E-cadherin. Treatment of HK2 cells with SFN caused HG-induced attenuation in EMT markers with activated Nrf2-HO-1. We found that SFN decreased HG-induced production of reactive oxygen species (ROS), phosphorylation of PI3K/Akt at serine 473, and inhibitory phosphorylation of serine/threonine kinase glycogen synthase kinase-3β (GSK-3β) at serine 9. Subsequently, these signaling led to the downregulation of the Snail-1 transcriptional factor and the recovery of E-cadherin. Conclusion. The present study suggests that Nrf2-HO-1 signaling has an inhibitory role in the regulation of EMT through the modulation of ROS-mediated PI3K/Akt/GSK-3β activity, highlighting Nrf2-HO-1 and GSK-3β as potential therapeutic targets in diabetic nephropathy.

Pharmacotherapy against Oxidative Stress in Chronic Kidney Disease: Promising Small Molecule Natural Products Targeting Nrf2-HO-1 Signaling

“…HG increased ROS-induced PI3K/Akt/GSK3β activity and accelerated EMT in HK-2 cells. Further, the accumulation of EMT was reduced by treatment with an Nrf2 activator, sulforaphane, emphasizing the therapeutic potential of targeting Nrf2-HO-1 signaling [65]. The TGFβ1-increased ROS levels, EMT genes, FN, and collagen 1, were further increased by Nrf2 knockdown and suppressed by Keap1 knockdown in HK-2 cells [68], suggesting a critical role of Nrf2 in kidney fibrosis.…”

“…Subsequently, these ROS reduced cytochrome C oxidase activity and caused mitochondrial dysfunction by interrupting mitochondrial oxidative phosphorylation [64]. HG increased ROS-induced phosphoinositide 3-kinase (PI3K)/ protein kinase B (Akt)/ glycogen synthase kinase 3β (GSK3β) activity and accelerated epithelial-to-mesenchymal transition (EMT) in HK-2 cells [65]. Treatment with 4-hydroxy-2-hexenal increased ROS levels and increased ERK and JNK expression, triggering NF-kB activation and IkBα degradation in HK-2 cells.…”

Eat Your Broccoli: Oxidative Stress, NRF2, and Sulforaphane in Chronic Kidney Disease

“…SFN also decreased expression levels of the fibrosis-associated proteins collagen-I, alpha-smooth muscle actin, and vimentin [ 123 ]. In an HK2 cell culture model of hyperglycemia, treatment with SFN prevented high glucose-induced epithelial-to-mesenchymal transition, likely via an NRF2-dependent mechanism [ 124 ]. Pretreatment of HK2 cells with SFN protected against hypoxia–reoxygenation-induced cytotoxicity in a dose-dependent manner, and significantly augmented mRNA expression of phase 2 enzymes [ 118 ].…”

Glycogen synthase kinase-3β: a promising candidate in the fight against fibrosis

“…Besides, HG stimulates ROS production, leading to the inhibition of GSK-3β and the promotion of EMT process. Meanwhile, the expression level of epithelial cell marker E-cadherin is decreased, the mesenchymal markers vimentin and α-SMA are both promoted, as evidenced by induced renal fibrosis generation 42 , 79 . Moreover, knockdown of endogenous Nox2 significantly activates GSK-3β by inhibiting its phosphorylation in a rabbit CAVD model, along with significant attenuation of aortic valve ROS production and fibrosis 80 .…”