TGF-β1 induces autophagy and promotes apoptosis in renal tubular epithelial cells

Transforming growth factor-beta 1 (TGF-beta 1) is a multifunctional cytokine that regulates cell growth, differentiation, apoptosis and autophagy in various cell types. It has been shown that TGE-beta 1-driven autophagy represents a novel mechanism of tubular decomposition, leading to renal interstitial fibrosis. However, the exact mechanism by which TGF-beta 1 regulates autophagy is still poorly understood. In the present study, we investigated the effects of exogenous TGF-beta 1 on cultured human renal proximal tubular epithelial cells (HRPTEpiCs). Presence of TGF-beta 1 in the medium induced accumulation of autophagosomes in a time- and dose-dependent manner as seen by monitoring the marker LC3 by confocal fluorescence microscopy and immunoblotting. In addition, TGF-beta 1 induced upregulation of autophagy-related genes, Atg5, Atg7 and Beclin1. Importantly, increased generation of reactive oxygen species (ROS) and enhanced expression of NADPH oxidases were found to be associated with the TGF-beta 1-induced autophagy. Conversely, treatment with inhibitors of NADPH oxidase markedly reversed the autophagic effects of TGF-beta 1. Apoptotic effects were evaluated by the TUNEL assay, measuring mitochondrial membrane potential and monitoring expression of the pro- and anti-apoptotic genes, Ban and Bcl-2, respectively. Transcriptional silencing of the above three autophagy-related genes in HRPTEpiCs caused attenuation of TGF-beta 1-mediated apoptosis. Similarly, when autophagy was prevented at an early stage by application of 3-methyladenine, the pro-apoptotic effects of TGF-beta 1 were attenuated. These observations suggest that in HRPTEpiCs TGF-beta 1 promotes autophagy through the generation of ROS, which contributes to its proapoptotic effect.