Nutlin-3a-induced methuosis-like cell death is associated with the HBP in KRAS MT/p53 WT NSCLC cells. Cells were supplemented with GlcNAc (40 mM) or GlcN (200 μM) for 24 h with or without nutlin-3a (30 μM). A, B Both GlcNac and GlcN rescue cell proliferation and death, which was analyzed by cell counting (A), and flow cytometry using annexin V/PI staining (B). Representative flow cytometry graph, and statistical histogram of four independent experiment are shown. C Live cell images captured by holotomography microscopy show that GlcNAc supplementation rescues the formation of huge cytoplasmic vacuoles induced by nutlin-3a. D Cells were incubated with dextran and LysoTracker for 2 h with GlcNAc or nutlin-3a. GlcNAc supplementation with nutlin-3a recovers dextran uptakes compared to only nutlin-3a treated cells (n ≥ 22). E Autophagy flux disruption recovered by supplementation of GlcNAc with nutlin-3a, which was detected after transfection of mRFP-GFP-LC3 tandem vector (n ≥ 20). Representative images were obtained by confocal laser scanning microscopy and quantitated from the captured image (D, E). Scale bar: 7 μm for (C), and 10 μm for (D-E). *p < 0.05, **p < 0.01, ***p < 0.001 compared with control
|
