2.4 | In vitro cell membrane injury assay
We performed an in vitro cell membrane injury repair assay following the protocols described previously.26, 29 HNECs were treated with TGF-β1/TGF-β1 inhibitor or transfected with Ad-MG53 for 72 h, and then suspended in PBS. Next, 100 μL of the cell suspension (5 × 104 cells/well) was added to a 96-well plate along with acid-washed glass microbeads. The plate was shaken at 200 rpm for 5 min to induce cell membrane injury, followed by centrifugation at 3000 rpm for 10 min, and 80 μL of the supernatant was collected. Lactate dehydrogenase (LDH) activity in the supernatant was determined using the LDH Cytotoxicity Detection Kit (MeiKe, MK0354A, China).