L‐Canavanine (CAV) is a higher plant nonprotein amino acid and a potent L‐arginine antimetabolite. CAV can inhibit the proliferation of tumor cells in vitro and in vivo, but little is known regarding the molecular mechanisms mediating these effects. We demonstrated that the treatment of human lung adenocarcinoma A549 cells with CAV caused growth inhibition; G1 phase arrest is accompanied by accumulation of an incompletely phosphorylated form of the retinoblastoma protein, whose phosphorylation is necessary for cell cycle progression from G1 to S phase. In addition, CAV induces the expression of p53 and subsequent expression of a cyclin‐dependent kinase inhibitor, p21/WAF1. The p53–dependent induction of p21/WAF1 and the following dephosphorylation of the retinoblastoma protein by CAV could account for the observed CAV‐mediated G1 phase arrest.

A p53-inducible gene, the growth arrest and DNA damage-inducible gene 45 (gadd45), is associated with cell growth inhibition, DNA damage response and DNA repair. Here we report that Delta12-prostaglandin J2 (Delta12-PGJ2), an in vivo metabolite of arachidonic acid, which inhibits cell proliferation, induces gadd45 mRNA in HeLa cells. Because the p53 protein in HeLa cells is inactivated by papilloma virus E6, this type of gadd45 induction appears to be p53-independent. The induction was dose-dependent, and the maximum induction was observed at a concentration of 7.5 microgram/ml. In a time course study, gadd45 mRNA was induced 3 h after the addition of 7.5 microgram/ml Delta12-PGJ2. To investigate the transcriptional mechanism of gadd45 mRNA induction, we cloned a human genomic DNA fragment containing the gadd45 promoter region, and investigated the effect of Delta12-PGJ2 on the gadd45 promoter activity. In HeLa cells, 7.5 microgram/ml Delta12-PGJ2 markedly stimulated the gadd45 gene promoter about 20-fold or more. On the other hand, Delta12-PGJ2 did not stimulate the promoter activity of a reporter plasmid containing only p53 binding sites in HeLa cells, indicating that the gadd45 promoter activation by Delta12-PGJ2 was not mediated by p53. These results suggest that one of the mechanisms of cell growth arrest by Delta12-PGJ2 is mediated through a p53-independent induction of gadd45.

Recent studies have indicated that monochloramine (NH2Cl), a reaction product of NH3 and hypochlorous acid, is involved in the pathogenesis of Helicobacter pylori-associated gastric mucosal damage, but how NH2Cl contributes to lesions is unclear. In the present study, the effects of NH2Cl on mucosal cell growth and the cell cycle were evaluated in vitro using a normal rat gastric mucosal cell line RGM-1. Cell viability was assessed by the Trypan Blue dye exclusion test and cell cycle patterns were determined by DNA labeling with propidium iodide and flow cytometric quantification. NH2Cl inhibited the growth of RGM-1 cells in a concentration-dependent manner. Exposure of cells to NH2Cl caused a time- and dose-dependent loss of G1-phase cells with accumulation of G2/M-phase cells, and produced a fraction of subdiploid cells with oligonucleosomal DNA degradation characteristic of apoptosis. NH2Cl-induced apoptosis was confirmed by fluorescent microscopy with Hoechst 33342 and propidium iodide. These results suggest that NH2Cl inhibits gastric mucosal cell growth and induces apoptosis in RGM-1 cells, events that may be important in gastric mucosal damage or atrophy induced by H. pylori infection.