N-Methyl-N'-nitro-N-nitrosoguanidine (
MNNG), found in pickled foods and in chlorinated water, has been used to induce malignant transformation and
gastrointestinal cancer in rats. Helicobacter pylori (HP) is implicated in human
gastric cancer and possibly also in
esophageal cancer. These two agents - one chemical and the other biological - might act together to induce
esophageal cancer. In this study, human esophageal epithelial cells (HEECs) were divided into four groups: HP,
MNNG, HP + MNNG, and control. The HP-to-HEEC ratio was 100:1. Cells were exposed for 6 h and then passaged until malignant transformation. HEEC at early, intermediate, and late stages of malignant transformation were used for proliferation, cell-cycle, and invasion assays. The alkaline comet assay was performed and expression of
proteins, including γ-H2AX and
PAXX, was studied by western blotting, to explore DNA damage and repair processes. Measurements of cell morphology, soft-
agar clone formation, and invasiveness, and a nude mouse xenograft model, were used to examine
malignancy. The effect of HP was stronger than that of
MNNG. The combination HP + MNNG exerted a stronger malignant transformation effect than either HP or
MNNG alone. Mechanisms of this combined
carcinogenesis may include promotion of cell proliferation, perturbation of the cell cycle, promotion of invasiveness,
DNA double-strand break induction, or
PAXX inhibition.