Lipotoxicity, intracellular lipid metabolites accumulation, has been proposed as an important pathogenic mechanism contributing to kidney dysfunction and disease. This study was aimed to investigate the effect of palmitic acid, the dominant saturated fatty acid, on apoptosis and exosome release by renal tubular epithelial cells. A rat proximal-tubule epithelial cell line, NRK-52E, was incubated with 1% bovine serum albumin (BSA), BSA-conjugated palmitic acid (PA, 250 µM), or BSA-conjugated oleic acid (OA, 250 µM) for 24-48 hours. Lipotoxicity was determined by fluorescence microscopy after staining with BODIPY 493/503. Cell proliferation and viability were examined via cell cycle analysis using Cellometer imaging cytometry, Annexin-V, and MTT-assays. Western blot analysis of apoptotic proteins and exosome-marker proteins was performed to evaluate the effects of PA. Confocal microscopy confirmed an increase in lipid accumulation in both treatments. Incubation of NRK-52E cells with PA resulted in a timed-dose-dependent increase in cell apoptosis, whereas OA had a stimulatory effect on cell proliferation. Compared to BSA control, PA treatment significantly increased cleaved caspase-3 in cell-lysates and Alix and TSG101 proteins in exosomes of NRK-52E cells. Our results indicate that saturated palmitic acid stimulates apoptosis and exosome release from cultured tubular epithelial cells. To conclude, the results suggest evaluating signaling pathways contributing to nephropathy-progression.
