SUN Feng-mei, ZHANG Meng-ying, ZHANG Hong, PU Yue-pu, ZHANG Juan. Effects and mechanism of autophagy induced by 1, 4-benzoquinone in K562 cells[J]. Journal of Environmental and Occupational Medicine, 2018, 35(5): 411-417. DOI: 10.13213/j.cnki.jeom.2018.17564
Citation: SUN Feng-mei, ZHANG Meng-ying, ZHANG Hong, PU Yue-pu, ZHANG Juan. Effects and mechanism of autophagy induced by 1, 4-benzoquinone in K562 cells[J]. Journal of Environmental and Occupational Medicine, 2018, 35(5): 411-417. DOI: 10.13213/j.cnki.jeom.2018.17564

Effects and mechanism of autophagy induced by 1, 4-benzoquinone in K562 cells

  • Objective To investigate the effects and mechanism of 1, 4-benzoquinone (1, 4-BQ) on autophagy activity in K562 cells.

    Methods Human chronic myelogenous leukemia K562 cells were divided into control group, l, 4-BQ (5, 10, and 20 μmol/L) groups, early-stage autophagy inhibitor LY294002 (10 μmol/L) group, late-stage autophage inhibitor chloroquine (CQ) (10 μmol/L) group, 1, 4-BQ (20μmol/L)+LY294002 (10μmol/L) group, and 1, 4-BQ (20μmol/L)+CQ (10μmol/L) group. The proliferation of K562 cells was evaluated by MTT assay, qualitative and quantitative autophagy level by fluorescent staining and flow cytometry assay using CYTO-ID fluorescence probe, lysosomal pH value by acridine orange staining method, and the gene and protein expressions of LC3-Ⅱ and P62 by real-time quantitative PCR and Western blot, respectively.

    Results The relative proliferation rates of the K562 cells treated with 10 and 20 μmol/L 1, 4-BQ remarkably decreased at 24, 48, and 72 h (P < 0.05). CYTO-ID dye was typically accumulated in the cytoplasm of K562 cells, and the fluorescent signals raised with increasing 1, 4-BQ concentration. The flow cytometry results revealed that, compared with the control group, the autophagy level was significantly increased in the cells treated with 5, 10, and 20 μmol/L 1, 4-BQ (P < 0.05). The acridine orange staining results showed stronger red fluorescence of the 1, 4-BQ groups than that of the control group, indicating decreased lysosomal pH values of the 1, 4-BQ groups. The results of Western blot showed that the LC3-Ⅱ and P62 protein expression levels were increased in the 10 and 20μmol/L 1, 4-BQ groups compared with the control group (P < 0.05), and peaked at 24 h. After the designed intervention, the autophagy level and LC3-Ⅱprotein expression level were lower in the 1, 4-BQ+LY294002 group than that of the 1, 4-BQ group (P < 0.05). The autophagy level in the 1, 4-BQ+CQ group was higher than that in the 1, 4-BQ group (P < 0.05), but not different from that in the CQ group. Additionally, the protein expression level of LC3-Ⅱ in the CQ group was higher than that in the control group (P < 0.05). There was no difference in the protein expression level of LC3-Ⅱ between the 1, 4-BQ group and the 1, 4-BQ+CQ group (P>0.05).

    Conclusion 1, 4-BQ can increase LC3-Ⅱ and P62 protein expression levels, indicating elevated autophagy level, with a potential pathway of increasing autophagic synthesis and decreasing autophagic degradation.

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