Extensive DNA damage induced by monochloramine in gastric cells

Colonization of Helicobacter pylori (Hp) to gastric mucosa plays an important role for the pathogenesis of gastric mucosal lesions. We previously reported the importance of monochloramine (NH₂Cl), which was derived from the interaction between Hp-urease and infiltrated leukocytes, in the course of Hp-associated gastric mucosal injury. While the long-term infection of Hp in the gastric mucosa is known to be one of the virulent factors which closely link to the gastric carcinogenesis, the details of its pathogenetic mechanisms remain speculative. The present study is designed to examine whether a NH₂Cl could damage the DNA of gastric cells. Rabbit gastric mucosal cells (RGMC) or KATO III cells were cultured and suspended. Cell suspensions were exposed to HOCl, NH₃ or NH₂Cl for 15 min to give a final concentration of 0.1 mM. The magnitude of a double strand break of DNA was quantified by measuring the remnant double strand stained by ethidium bromide (EB), and the fluorescence intensity of EB was analyzed by spectrophotometer. Separately, cell nuclei were stained by fluorescent dye (Hoechst No. 33258) in order to evaluate the levels of chromatin condensation evoked by DNA fragmentation. The number of cells with chromatin condensation was counted. During the entire experimental period, more than 85% of cells were persistently viable in all groups. NH₂Cl significantly induced the DNA double strand break as well as chromatin condensation in RGMC and KATO III cells (P < 0.05). However, NH₃ or HOCl did not induce the DNA double strand break as well as chromatin condensation in both cells. NH₂Cl, but not its precursors (NH₃ or HOCl), enhanced the levels of DNA injury, suggesting the possible involvement in the carcinogenesis of Hp-associated gastric mucosa.