Autophagy protects osteoblasts from advanced glycation end products-induced apoptosis through intracellular reactive oxygen species

Patients with type II diabetes are susceptible to fracture; however, these patients typically have normal bone mineral density. Thus, such fractures cannot be entirely explained by advanced glycation end products (AGEs)-induced osteoblast apoptosis. Autophagy is a molecular process allowing cells to degrade unnecessary or dysfunctional cellular organelles, and closely interacts with apoptosis. The aim of this study was to determine whether autophagy participated in the pathology of AGEs-treated osteoblasts, and the possible mechanism of such an involvement. Osteoblastic MC3T3-E1 cells were used. Autophagy was evaluated by detecting the level of LC3 via western blotting and immunofluorescence. p62/SQSTM1 expression was also assessed by western blotting. The autophagy inducer rapamycin (RA) and the autophagy inhibitor 3-methyladenine were used to determine whether autophagy has effect on AGEs-induced apoptosis. N-Acetylcysteine (NAC), reactive oxygen species (ROS) inhibitor, was used to determine whether ROS and mitochondrial damage were involved in autophagy regulation. The results showed that the autophagy level was increased in MC3T3-E1 cells treated with AGEs, as represented by an increase in both the total LC3 level and the LC3II/LC3I ratio, as well as a decrease in p62/SQSTMI expression. Further inducing autophagy by RA attenuated AGEs-induced apoptosis. The antioxidant NAC suppresses AGEs-induced autophagy in osteoblastic MC3T3-E1 cells. These results demonstrate that autophagy participates in the pathology of AGEs-treated osteoblasts, and may play a protective role in AGEs-induced apoptosis in osteoblastic MC3T3-E1 cells. ROS and mitochondrial damage are essential in upregulating AGEs-induced autophagy.