Figure 4. Sestrin 1 knockdown significantly inhibited autophagy in KGN cells. KGN cells were transfected with negative control (siRNA-ctrl) or Sestrin 1 siRNA2, and rapamycin was added 48 h after transfection. (A) Immunofluorescence staining for LC3 was used to evaluate the effects of sestrin 1 downregulation on autophagy in KGN cells by calculating the percentage of cells that were LC3 positive. (B) Western blots were performed to determine levels of Atg7, p62, p-mTOR, mTOR, Beclin 1 and LC3 I/II. β-actin was used as an internal control. (C–G) Atg7, p62, p-mTOR, Beclin 1 and LC3 I/II expressions were quantified. (H) The formation of autophagosome in each group was observed by the electron microscope. **P < 0.01 compared to the blank control. ##P < 0.01 compared to the sestrin 1 siRNA2 group.