Klotho Prevents Epithelial-Mesenchymal Transition through Egr-1 Down-Regulation in Diabetic Kidney Disease

Background: As a key event leading to tubulointerstitial fibrosis in diabetic kidney disease (DKD), epithelial-mesenchymal transition (EMT) has drawn increasing attention from researchers. Klotho attenuates renal fibrosis in part by inhibiting ERK1/2 signaling in DKD. Early growth response factor 1 (Egr-1), activated mainly by ERK1/2, has been shown to play an important role in EMT. However, whether Klotho prevents EMT by inhibiting ERK1/2-dependent Egr-1 expression in DKD is unclear. The aim of this study was to investigate whether Klotho prevents EMT through Egr-1 down-regulation by inhibiting the ERK1/2 signaling pathway in DKD.

Methods: Male C57BL/6J mice fed an HFD for 4 weeks received 120 mg/kg STZ. Klotho and Egr-1 expression was detected in the renal cortices of these mice upon their sacrifice at 6 and 12 w after STZ treatment. In vitro studies, we incubated HK2 cells under high-glucose (HG) or TGF-β1 conditions to mimic DKD. We then transfected the cells with an Egr-1-containing plasmid, Egr-1 siRNA, a Klotho-containing plasmid or Klotho siRNA. Klotho, Egr-1, E-cadherin, α-smooth muscle actin (α-SMA), fibronectin(FN), ERK1/2 and p-ERK1/2 were determined.

Results: Klotho expression was significantly decreased in the renal cortices of diabetes mellitus (DM) mice compared to control mice at 6 w and was even more significantly decreased at 12 w. In contrast, Egr-1 expression was significantly increased in DM mice compared to control mice only at 12 w. We also found that Klotho overexpression down-regulated Egr-1 expression and the (p-ERK1/2)/(ERK1/2) ratio in HG- or TGFβ1-treated HK2 cells. Conversely, Klotho silencing up-regulated Egr-1 expression and the (p-ERK1/2)/(ERK1/2) ratio. Moreover, the effects of si-Klotho were abolished by the ERK1/2 inhibitor PD98059.

Conclusions: Klotho prevents EMT during DKD progression, an effect that has been partially attributed to Egr-1 down-regulation mediated by ERK1/2 signaling pathway inhibition.