Fig. 6

MT is involved in MLT-mediated autophagy regulation in cells undergoing long-term ex vivo passaging. (CON: P15 cells without any treatment; MLT: MLT-treated P15 cells; LUZ: LUZ-treated P15 cells; MLT + LUZ: MLT plus LUZ–treated P15 cells). *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001 represent significant differences between the indicated columns, while NS represents no significant difference. a Representative confocal images of autophagosomes (yellow) and their matured form, autolysosomes (red), in mRFP-GFP-LC3 adenovirus-transfected P15 cells with or without MLT/LUZ treatment (scale bar = 50 μm, ×200 magnification). b Quantitative analysis of the percentage of red puncta out of the total puncta based on confocal examination. c Cell autophagy-related proteins Atg7, p62, Beclin-1, and LC3-II were analyzed by Western blot assay. d Quantitative analysis of Atg7, p62, Beclin-1, and LC3-II expression in P15 cells with or without MLT/LUZ treatment. e SA-β-gal in P15 cells with or without MLT/LUZ treatment (top: representative images showing SA-β-gal expression by immunocytochemical staining; positively stained cells were dyed blue, scale bar = 100 μm; bottom: percentage of SA-β-gal-positive cells among all cells and quantitative analysis). f Cell senescence-related protein expression in P15 cells with or without MLT/LUZ treatment as determined by Western blot assay. g Quantitative analysis of p53, p21, p16, and γ-H2AX expression in P15 cells with or without MLT/LUZ treatment