PBX1 attributes as a determinant of connexin 32 downregulation in Helicobacter pylori-related gastric carcinogenesis

Figure 1 Comparison of PBX1 and connexin 32 expression among developmental stages of Helicobacter pylori-associated gastric carcinoma in clinical specimens. Quantitative RT-PCR analysis of PBX1 (A) and Cx32 (B) in H. pylori (-) NGM and H. pylori (+) CAG and GC patients (n = 15). C: Representative images of the PBX1 and Cx32 protein levels in H. pylori (-) NGM and H. pylori (-/+) CAG, IM and GC tissues. ^bP < 0.01, ^eP < 0.001. NGM: Normal gastric mucosa; CAG: Chronic atrophic gastritis; GC: Gastric cancer; IM: Intestinal metaplasia.

Figure 2 Effects of Helicobacter pylori infection on PBX1 and connexin 32 expression in GES-1 cells. Quantitative RT-PCR analysis of PBX1 (A) and Cx32 (B) in GES-1 cells with 24- and 48-h H. pylori infection; C: Western blot analysis of PBX1 and Cx32 protein levels in GES-1 cell under 24- and 48-h H. pylori infection. ^bP < 0.01, ^eP < 0.001.

Figure 3 PBX1 negatively regulates connexin 32 by directly binding to its promoters. Quantitative RT-PCR analysis of PBX1 (A) and Cx32 (B) levels in GES-1 cells enforced with the PBX1 overexpressing (OE) vector and transfected with a negative control (NC) vector. Quantitative RT-PCR analysis of PBX1 (C) and Cx32 (D) levels in human adenocarcinoma BGC823 cells treated with a scrambled control, PBX1 siRNA-1, PBX1 siRNA-2, and PBX1 siRNA-3. E: PBX1 and Cx32 protein levels in BGC823 treated with a scrambled control, PBX1 siRNA-1, PBX1 siRNA-2, and PBX1 siRNA-3. F: PBX1 directly binds to Cx32 promoters. HEK293T cells co-transfected with pDoubleEx-EGFP-PBX1 and Cx32-pGL3 vectors. The results are displayed as the ratio of firefly luciferase activity in pGL3-Cx32-PBX1-transfected cells to the activity in Cx32-pGL3 controls. ^bP < 0.01, ^eP < 0.001.

Figure 4 Effects of Helicobacter pylori on PBX1 and connexin 32 expression in Mongolian gerbils. Alteration in the PBX1 (A) and Cx32 (B) mRNA expression levels in Mongolian gerbils with 48-wk H. pylori infection (n = 21) compared with those in the control group (n = 4). ^eP < 0.001; C: Correlation between the PBX1 and Cx32 mRNA levels in Mongolian gerbils (21 from the experimental group and 4 from the control group); D: Pathological outcome of Mongolian gerbils (HandE staining 400 ×) after 48-wk H. pylori infection: NGM with no H. pylori infection; NAG (mild/moderate/severe inflammation), CAG, and IM in the experimental group. Scale bar: 20 μm. NGM: Normal gastric mucosa; NAG: Non-atrophic gastritis; CAG: Chronic atrophic gastritis; IM: Intestinal metaplasia.