1. Trans-fat diet increases hepatic protein tyrosine nitration Gabriella DiOdoardo and Steven A. Bloomer Department of Biology, Penn State Abington METHODS -Experiments were performed at Rutgers University and were approved by the Rutgers University IACUC. -Duration of control and TFA diets were 22 weeks (young) or 57 weeks (old). - All diets had the same macronutrient composition ratios, the only change was a replacement of 2% the fat energy replaced with a partially hydrogenated oil, when the controls had none (1). -Hepatic nitrotyrosine was evaluated by immunoblotting. ABSTRACT Aging is associated with hepatic oxidative damage, which is due, in part, to chronic exposure to exogenous stressors. One environmental component that affects overall physiology is diet composition, especially the amount and type of fats in the diet. In recent years, the consumption of trans-fatty acids (TFA) has increased, due to an increased availability of processed foods. In aged organisms, a diet containing TFA has negative effects on blood cholesterol. Since the liver is a major regulator of cholesterol, it is likely that TFA have a direct impact on the liver. However, it is unknown whether aging affects the hepatic response to TFA. Therefore, a diet containing 2% TFA was utilized to evaluate the combined effects of aging and TFA on hepatic oxidative stress. Nitrotyrosine was used as a marker of oxidative/nitrosative stress, and the amount of hepatic nitrotyrosine was determined via immunoblot and densitometry. While there was no significant effect of the TFA diet in the young groups, the TFA diet increased nitrotyrosine by 36% (p<0.05) compared to the control diet in the old groups. Our results suggest that aging impairs the ability to repair TFA-induced hepatic injury. Based on these results, we recommend decreasing the amount of TFA in the diet, especially in aged populations. INTRODUCTION - In the Western diet, trans- fatty acids (TFA) are a prevalent part of the diet, with average intake making up 2-3% of the total energy intake. (1) - TFA can cause fatty liver disease and myocardial infarction; both of these conditions are associated with oxidative stress. (1) - Oxidative stress is a condition of excess free radicals, which are molecules with unpaired electrons. - These molecules can cause damage to cellular macromolecules. - Aging is also associated with oxidative stress. - Research Question: Does a trans fat diet further increase oxidative damage with aging? - Hypothesis: TFA diet will increase Nitrotyrosine (Nty) abundance. SIGNIFICANCE - Because the trans fat diet augmented cellular injury in the old but not in the young group, this can indicate that the young are better at repairing the oxidative damage induced with TFA diet. FUTURE DIRECTIONS - Evaluate inducible nitric oxide synthase (iNOS) protein, which produces NO, and is induced by reactive oxidative species. - An increase in iNOS would further support our result of increased NTy abundance. - Determine protein amounts of superoxide dismutase (SOD). Because TFA diet leads to an increase in the superoxide radical(__), it is likely that TFA will increase SOD to compensate for the increased production of superoxide. RESULTS - Aging resulted in a trend towards an increase in hepatic nitrotyrosine. - In the young group, TFA did not cause an increase in Nty, whereas Nty was elevated in the old group on the TFA diet. FIGURE 2 : Effect of trans-fat diet on hepatic nitrotyrosine in the young and old groups. (A) Top panel: Representative immunoblot for nitrotyrosine. Bottom panel: Ponceau-stained membrane demonstrating equal protein loading and transfer. (B) Quantification of nitrotyrosine abundance normalized to the Ponceau stain. REFERENCES 1.Liou et al., 2013, Moderate Compared to Low Dietary Intake of trans- fatty Acids Impairs Stength of Old and Aerobic Capacity of Young SAMP8 Mice in Both Sexes. Lipids 48; (11) 1135-1143. 2.Craig-Schmidt, 2006, World –wide consumption of trans fatty acids. Atheroscler Suppl 7: 1-4. 3.Dhibi et al., 2011, The intake of high fat diet with different trans fatty acid levels differentially induces oxidative stress and non alcoholic fatty liver disease (NAFLD) in rats. Nutr Metab 8 (65). ACKNOWLEDGMENTS -Dr. Gregory Henderson at Newomics (formerly Rutgers University) -The ACURA program at Abington College -Faculty Development Grant to SAB Young Control Diet Young TFA Diet Old Control Diet Old TFA Diet Nitrotyrosine Abundance William A. Pryor et al. Am J Physiol Regul Integr Comp Physiol 2006;291:R491-R511 NO ● + O 2 ● - → OONO - + CO 2 → ● NO 2 + CO 3 ● - Trans-fatty acids Cis-fatty acids Partial hydrogenation From: “Fat and why it matters,” Indiana University http://www.indiana.edu/~oso/Fat/trans.html 250 kDa 100 kDa 250 kDa 100 kDa NTy Ponceau Partial hydrogenation p < 0.05