Diet in the promotion or prevention of colorectal cancer

Colorectal cancer incidence is highly associated with one’s lifestyle, such as lack of physical activity, which leads to obesity, smoking habits, and most importantly, diet. Diet is an important risk factor for colon carcinogenesis and several studies have shown that high red meat and saturated fat intake increases the incidence of this disease significantly. On the other hand, a healthy balanced diet with intake of fruits and vegetables can decrease the risk of this disease. Dietary strategies for colon cancer chemoprevention, and even during treatment, are needed to help reduce the incidence of colorectal cancer. The aim of this work was to investigate dietary compounds that can be used in these dietary strategies for colon cancer prevention. Also, we initiated the development of new models to be used for compound screening processes. Initially, we focused on the effects of two bile acids, deoxycholic acid (DCA), with colon cancer promoting capacity, and ursodeoxycholic acid (UDCA), a chemopreventive compound, induced in Caco-2 cell line. We found that DCA in fact increase DNA damage and apoptosis in Caco-2 cell line. Also, activation of MAPK/ERK and PI3K/AKT pathways was also observed. UDCA did not induce DNA damage, but did induce the same act ivation in the signaling pathways. So, it seems that DCA increase cell turnover by increasing apoptosis and also cell proliferation in the remaining cells. UDCA only increase cell proliferation. When UDCA was administered as a pretreatment before DCA treatment, apoptosis was increased and this increase was accompanied by a constant activation of the JNK signaling pathway. Also, pretreatment with UDCA significantly decreased expression of the repair proteins MGMT and MLH1. One of the aims of this work was to develop an in vitro model of the in vivo azoxymethane (AOM)-induced colon cancer model. With the conditions tested we were able to induce a slight increase in cell proliferation in Caco-2 cells. This increase of cell proliferation could possibly be explained by the activation of the MAPK/ERK pathway, which was also activated with AOM treatment. Although we observed this increase in cell proliferation, we found no induction of O6-methylguanine lesions by our CoMeth assay or DNA damage observed by the comet assay. In the in vivo assays, the potential of an herbal tea, sage, and two isolated compounds found in foods from our diet, ursolic acid (UA) and EGCG, were evaluated for their chemopreventive effects against colorectal cancer. In the first study, sage tea was given to Fischer 344 rats before or after AOM treatment. Sage tea was able to reduce the number of pre-neoplastic lesions when given before AOM treatment, demonstrating chemopreventive potential. This reduction of pre-neoplastic lesions was accompanied by a reduction of the number of proliferating cells in colon crypts, as seen by Ki67 marker. Also, it conferred protection against DNA damage induced by AOM and by H2O2 ex vivo in colonocytes and lymphocytes. In the second study, UA and EGCG were added to the diet of healthy Fischer 344 rats. The potential of these compounds to protect against DNA damage was assessed. We found that both compounds protected against endogenous DNA damage in colonocytes and lymphocytes. The effects of the two compounds on protection against alkylating DNA damage induced ex vivo was also evaluated. UA and EGCG conferred protection against this type of damage in colonocytes, but not in lymphocytes. Finally, it has been shown that epigenetics has an important role in colon carcinogenesis, so we tried to develop a new, simple method to evaluate demethylating agents. We used the CoMeth assay developed in our group in a MMR-deficient cell line, in which one of the intervenients, MLH1, is epigenetically silenced by hypermethylation. Using 5-azacytidine, we were able to revert the hypermethylation and induce DNA damage in these cells. We further characterized the model, showing increase in apoptosis, and effects on reexpression of MLH1 protein levels by western blot. We also tested a few natural compounds with the model and found that EGCG, which is well-known that it has demethylating ability, induced similar levels of DNA damage as 5-azacitidine, suggesting that the model is functional. In conclusion, this work demonstrated the potential of sage tea as a chemopreventive agent and UA and EGCG as compounds with interest for chemopreventive strategies. The AOM in vitro model needs to be improved, but the adapted CoMeth assay for demethylating compounds is functional. Altogether, but with some additional studies, these natural compounds could be considered as chemopreventive agents and have possible interest in dietary strategies for cancer prevention.

Name thesis:degree_name

Doutoramento em Biologia Molecular e Ambiental

Year dc:date.issued

2015

Author dc:creator

• Pedro, Dalila Fernanda Neto

Advisor dc:contributor.advisor

• Wilson, Cristina Pereira

Statement dc:rights

• openAccess

Language dc:language.iso

eng

Handle dc:identifier.uri

https://hdl.handle.net/1822/36622

OAI record oai:identifier

http://repositorium.sdum.uminho.pt/oai/request?verb=GetRecord&metadataPrefix=dim&identifier=oai%3Arepositorium.uminho.pt%3A1822%2F36622