miR-21 Targets ASPP2 to Inhibit Apoptosis via CHOP-Mediated Signaling in Helicobacter pylori-Infected Gastric Cancer Cells

Figure 3

ASPP2 is a miR-21 target in HPGC. (a) AGS and MKN45 cells were transfected with control, miR-21 mimic, or anti-miR-21 oligonucleotides. ASPP2 protein levels in the cell lysates were analyzed via immunoblotting. (b) The binding sites between miR-21 and ASPP2 3′UTR were evaluated using TargetScan analysis (upper). Subsequently, AGS cells transfected with the indicated vectors and relative luciferase activity in AGS cells with wild-type or mutant ASPP2 3′UTR were analyzed by luciferase reporter assay (lower). (c) AGS and MKN45 cells transfected with control or anti-miR-21 oligonucleotides were incubated with H. pylori at 30 and 90 MOI or PBS for 6 hours. Relative ASPP2 mRNA levels were measured by qPCR. All the data were presented as mean ± SD of 3 replicates. (d) Relative ASPP2 expression levels in HPGC tumor tissues (n = 35) as compared to adjacent normal tissues (n = 35) by IHC and the correlation with miR-21 levels. Upper left, representative images were captured at 200x magnification. Scale bar, 50 μm. Lower left, relative quantitation of ASPP2 levels by H score ranking. The significance of differences in H score between the two groups was compared using the nonparametric Mann–Whitney U test. Right, Spearman correlation analysis between miR-21 and ASPP2 levels in HPGC tumor tissues (n = 35).  < 0.05,  < 0.01, and  < 0.001.

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