No abstract

Death-associated protein kinase (DAPK) is a serine/threonine kinase that contributes to pro-apoptotic signaling on cytokine exposure. The role of DAPK in macrophage-associated tumor cell death is currently unknown. Recently, we suggested a new function for DAPK in the induction of apoptosis during the interaction between colorectal tumor cells and tumor-associated macrophages. Using a cell-culture model with conditioned supernatants of differentiated/activated macrophages (U937) and human HCT116 colorectal tumor cells, we replicated DAPK-associated tumor cell death; this model likely reflects the in vivo tumor setting. In this study, we show that tumor necrosis factor-alpha exposure under conditions of macrophage activation induced DAPK-dependent apoptosis in the colorectal tumor cell line HCT116. Simultaneously, early phosphorylation of p38 mitogen-activated protein kinase (phospho-p38) was observed. We identified the phospho-p38 mitogen-activated protein kinase as a novel interacting protein of DAPK in tumor necrosis factor-alpha-induced apoptosis. The general relevance of this interaction was verified in two colorectal cell lines without functional p53 (ie, HCT116 p53(-/-) and HT29 mutant) and in human colon cancer and ulcerative colitis tissues. Supernatants of freshly isolated human macrophages were also able to induce DAPK and phospho-p38. Our findings highlight the mechanisms that underlie DAPK regulation in tumor cell death evoked by immune cells.

Helicobacter pylori are responsible for the induction of chronic gastric inflammation progressing to atrophy, metaplasia, and gastric cancer. The overexpression of Cathepsin X/Z (Ctsz) in H. pylori-infected mucosa and gastric cancer is mediated predominantly by an augmented migration of ctsz−/−positive macrophages and the up-regulation of Ctsz in tumor epithelium. To explore the Ctsz-function in the context of chronic inflammation and the development of preneoplastic lesions, we used Ctsz-deficient mice in a H. pylori gastritis model. Ctsz −/− and wild-type (wt) mice were infected with H. pylori strain SS1. The mice were sacrificed at 24, 36, and 50 weeks post infection (wpi). The stomach was removed, and gastric strips were snap-frozen or embedded and stained with H&E. Tissue sections were scored for epithelial lesions and inflammation. Ki-67 and F4/80 immunostaining were used to measure epithelial cell proliferation and macrophage infiltration, respectively. The upregulation of compensating cathepsins and cytokines were confirmed by Western blotting and quantitative RT-PCR. SS1-infected wt and ctsz −/− mice showed strong inflammation, foveolar hyperplasia, atrophy, and cystically-dilated glands. However, at 50 wpi, ctsz −/− mice developed significantly more severe spasmolytic polypeptide-expressing metaplasia (SPEM), showed enhanced epithelial proliferation, and higher levels of infiltrating macrophages. Induction of cytokines was higher and significantly prolonged in ctsz −/− mice compared to wt. Ctsz deficiency supports H. pylori-dependent development of chronic gastritis up to metaplasia, indicating a protective, but not proteolytic, function of Ctsz in inflammatory gastric disease.