| AIM: To investigate gastric cancerogenesis induced by Helicobacter pylori(H. pylori)-through mitochondria- cytochrome c apoptotic pathway.METHODS: (1) Specimens of 95 patients (72 males and 23 females, age range 31 to 84 years, mean 56 years), who had undergone gastric adenocarcinoma resection surgery at the Southwest Hospital in Chongqing and had not taken anti-H. pylori drugs before operation, were collected from 2001 to 2002. Gatric adenocarcinoma tissues were examined microscopically. Resection margin tissues were also examined to verify that they did not contain malignant cells. The histological diagnosis was confirmed by a professional pathologist. The remaining specimens were snap-frozen and stored at -80℃ until assayed. Warthin-Starry silver staining and polymerase chain reaction (PCR) analysis for H. pylori urease gene A (ureA) were performed to detect H. pylori infection. PCR analysis for H. pylori cagA gene was performed to verify cagA+ H. pylori infection. Fifty-eight patients whose both Warthin-Starry staining and PCR for ureA showed positive results were diagnosed as suffering from H. pylori infection and 37 suffered from cagA+ H. pylori. (2) H. pylori strains NCTC 11637 were cocultured with human gastric adenocarcinoma epithelial cell lines SGC-7901 and AGS cells, and apoptosis were assessed by Flow cytometry. Semi-quantitative RT-PCR was used to measure Bid, Bax, Bcl-2, IL-1β, cytochrome c, Caspase-9 and Caspase-3 mRNA expressions. Western blotting analysis was used to measure Bid, Bax, Bcl-2, cytochrome c, Caspase-9 and Caspase-3 protein expressions. RESULTS: (1) Expressions of Bid and Bax in gastric adenocarcinoma tissues without H. pylori infection, with cagA- H. pylori infection and cagA+ H. pylori infection increased significantly in turn (Bid, 0.30, 0.42 and 0.85 respectively, P<0.05; Bax, 0.30, 0.65 and 0.97 respectively, P<0.05). Bcl-2 mRNA levels increased significantly in gastric adenocarcinoma tissues with cagA- H. pylori infection and cagA+ H. pylori infection, compared with those without H. pylori infection (0.69 and 0.84 vs. 0.41, P<0.05). Expressions of Bid, Bax and Bcl-2 in resection margin tissues without H. pylori infection, with cagA- H. pylori infection and cagA+ H. pylori infection increased significantly in turn (Bid, 0.37, 0.68 and 0.93 respectively, P<0.05; Bax, 0.35, 0.64 and 1.00 respectively, P<0.05; Bcl-2, 0.37, 0.48 and 0.61 respectively, P<0.05). In H. pylori negative specimens, expressions of Bid and Bax correlated negatively with that of Bcl-2 respectively in adenocarcinoma tissues (Bid vs. Bcl-2, r = -0.40, P<0.05; Bax vs. Bcl-2, r = -0.45, P<0.05). In H. pylori positive specimens, expressions of Bid and Bax did not correlate with that of Bcl-2 in adenocarcinoma tissues (Bid vs. Bcl-2, r = 0.18, P>0.05; Bax vs. Bcl-2, r = 0.20, P>0.05), but correlated positively with that of Bcl-2 respectively in resection margin tissues (Bid vs. Bcl-2, r = 0.33, P<0.05; Bax vs. Bcl-2, r = 0.29, P<0.05). H. pylori could upregulate IL-1β mRNA levels, especially cagA+ H. pylori. Expressions of IL-1β mRNA correlated positively with that of Bid and Bax in adenocarcinoma tissues infected by H. pylori (IL-1β vs. Bid,r = 0.50,P<0.05;IL-1β vs. Bax,r = 0.46,P<0.05). Cytochrome c, Caspase-9 and Caspase-3 mRNA and protein expression did not have significant differences among gastric adenocarcinoma and resection margin tissues without H. pylori infection, with cagA- H. pylori infection and cagA+ H. pylori infection, but active form of Caspase-9 and Caspase-3 in gastric adenocarcinoma and resection margin tissues without H. pylori infection, with cagA- H. pylori infection and cagA+ H. pylori infection increased in turn. (2) NCTC 11637 could induce SGC-7901 and AGS cells apoptosis in dose- and time-depend manner. NCTC 11637 could upregulate Bid, Bax, Cytochrome c, Caspase-9 and Caspase-3 mRNA expression in a time-depend manner after coculturing with SGC-7901 and AGS cells, but could not regulate Bcl-2 mRNA expression. Apoptosis could be inhibited almost completely by prior incubati... |
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