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. 2023 May 2;299(6):104779. doi: 10.1016/j.jbc.2023.104779

Figure 4.

Figure 4

Inhibiting DNA methylation in vitro restores autophagy.A, Oil red O staining confirmed increased oil globules in palmitate (500 μM) and LPS (100 ng/ml)-treated compared with control RAW 264.7 macrophages (the scale bar represents 200 pixels). B, RAW 264.7 cells with or without palmitate and LPS treatment are subjected to Western blotting against p-ULK, T-ULK, SIRT-1, p-AMPK, and T-AMPK, and β-actin. C, bar graphs represent the densitometric ratio of p-ULK to T-ULK. D, bar graphs represent the densitometric ratio of SIRT-1 to the corresponding β-actin. E, bar graphs represent the densitometric ratio of pAMPK to T-AMPK. F, RAW 264.7 cells pretreated with palmitate and LPS treated with Zebularine (0.5 μM and 5 μM) and Azacytidine (0.25 μM and 0.5 μM) where subjected to Western blotting against protein expression of SIRT1, p-ULK, p-AMPK, and T-AMPK. G, bar graphs represent the densitometric ratio of p-ULK to the corresponding β-actin. H, bar graphs represent the densitometric ratio of SIRT-1 to the corresponding β-actin. I, bar graphs represent the densitometric ratio of pAMPK to T-AMPK. J, RAW 264.7 cells pretreated with palmitate and LPS treated with Zebularine (0.5 μM and 5 μM) and Azacytidine (0.25 μM and 0.5 μM) were subjected to Western blotting against DNMT1 and autophagic proteins (ATG-7, ATG-5, Beclin1, and LC3I/II). K, bar graphs represent the densitometric ratio of DNMT1 to the corresponding β-actin. L, bar graphs represent the densitometric ratio of ATG-7 to the corresponding β-actin. M, bar graphs represent the densitometric ratio of ATG5 to the corresponding β-actin. N, bar graphs represent the densitometric ratio of Beclin-1 to the corresponding β-actin. O, bar graphs represent the densitometric ratio of LC3-II to the corresponding β-actin. Data are represented as mean ±SD, ∗∗p < 0.01, and ∗∗∗p < 0.001 versus Control group; #p < 0.05, ##p < 0.01, and ###p < 0.001 versus Palmitate and LPS-treated group. Error bars (wherever applicable in the figure) represent standard deviations from 3 to 4 independent experiments.