Examining signaling mechanisms by which colonic pro-inflammatory cytokines modulate obesity-promoted colonic carcinogenesis

Colorectal cancer (CRC) is the third most common cancer and third most common cause of cancer deaths in the United States. Amongst the many risk factors for this disease is obesity: those with a BMI of 25-29.9 have a relative risk of 1.2 and 1.5 for developing CRC, while those with a BMI of 30 have a relative risk of 1.5 and 2.0 for females and males, respectively. Recent evidence suggests that the low-grade chronic inflammatory state that accompanies obesity can have multiple promotional effects on pro-carcinogenic cell signaling cascades, and thus may be an important avenue by which excess adiposity promotes the risk of CRC.;The intent of the project described in this thesis was to define some of the mechanistic links between inflammation and colon carcinogenesis in obese rodent models as well as in humans. I aimed to: 1) elucidate the associations between pro-inflammatory cytokines in the colon and pro-carcinogenic signaling pathways in obese individuals; 2) delineate the mechanistic roles of colonic pro-inflammatory IL-1beta on the activation of Akt, NFkB and Wnt; and 3) define differences in colonic pro-inflammatory cytokines and gene expression signatures in the colonic epithelium in diet-induced (DIO) and genetically-induced (GIO) obese mouse models.;To demonstrate the clinical relevance of these principles, and to begin understanding how obesity might generate biochemical inflammation in the human colon, we compared the concentrations of TNF-alpha, IL-1beta, IL-6 and IFNgamma in the colonic mucosa of 16 lean and 26 obese individuals. In a general linear model, colonic TNF-alpha (r=0.41; p=0.01) and IL-6 (r=0.41; p=0.01) concentrations increased incrementally with BMI. Among individuals with a BMI of >34 the mean colonic concentrations of TNF-alpha and IL-6 were two-fold greater than in lean subjects (p<0.03); those with milder obesity (30 To demonstrate the clinical relevance of these principles, and to begin understanding how obesity might generate biochemical inflammation in the human colon, we compared the concentrations of TNF-alpha, IL-1beta, IL-6 and IFNgamma in the colonic mucosa of 16 lean and 26 obese individuals. In a general linear model, colonic TNF-alpha (r=0.41; p=0.01) and IL-6 (r=0.41; p=0.01) concentrations increased incrementally with BMI. Among individuals with a BMI of >34 the mean colonic concentrations of TNF-alpha and IL-6 were two-fold greater than in lean subjects (p<0.03); those with milder obesity (30≤BMI≤33.9) had intermediate values. Regular use of NSAIDs diminished the concentration of colonic TNF-a and IL-6 by a constant amount (p<.05) at each BMI. RNA sequencing analysis identified 182 differentially expressed genes in the obese colons compared to lean. Pathway analysis indicated gene enrichment for biologic functions regulating cell cycle control, apoptosis and proliferation, as well as immune function. Further, the Wnt, NFeB, and ERK signaling cascades assumed central positions in the two gene expression networks that were most tightly linked to the altered gene signatures, and the direction of change in gene expression among the regulatory molecules surrounding these cascades was consistent with their activation.;To examine the role of IL-1beta in mediating the early biochemical and molecular events leading up to CRC, male C57BL/6 (WT) mice were randomized to either low-fat or high-fat diets, as were two groups of male mice lacking a functional IL-1 receptor (IL1RKO) resulting in lean and obese mice. WT obese mice displayed significantly elevated levels of IL-1beta and TNF-alpha in the colonic mucosa (p<0.05), an increase in active ?-catenin in isolated colonocytes compared to lean WT mice (p<0.05), and a significant expansion of the proliferation zone in the colonic crypt (p<0.05); in contrast, in the absence of IL-1 signaling obesity had no effect on colonic IL-1beta, TNF-alpha, proliferation or Wnt activation.;Lastly, to better understand whether obesity-induced inflammation and elevation in pro-carcinogenic signaling pathways is due to high-fat consumption or excess adiposity, we used a diet-induced (HF) mouse model of obesity in Apc1638 mice and compared them to Apc1638Db/Db mice lacking the leptin receptor. Intestinal tumors were increased in both diet- and genetically-induced obesity, but we found no difference in colonic or small intestinal cytokines in HF or DbDb mice. A transcriptome analysis identified 266 and 584 genes differentially expressed in the HF and DbDb compared to lean, respectively. Pathway analysis indicated that the genes altered in both obese models were related to immune function and cellular proliferation and cancer. Furthermore, the top-ranking networks of interacting genes identified in both gene sets centered around Protein kinase B (Akt).;In conclusion, observations from this thesis indicate that even moderate degrees of adiposity result in elevations in colonic cytokines and cytokinetic and molecular changes relevant for the development of colonic carcinogenesis. Furthermore, it is important to emphasize that obesity produces chronic exposure to this inflammatory environment, which over time would likely magnify its impact on the colonic epithelium. These findings suggest that strategies to attenuate the effects of cytokine signaling, in particular the impact of IL-1, may be an effective mechanism to alleviate the burden of obesity-promoted CRC.