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. 2021 Sep 27;9:722960. doi: 10.3389/fcell.2021.722960

FIGURE 4.

FIGURE 4

SESN2 regulates autophagy and reduces the sensitivity of osteosarcoma cells to chemotherapy. In the presence or absence of 3-MA (5 mM), SESN2-overexpressing and control HOS cells were treated with Dox (0.2 μg/mL) for 24 h, and apoptosis was analysed by flow cytometry (A) (n = 3). After treatment with Cis (20 μmol/L) or Dox (0.2 μg/mL), the expression levels of LC3 and P62 in SESN2-overexpressing and control HOS cells were detected by western blot (B) (n = 3). The expression levels of LC3 and P62 in SESN2-overexpressing and control HOS cells treated with 3-MA (5 mM) were detected by western blot (C) (n = 3). In the presence or absence of 3-MA (5 mM), SESN2-overexpressing and control HOS cells were treated with Dox (0.2 μg/mL) for 24 h, cell activity was detected by CCK-8 (D) (n = 3), intracellular autophagosomes were observed by TEM (E) (n = 3, scale bar = 2 μm), and intracellular LC3 puncta formation was analysed by immunofluorescence (F) (n = 3, scale bar = 20 μm). After treatment with Cis (20 μmol/L) or Dox (0.2 μg/mL), autophagosome formation in HOS cells with ectopic SESN2 expression was monitored by immunofluorescence through transfection with RFP-GFP-LC3 lentivirus after upregulating SESN2 (G) (n = 3, scale bar = 10 μm). The data are presented as the mean ± SD. *p < 0.05 vs. the pHBLV control group.