Silencing miR-208a-3p promotes autophagy and attenuates high glucose-triggered podocyte injury by activating VAV3/AKT/mTOR pathway.

Abstract

Dysregulation of microRNAs (miRNAs) has been associated with podocyte injury in diabetic nephropathy (DN). This study aimed to investigate the role and underlying mechanism of miR-208a-3p in the development of DN. An in vitro DN model was established by treating podocytes with high glucose (HG). Gene expression was analyzed using reverse transcription quantitative polymerase chain reaction, and protein levels were determined by Western blotting. Immunofluorescence staining was performed to assess microtubule-associated protein 1A/1B-light chain 3 (LC3) intensity as an autophagy marker. Podocyte viability and apoptosis were evaluated via Cell Counting Kit-8 assay and flow cytometry, respectively. Intracellular reactive oxygen species (ROS) levels were measured using 2',7'-dichlorofluorescin diacetate staining. miR-208a-3p was significantly upregulated in HG-stimulated podocytes. HG exposure promoted apoptosis and oxidative stress, while suppressing autophagy and cell viability, effects that were reversed by miR-208a-3p inhibition. Mechanistically, miR-208a-3p directly targeted vav guanine nucleotide exchange factor 3 (VAV3). HG suppressed AKT/mTOR signaling, whereas miR-208a-3p silencing activated it. Conversely, VAV3 knockdown inactivated AKT/mTOR signaling. VAV3 downregulation or treatment with LY294002, an AKT/mTOR inhibitor, abolished the protective effects of miR-208a-3p silencing against HG-induced podocyte injury. These findings indicate that miR-208a-3p silencing attenuates HG-induced podocyte injury by promoting autophagy via targeting the VAV3/AKT/mTOR pathway.