Impact of klotho on the expression of SRGAP2a in podocytes in diabetic nephropathy

Diabetic nephropathy (DN) is the major cause of kidney failure, and glomerular podocytes play critical roles in the pathogenesis of DN by maintaining the glomerular structure and filtration barrier. Klotho and Slit-Robo GTP activating protein 2a (SRGAP2a) have been indicated to play protective roles in reducing kidney injury, but whether there is an internal relationship between these two factors is unclear.

In this study, we cultured differentiated rat podocytes in vitro and measured the SRGAP2a expressions by immunofluorescence staining, quantitative real-time PCR (qRT-PCR) and western blotting, after siRNA-mediated transforming growth factor β1 (TGF-β1) silencing, TGF-β1 overexpression and in the presence of a reactive oxygen species (ROS) inhibitor. And we detected the expressions of SRGAP2a, small mother against decapentaplegic (Smad)2/3, phosphorylated-Smad2/3 (p-Smad2/3), Smad7, and NAD(P)H oxidase 4 (NOX4), ROS levels and podocyte cytoskeletal remodelling under high glucose (HG) and exogenous klotho conditions. In addition, we performed haematoxylin-eosin (HE) staining and immunohistochemistry with diabetic rat models to confirm the in vitro results.

The results indicated that SRGAP2a expression was significantly upregulated under siRNA-mediated TGF-β1 silencing conditions or after adding a ROS inhibitor, but significantly downregulated with TGF-β1 overexpression, in the presence of HG. The supplementation of exogenous klotho under HG conditions significantly increased the SRGAP2a expression, remodelled the actin cytoskeleton and altered the expressions of Smad2/3, p-Smad2/3, Smad7 and NOX4 and reduced the ROS generation in podocytes. Moreover, klotho administration protected kidney injury in DN rats.

This study indicated that klotho may modulate the expression of SRGAP2a by regulating the ROS and TGF-β1 signalling pathways and provided theoretical support for klotho protein as a novel therapeutic strategy for treating DN patients.