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. 2018 Jan 15;17(2):250–262. doi: 10.1080/15384101.2017.1417708

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Figure 5. — ZEB2 expression pattern and MiR-200a directly targets and inhibits ZEB2 424 expression

(a) The expression of ZEB2 in ethanol treated mice was analyzed by immunofluorescence assay (original magnification, × 20). (b) Total mRNA and protein of ZEB2 extracted from the primary hepatocyte and were analyzed by qPCR and western blot. The results are shown as relative expression against control expression without treatment. (c) The expression of ZEB2 was analyzed by qPCR and western blot in ethanol stimulated AML-12 cells. The results are shown as relative expression against control expression without treatment. (d) Computational analysis of the ZEB2 3′UTR revealed putative conserved miR-200-binding sites. (e) Luciferase reporter assay was performed on AML-12 cells to detect the relative luciferase activities of Wild type and mutation ZEB2 reporters. Data shown are the mean ± SD from 3 independent experiments. (f) The successful over-expression of ZEB2 was verify by western blot and qPCR. The results are shown as relative expression against control expression without treatment. (g) ZEB2 protein and mRNA expression were analyzed by western blot and qPCR in AML-12 cells transfected with miR-200a inhibitor or mimics. Data were presented as mean ± SD from three independent experiments. * P < 0.05, ** P < 0.01 versus control. # P< 0.05, ## P< 0.01 versus ethanol treatment group, ns, not significant versus ethanol treatment group.

graphic file with name kccy-17-02-1417708-g007.jpg — ZEB2 expression pattern and MiR-200a directly targets and inhibits ZEB2 424 expression

(a) The expression of ZEB2 in ethanol treated mice was analyzed by immunofluorescence assay (original magnification, × 20). (b) Total mRNA and protein of ZEB2 extracted from the primary hepatocyte and were analyzed by qPCR and western blot. The results are shown as relative expression against control expression without treatment. (c) The expression of ZEB2 was analyzed by qPCR and western blot in ethanol stimulated AML-12 cells. The results are shown as relative expression against control expression without treatment. (d) Computational analysis of the ZEB2 3′UTR revealed putative conserved miR-200-binding sites. (e) Luciferase reporter assay was performed on AML-12 cells to detect the relative luciferase activities of Wild type and mutation ZEB2 reporters. Data shown are the mean ± SD from 3 independent experiments. (f) The successful over-expression of ZEB2 was verify by western blot and qPCR. The results are shown as relative expression against control expression without treatment. (g) ZEB2 protein and mRNA expression were analyzed by western blot and qPCR in AML-12 cells transfected with miR-200a inhibitor or mimics. Data were presented as mean ± SD from three independent experiments. * P < 0.05, ** P < 0.01 versus control. # P< 0.05, ## P< 0.01 versus ethanol treatment group, ns, not significant versus ethanol treatment group.