Measurement of cellular reactive oxygen species (ROS)

Cells were incubated with fluorescent ROS probe dichloro-dihydro-fluorescein diacetate (DCFH-DA, S0033M-1, Beyotime, Shanghai, China) for 30 min. Then, cells were washed with DPBS (14190144, Thermo Fisher Scientific, Waltham, MA, USA). A fluorescence microscope (DMIL LED fluo, Leica, Wetzlar, Germany) was applied to observe and capture cell images. ImageJ software was then used to calculate the integrated density and cell area, with the integrated density subsequently normalized by the cell area.