Renal tubular injury is associated with the development of diabetic nephropathy (DN) and the end-stage renal disease (ESRD). Free fatty acids (FFAs)-associated lipotoxicity contributes to injury of proximal renal tubular epithelial (HK-2) cells in diabetes. Palmitic acid (PA) which is the most abundant saturated fatty acid in FFAs is closely associated with the gradual decline of renal function. Astragalosides IV (AS-IV) has a variety of pharmacological effects such as anti-inflammation and anti-oxidation. In the current study, we investigated the effects of AS-IV on PA-induced apoptosis of HK-2 cells and the underlying mechanisms. The results showed that AS-IV (10, 20, 40 μmol/L) could alleviate PA-induced apoptosis of HK-2 cells. We found that AS-IV reduced the expression of Bax and cleaved-caspase3, but increased the expression of Bcl-2 and phosphorylated Nrf2 in HK-2 cells. Moreover, AS-IV reduced the level of reactive oxygen species (ROS) in the cells. Our study suggests that AS-IV could protect against PA-induced apoptosis in HK-2 cells by inhibiting ROS generation and apoptotic protein expression. This study may provide a new theoretical option for the patients with type 2 diabetes.