1 Department of Systems Biology, Technical University of Denmark2 Center for Biological Sequence Analysis, Department of Systems Biology, Technical University of Denmark3 Department of Management Engineering, Technical University of Denmark4 Systems Metabolic Lipidology, Department of Biotechnology and Biomedicine, Technical University of Denmark5 Disease Systems Immunology, Department of Biotechnology and Biomedicine, Technical University of Denmark
The still growing obesity epidemic is a major risk for our society, as it is associated with the development of the so called metabolic syndrome, which is a clinical diagnosis correlated to development of metabolic disorders. Lack of physical activity, excess energy intake, and nutritional factors e.g. fatty acid composition of the diet, are important factors with regard to development of metabolic syndrome. There is a controversy between the fact that several studies has shown that intake of saturated fatty acids are strongly correlated to the development of metabolic related diseases, such as cardiovascular diseases and type 2 diabetes, and against the fact that other studies has shown that intake of dairy fat, which has high saturated fatty acid content, correlates negatively with risk factors. Hence, it has even been suggested that dairy fat might have beneficial impacts in relation to metabolic disorders. Dairy fat is the most complex type of fat occurring in the nature, with more than 400 identified fatty acids. Several of these fatty acids that occur in low amounts have been suggested to have beneficial properties with regard to metabolic disorders. The concentrations of certain of these minor fatty acids are raised in dairy fat along with the amount of green plant material intake of the cattle. Phytanic acid is one of these minor fatty acids, due to agonist activities for nuclear receptors with central roles in among others the lipid and glucose metabolism. To determine the effects of both dairy fat in general and phytanic acid on metabolic parameters, we performed several studies. First, we investigated effects on hepatic lipid metabolism, glucose homeostasis, and circulating metabolic markers, of high fat diets based on butter from high- or low-yield production, a diet based on high oleic acid sunflower oil, and a diet based on grape-seed oil with high amount of linoleic acid, in diet induced obese mice. Second, we investigated phytanic acid effects on similar parameters in obese mice, both as dose response in butter based diets, and in grape-seed oil based diets with and without addition of phytanic acid. Third, we investigated butter and phytanic acid effects on human T-cell polarization, both by in vitro incubation with phytanic acid, and by a 12 weeks intervention with intake of butter. Finally, we performed two human interventions, first one with intake of butter and cheese, and the second with intake of butter. In these studies we investigated whether it is possible to alter the human plasma concentration of phytanic acid due to dairy fat intake, and if butter from different feeding regimes, and production forms has different effects on metabolic parameters upon intake. Fat type intervention in mice Obesity was induced in mice, by addition of sucrose to the drinking water, and giving high fat diets, based on butter from either grazing or conventional fed cattle, high oleic acid (monounsaturated fatty acid) sunflower oil, or finally from grape-seed oil with high content of the n-6 poly unsaturated fatty acid linoleic acid, along with having a lean reference group. Oral glucose tolerance test was performed after 10 weeks intervention, and animals sacrificed two days later. Parameters relevant to glucose metabolism, and hepatic lipid metabolism e.g. lipid deposition, were measured, just as RT-qPCR were used to measure expression of genes relevant for lipid metabolism in the liver. Plasma lipids, adipokines, and a marker of inflammation were also measured. We found that the hyper caloric diet based on oleic acid had the most detrimental effects on metabolic parameters, of the tested fats, as it led to increased hepatic lipid deposition, and reduced glucose tolerance. The butter based diets had more unfavorable effects on concentration of blood lipids, observed as raised triacylglycerol and total cholesterol. Compared to the literature the results with regard to oleic acid are controversial, as the common advice is to substitute SFA by MUFA in the diet. Phytanic acid effects in mice Production of phytanic acid by organic synthesis, allowed us to investigate isolated effects of phytanic acid intake. Obesity were induced in similar manner as in the fat type intervention described above, with different amounts of phytanic acid ethyl-esters added to either butter or grape-seed oil based diets, to investigate the effects from phytanic acid intake, on parameters similar to those in the fat type intervention. We saw that PA intake have aggravating effect on glucose homeostasis in dosages of 1.0 % of total fat. We did se limited up regulation of PPARa and ACOX1 due to 1.0 % phytanic acid in butter. As we are the first to perform interventions with physiological realistic amounts of phytanic acid, which have been proposed to have protective effects due to its agonist activities for central nuclear receptors, our results most definitely, add to the knowledge of the field. Butter and phytanic acid effects in humans, and on T-cell polarization Two human dairy fat interventions was conducted, with healthy subjects divided into groups and given dairy fat (as butter and cheese) from cattle under different feeding regimes, resulting in among others difference in phytanic acid content. From the first intervention, we found that it is possible to alter the human plasma phytanic acid concentration due to four weeks dairy fat intervention. From the second intervention we found that butter from grazing cattle, which among others have increased phytanic acid content, increase plasma LDL cholesterol and insulin, compared to conventional butter. From a subpopulation of the second intervention, T-cells were isolated from blood before and after the intervention, to analyze the effect on T-cell polarization. Furthermore we performed an in vitro incubation of T-cells, from eight donors, with phytanic- and palmitic acid, to investigate if phytanic acid affects T-cell polarization as hypothesized. Phytanic acid was not found to change the T-cell polarization, neither in the incubation study nor due to the difference in concentrations in the butter intervention. We saw up regulation in mRNA expression of both IL-4 and IFN-gamma due to the butter intervention, when the groups were regarded as one. This was more pronounced for IL-4 than IFN-gamma, and we observed increase in the ratio IL-4: IFN-gamma due to the intervention. This is pointing towards a general effect towards Th2 polarization of human T-cells due to increased intake of butter. These results add to the understanding of potential phytanic acid and butter effects, on the immune system as similar studies have not been performed on T-cells before.