Modulation of colon carcinogenesis by dietary ω-6/ω-3 fatty acid ratios : a chemopreventive strategy?
The aim of this study was to determine whether dietary fats constituting specific ω-6/ω-3 fatty acids (FA) ratio has chemopreventive modulating effects on the development of colon cancer. Western diets intake of saturated FA (SATS) and ω-6 polyunsaturated FA (PUFA) are very high relative to low ω-3 PUFA consumption. This high ω-6 and low ω-3 FA intake, resulting in a high ω-6/ω-3 FA ratio, appears to have a promoting effect on disease outcome, whilst increased ω-3 FA intake exhibiting anti-cancer effects. An animal cancer model was employed to evaluate the effects of dietary fat ratios on chemically induced carcinogenesis during cancer promotion. This was to determine whether the FA diets have a promoting or inhibitory effect on early neoplastic lesions by quantifying aberrant crypt foci (ACF) development and monitoring the crypt cells proliferative and apoptotic indices. The expressions of genes associated with changes in cells redox balance were also assessed. Common dietary fats were combined to produce the dietary fat ratios: sunflower oil (S), borage oil (B) and fish oil (F). Combinations of these oils generated the different ω-6/ω-3 FA ratios: SB (ω-6/ω-3: 38:1), SF (ω-6/ω-3: 13:1) and SBF (10:1). To represent the Western diet's high ω-6/ω-3 FA ratio profile, S (ω-6/ω-3: 501: 1) was used as a control, and canola oil and olive oil as additional reference. The dietary fats had no toxic effects on the liver and kidney based on serum clinical biochemical measurements. Diets containing borage oil (SB and SBF diets), canola and olive oil decreased (p<0.05) the crypt multiplicity of large (≥7 crypts/focus) ACF, exhibiting anti-cancer effects by decreasing (p<0.05) the proliferative activity of the rat colon crypts. Borage oil's protective effect resulted from the enhanced supply of C18:3ω-6 that has anti-inflammatory and anti-proliferative properties. The observed decrease (p<0.05) in apoptosis in the ACF was also facilitated by the up- and down-regulation of DNA repair and DNA replication associated genes, Xpa and Ercc2 by borage oil, respectively. Canola oil and olive had the largest inhibitory effect on suppressing crypt multiplicity by reducing (p<0.05) proliferation in the colon. Both oils effected the up-regulation (p<0.05) of the expression of several oxidative stress and anti-oxidant defence genes mediating the regulation of cell proliferation. The increased supply of C18:1ω-9 (canola and olive) and total polyphenolic content (olive) protected cells against oxidative stress induced apoptosis, which provided interesting interactive effects between FA and polyphenolic oil constituents that should be further elucidated. In contrast, the fish oil containing (SF diet) and the control sunflower (S diet) increased (p<0.05) the total ACF and colon crypt multiplicity (≥7 crypts/focus) when compared to the SB, SBF, olive oil and canola oil diets. An increased resistance to oxidative stress induced apoptosis appears to facilitate fish oil’s enhancing effect on crypt multiplicity despite the increased supply of LC ω-3 FA, which are prone to oxidation and leads to increased oxidative stress. This protective effect on crypt multiplicity and ACF development was mainly due to enhanced cellular antioxidant and DNA repair responses through the up-regulation (p<0.05) of Gpx4 and Nudt1, which favoured the increase (p<0.05) of crypt cells proliferation.The in vitro study demonstrated that oil ratio emulsions (S: ω-6/ω-3 = 249:1; SB: ω-6/ω-3 = 28:1; SF: ω-6/ω-3 = 12:1 and SBF: ω-6/ω-3 = 12:1) had differential effects on the survival indices of HT-29 and Caco-2 colon cancer cells. Contrary to the in vivo model, fish oil (SF and SBF emulsions) significantly (p<0.05) reduced the viability and proliferation of both cell lines, with the HT-29 cells showing greater sensitivity to the oil’s anti-proliferative effect. The HT-29 cells exposure to increased levels of C20:5ω-3 and C22:6ω-3 predisposes it to lipid peroxidation that increases the potential for cell removal via apoptosis. However, apoptotic effects were absent due to the HT-29 cells removal via necrosis as the cells energy status (ATP production) was significantly (p<0.05) depleted. Similar to the animal cancer model, borage oil (SB and SBF emulsions) had a reducing (p<0.05) effect on cell proliferation in both cell lines. However, as ATP was decreased (p<0.05), the S, SF and SBF emulsions resulted in an increased (p<0.05) apoptotic response in the Caco-2 cells in a dose dependent manner. This response resulted from the altered FA and lipid composition effected by the oil emulsions. Increased (p<0.05) incorporation of C20:5ω-3 and C22:6ω-3 in membrane phospholipid, phosphatidylethanolamine (PE), resulted in a significant decrease (p<0.05) in total SATS and MUFA content. A decrease (p<0.05) in membranes ω-6/ω-3 FA ratio was noted as well. This effect seems to selectively favour the induction of apoptosis by borage oil (SB and SBF). Similarly, an increase (p<0.05) in the PC/PE ratio by all oil emulsions, and a decrease (S and SB) and increase (SF and SBF) (p<0.05) in the chol/PL ratio appears to facilitate apoptosis too. A different threshold of the FA and lipid composition parameters elicits the inhibition of cell proliferation utilising lower oil emulsion concentrations. Therefore, the dietary supply of fats characterised by a defined low ω-6/ω-3 FA ratio can selectively modulate the growth indices of colon cancer. Specific oil ratio combinations by incorporating borage oil and fish oil hereby provide a selective strategy for chemoprevention in the colon, although underlying interactions and threshold effects of specific FA seems to prevail that should be further unravelled.