H. PYLORISTRAINS ISOLATED FROM PATIENTS WITH GASTRIC CANCER EXPRESS SIGNIFICANTLY HIGHERN-NITROSATING ACTIVITIES THAN STRAINS COLLECTED FROM DUODENAL ULCER AND NON-ULCER DYSPEPSIA

Abstract

Background: Chronic Helicobacter pylori infection and exposure to N-nitroso compounds (NOC) are considered the two most important contributing factors in gastric carcinogenesis. It has been shown in a previous study that H. pylori also catalyzes N-nitrosation in vitro. Whether the N-nitrosating activity of H. pylori plays any role in the development of gastric adenocarcinoma has yet to be determined. Methods:H. pylori strains isolated from patients with different gastroduodenal pathologies were subcultured with the addition of morpholine (a nitrosable chemical) and sodium nitrite at a concentration of 25mM and 16mM, respectively. The bacteria were allowed to grow for 6 to 8 hours until OD600 of the culture media reached a value between 0.6 and 0.8. The culture-supernatants were then separated at 4°C, and the quantity of N-nitroso-morpholine formed was assayed using gas chromatography, as described by Suzaki and Mitsuoka. The nitrosating activity of individual H. pylori strain was calculated by adjusting the OD600 to unity. Bacterial genotypes, cagA, iceA and vacA of the H. pylori isolates were also determined by polymerase chain reaction based on published primer sequences. N-nitrosating activities of the H. pylori strains were compared with regard to the patient characteristics, the bacterial genotypes, and their related disease categories. Results: 20 strains from gastric cancer (GC), 27 from duodenal ulcer (DU) and 22 from non-ulcer dyspepsia (NUD) were studied. Though patients with GC were significantly older than those with DU and NUD, the 3 groups were comparable with regard to patient’s gender and bacterial genotypes. All 69 H. pylori strains expressed N-nitrosating activities with a uni-modal distribution (median 41.3 μg/L; range: 9.2–152.3 μg/L). Strains isolated from GC patients had a significantly higher mean N-nitrosating activity (65.3 ± 27.2 μg/L) than those collected from patients with DU (43.8 ± 24.9 μg/L) or NUD (33.8 ± 21.1 μg/L) (p = 0.003, one-way ANOVA), but there was no difference between the DU and the NUD groups. Neither the patient characteristics (age and gender) nor the bacterial cagA, iceA and vacA subtypes had any association with the nitrosating activity. Conclusions:H. pylori-catalyzed N-nitrosation may be important in gastric carcinogenesis as strains isolated from stomach cancer patients express a higher level of N-nitrosating activity than those found in non-cancer patients.