Abstract
Autophagy is a highly conserved degradation process that is involved in the clearance of proteins and damaged organelles to maintain intracellular homeostasis and cell integrity. Type 2 diabetes is often accompanied by dyslipidemia with elevated levels of free fatty acids (FFAs). Podocytes, as an important component of the filtration barrier, are susceptible to lipid disorders. The loss of podocytes causes proteinuria, which is involved in the pathogenesis of diabetic nephropathy. In the present study, we demonstrated that palmitic acid (PA) promoted autophagy in podocytes. We further found that PA increased the production of reactive oxygen species (ROS) in podocytes and that NAC (N-acetyl-cysteine), a potent antioxidant, significantly eliminated the excessive ROS and suppressed autophagy, indicating that the increased generation of ROS was associated with the palmitic acid-induced autophagy in podocytes. Moreover, we also found that PA stimulation decreased the mitochondrial membrane potential in podocytes and induced podocyte apoptosis, while the inhibition of autophagy by chloroquine (CQ) enhanced palmitic acid-induced apoptosis accompanied by increased ROS generation, and the stimulation of autophagy by rapamycin (Rap) remarkably suppressed palmitic acid-induced ROS generation and apoptosis. Taken together, these in vitro findings suggest that PA-induced autophagy in podocytes is mediated by ROS production and that autophagy plays a protective role against PA-induced podocyte apoptosis.
Highlights
Podocytes play a crucial role in maintaining the glomerular filtration barrier[5,6]
To examine the effect of lipid disorders on autophagy in podocytes, we treated MPC5 cells with 150 μmol/L palmitic acid (PA), a saturated free fatty acids (FFAs), for different lengths of time and found that the expression of Beclin[1], LC3-II/LC3-I and Lysosomal-associated membrane protein 2 (LAMP-2) in podocytes increased in a time-dependent manner, as demonstrated by western blot analysis (Fig. 1a and c)
There was no significant difference in Beclin[1], LC3-II/LC3-I and LAMP-2 expression found between podocytes treated with 150 μmol/L and 300 μmol/L palmitic acid for 24 hours (Fig. 1b and d), which suggested that 150 μmol/L was the appropriate dose of palmitic acid for the induction of autophagy
Summary
Apoptosis and detachment of podocytes lead to proteinuria in patients with diabetic nephropathy, and the reduction in the number of podocytes is an important index used to predict the progression of DN7,8. Autophagy is a highly conserved lysosomal degradation pathway that degrades cellular unfolded/misfolded proteins and damaged/unnecessary organelles, to maintain intracellular homeostasis and cell survival. A strong body of evidence suggests that autophagy plays a critical role in kidney maintenance and diseases[17] and that insufficient podocyte autophagy may be involved in the pathogenesis of podocyte loss, leading to massive proteinuria and the rapid progression of diabetic nephropathy, and autophagy is considered to be a potential novel therapeutic strategy for DN18–20. Podocytes are susceptible to saturated FFAs21, and it has been reported that high-fat feeding enhanced autophagy in beta cells[22] and that palmitic acid (PA) induced autophagy in hepatocytes[23]. We aimed to investigate the significance of autophagy in palmitic acid-induced podocyte apoptosis using an in vitro model and to clarify its mechanism
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