GAS virulence factor SLO is a critical inducer for LAP induction. HMEC-1 cells were infected with wild-type or SLO deletion (SW974) NZ131 at an MOI of 5 for 30 min, and then gentamicin was added to kill extracellular bacteria. (A and B) Cells were collected at 1 h postinfection, followed by staining of CM-H2DCFDA and flow cytometric analysis. The data are shown as the dot plots and the histogram (A), as well as quantified by the percentage of the high-fluorescence subset (Q2) and the mean fluorescence intensity (MFI) (B). Data represent the means ± SD from three independent experiments. **, P < 0.01; ***, P < 0.001. (C and D) Cells were collected at 1 h postinfection and stained with anti-ULK1 (C), anti-NOX2 (D), and anti-LC3 antibodies. DAPI was used for cell nuclear and bacterial DNA staining. Images were obtained by confocal microscopy. Scale bars, 10 μm. LC3-positive GAS surrounded with ULK1 (C) or NOX2 (D) was relative to total intracellular LC3-positive GAS. Data represent the means ± SD from three independent experiments, and over 100 cells were counted in each sample. *, P < 0.05, and **, P < 0.01, compared to wild-type NZ131-infected cells.