Starvation-induced NEDD4-mediated autophagy in BMSCs

Autophagy regulates the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) and mediates BMSC-mediated bone remodeling under pathological conditions (such as osteoporosis and infectious bone defects). However, the regulatory mechanisms controlling autophagy in BMSCs remain unclear. Neural precursor cell-expressed developmentally down-regulated protein 4 (NEDD4) has been implicated in autophagy regulation. In the present study, autophagy was induced in BMSCs through serum starvation and NEDD4 expression was silenced using lentiviral short hairpin RNA. Quantitative real-time polymerase chain reaction, western blotting, and immunofluorescence were used to examine the expression of the autophagy-related molecules, microtubule-associated protein 1 light chain 3 (LC3) and Beclin-1, to explore the underlying molecular mechanisms. It was determined that BMSCs exhibited significant autophagy activation 30 min after starvation, accompanied by marked increases in NEDD4, LC3, and Beclin-1 at both mRNA and protein levels. NEDD4 knockdown significantly attenuated the upregulation of LC3 and Beclin-1 and prevented the starvation-induced decrease in phosphorylated mTOR. These results indicated that NEDD4 positively regulates autophagy in BMSCs, likely through the mTOR signaling pathway. In conclusion, the present study demonstrated that NEDD4 is essential for BMSC autophagy and may serve as an important target for therapies aimed at modulating BMSC function.