Effect of genetic deletion or pharmacological antagonism of tumor necrosis factor alpha on colitis-associated carcinogenesis in mice.

BACKGROUND: Intestinal inflammation in inflammatory bowel diseases is driven by abnormal levels of proinflammatory cytokines, where tumor necrosis factor (TNF)-α seems to be particularly important. Chronic inflammatory signaling in the colon increases the risk of colorectal cancer, so we sought to evaluate the role of TNF-α in a mouse model of this condition. METHODS: TNF mice were treated with azoxymethane/dextran sulfate sodium to induce inflammation and tumorigenesis. Etanercept was used to produce pharmacological ablation of TNF-α in wild-type mice. Subsequent activation of procarcinogenic transcription factor NF-κB and relevant proinflammatory cytokines of the TNF superfamily were measured through immunohistochemistry and quantitative polymerase chain reaction methods. RESULTS: Results showed that the severity of colitis, as assessed by mortality, histological scoring, and cytokine expression levels, was similar or slightly higher in mice lacking TNF-α than in control mice. Activation levels of NF-κB were not influenced by the presence of TNF-α. We also observed upregulated expression of TNF family member TNF-ß, TNF receptors 1 and 2 and a variety of other proinflammatory factors in colitis-associated tumors of TNF mice, compared with levels in tumors of control mice. Neither genetic ablation of TNF-α nor pharmacological inhibition of the TNF family using etanercept reduced tumor number. CONCLUSIONS: Our results reveal a redundant role for TNF-α in a mouse model of colitis-associated tumorigenesis, indicating a high degree of redundancy in proinflammatory cytokine networks in this model.

Autophagosomal IkappaB alpha degradation plays a role in the long term control of tumor necrosis factor-alpha-induced nuclear factor-kappaB (NF-kappaB) activity.

Transcription factor NF-κB is persistently activated in many chronic inflammatory diseases and cancers. The short term regulation of NF-κB is well understood, but little is known about the mechanisms of its long term activation. We studied the effect of a single application of TNF-α on NF-κB activity for up to 48 h in intestinal epithelial cells. Results show that NF-κB remained persistently activated up to 48 h after TNF-α and that the long term activation of NF-κB was accompanied by a biphasic degradation of IκBα. The first phase of IκBα degradation was proteasome-dependent, but the second was not. Further investigation showed that TNF-α stimulated formation of autophagosomes in intestinal epithelial cells and that IκBα co-localized with autophagosomal vesicles. Pharmacological or genetic blockade of autophagosome formation or the inhibition of lysosomal proteases decreased TNF-α-induced degradation of IκBα and lowered NF-κB target gene expression. Together, these findings indicate a role of autophagy in the control of long term NF-κB activity. Because abnormalities in autophagy have been linked to ineffective innate immunity, we propose that alterations in NF-κB may mediate this effect.

Upregulation of DNA methyltransferase-mediated gene silencing, anchorage-independent growth, and migration of colon cancer cells by interleukin-6.

Inflammatory bowel disease is characterized by chronic inflammation which predisposes to colorectal cancer. The mechanisms by which inflammation promotes tumorigenesis are not fully known. We aimed to investigate the links between colonic inflammation and tumorigenesis via epigenetic gene silencing. Colon cancer specimens were assessed for the expression of DNA methyltransferase-1 (DNMT-1) using immunohistochemistry. Colorectal carcinoma cell lines were assessed for DNMT1 expression, methylcytosine content, promoter methylation, gene expression, and tumorigenesis in response to interleukin (IL)-6. DNMT1 was expressed at higher levels in both the peritumoral stroma and tumor in inflammatory bowel disease-associated cancers compared with sporadic colon cancers. IL-6 treatment of colon cancer cells resulted in an increase in DNMT1 expression, independent of de novo gene expression. IL-6 increased the methylation of promoter regions of genes associated with tumor suppression, adhesion, and apoptosis resistance. Expression of a subset of these genes was downregulated by IL-6, an effect that was prevented by preincubation with 5-azadeoxycytidine, a DNMT1 inhibitor. Anchorage-independent growth and migration of colon cancer cells was also increased by IL-6 in a 5-azadeoxycytidine-sensitive manner. Our results indicate that DNMT-mediated gene silencing may play a role in inflammation-associated colon tumorigenesis.