2007 Annual Report 1a.Objectives (from AD-416) LAB:Nutrition, aging, immune function and inflammatory response in health and disease Determine the mechanisms of vitamin E-induced enhancement of T cell function in the aged. Determine the efficacy of vitamin E in reducing the pathogenesis of secondary bacterial infection following influenza infection. Determine the role of adipocytes and obesity in age-related immune and inflammatory dysregulation and pathologies associated. Determine the effect and mechanisms of food components and their interaction with age and genetic background on immune function and infectious diseases. LAB:Vascular biology Determine at molecular and cellular levels the mechanisms by which the antioxidant components of foods (vitamins E, carotenoids and polyphenols such as catechins) singly and interactively modulate development of atherosclerosis and angiogenesis. Determine the effect of components of foods-vitamin E, carotenoids, lipids and polyphenols such as catechins, as well as their synergistic interactions in preventing and in reducing the risk of atherosclerosis, specifically when they are incorporated into the diet of animal models early in life compared to middle and later ages. 1b.Approach (from AD-416) LAB:Nutrition, aging, immune function and inflammatory response in health and disease The mechanism of vitamin E-induced enhancement of T cell function will be determined by investigating the effect of age and vitamin E supplementation on T cell receptor induced immune synapse formation using confocal microscopy of single and two-colored stained naïve T cells from young and old mice supplemented with or without vitamins. The mechanism of E-induced enhancement of effective immune synapse formation will further be determined by investigating the effect of age and vitamin E on the association of critical signaling molecules with membrane lipid domains known as lipid rafts using Western blots of phosphorylated and non-phosphorylated forms, of existing proteins, and S-methionine metabolic labeling of newly synthesized proteins. The mechanism of E-induced changes in association with the above key signaling molecules with lipid rafts will be determined by investigating the effect of age and vitamin E on palmitoylation of LAT (linker for activation of T Cells) and Lck (a Src family kinase) in young and old naïve T cells using metabolic labeling. In addition, sphingolipid and cholesterol composition of lipid rafts will be determined using HPLC and tandem mass spectrometry. The effect of vitamin E on downstream signaling molecules will be determined by focusing on Ca2+ mobilization and NFAT, NFKB, and AP-1 activities. The efficacy of vitamin E in preventing secondary bacterial infection in aged mice will be tested by feeding young and old C57BL/6 mice with adequate (30 ppm) or high (500 ppm) levels of E and determining viral and bacterial titers as well as morbidity and mortality after primary influenza A or S. aureus infection, or following a secondary S. aureus infection subsequent to an influenza infection. Preliminary experiments will be conducted to determine the underlying mechanisms. LAB:Vascular Biology The effect of mixed tocopherols at the concentrations commonly found in human plasma and after vitamin E supplementation will be examined in a cell culture system for modulation of immune and endothelial cell interaction and inflammatory cytokines. In this system, the interaction of vitamin E with avenathramide , a flavonoid found in oats, will be also tested for anti-atherogenic and anti-inflammatory action. The molecular mechanism of green tea catechins' inhibition of angiogenesis will be tested in a cell culture system using Matrigel and 3D gel. This effect of green tea will be investigated in vivo using the Matrigel plug assay in young and aged mice. The effect of vitamin E supplementation started from early, middle, and late age on the inhibition of atherosclerosis will be determined in LDL receptor null mice, which will be fed medium or high fat/cholesterol diets. The potential inhibitory effect of dietary vitamin E on the inhibition of atherosclerosis will be further tested in mice models when vitamin E is combined with lycopene, a carotenoid found in tomato, and with epigallocatechin gallate, the major catechin of green tea. 4.Accomplishments LAB: NUTRITIONAL IMMUNOLOGY 1) Vitamin E Improves the Age-related Defect in T cells. T cell function, important to immune system function, decline with age. Vitamin E restores T cell function in both aged animals and humans. To determine the underlying mechanism(s), we evaluated the effect of vitamin E on molecular reactions in T cells. We demonstrate that aged mouse cells have less of the active form (phosphorylated form) of the linker for activation of T cells (LAT), a key regulator of gene expression in T cells. We also demonstrate that the lower LAT phosphorylation in old T cells is because of less expression of the phosphatase SHP-1, which is a negative regulator of LAT phosphorylation. Vitamin E supplementation restores LAT phosphorylation, and reduces the level of SHP-1. These results indicate that Vitamin E improved T cell function in the aged mice by increasing the activity of key molecules. These results move us toward understanding the underlying causes of age-related decrease in T cell function and its reversal by supplementing the diet with vitamin E. NP107 Human Nutrition Component 5- Health Promoting Properties of Plant and Animal Foods. 2) Aging increases inflammation in fatty tissue. Obesity is a leading risk factor for type 2 diabetes (T2D). Aging is associated with increase in T2D occurrence even though we see a much lower prevalence of obesity in the elderly. Low-grade inflammation in fatty or adipose tissue (AT) is believed to be the cause of the development of insulin resistance and T2D. Thus, we conducted a study to determine if inflammation increases with age in AT. Results show that visceral AT from old C57BL mice have significantly higher expression of genes for pro-inflammatory cytokines, and cyclooxygenase -2, a key regulatory enzyme in inflammation than those of young. We further showed that among the cells found in fat tissue, fat cells or adipocytes (AD), and not macrophages cause increased inflammation of the aged AT, suggesting that the age-associated increase in AT inflammation is distinguished from that seen in obesity, in which macrophages are the main contributor. Sphingolipid ceramide was higher in old compared to young AD and contributed to the higher expression of inflammatory products in AD. These results have significant implication for understanding the underlying causes of age-associated increase in T2D and developing dietary strategies to reduce it. NP107 Human Nutrition Component 6-Prevention of Obesity and Disease: Relationship between Diet, Genetics, and Lifestyle. 3) Low Serum zinc and pneumonia in nursing home elderly. Pneumonia is a leading cause of death among elderly. Zinc plays an important role in immune system function and has been shown to reduce the number of pneumonias and death from pneumonia in children. The effect of Zinc on incidence of pneumonia in the elderly, however, has not been studied. Previously we reported that 30% of nursing home residents in the Boston area had low serum or blood levels of zinc. The objective of this study was to determine if serum zinc concentrations in nursing home elderly are associated with incidence and duration of pneumonia, antibiotic use, and pneumonia-associated and overall death. This observational study was conducted in residents from 33 nursing homes in Boston, MA, who participated in a one-year vitamin E supplementation trial. Subjects with normal final serum zinc concentrations had lower pneumonia incidence, total antibiotic use (by almost 50%), and shorter duration of pneumonia and antibiotic use (by 3.9 and 2.6 days, respectively) relative to those with low zinc concentrations. Normal baseline serum zinc concentrations were associated with decreased all-cause mortality. The results from our current study suggest that elderly people with low serum zinc concentrations might benefit from zinc supplementation. Such a measure has the potential to reduce the number of episodes and duration of pneumonia. It also may affect the amount and duration of antibiotic use due to pneumonia as well as decrease the number of overall deaths in the elderly. NP107 Human Nutrition Component 5-Health Promoting Properties of Plant and Animal Foods. 4) White Button Mushroom improves Natural Killer Cell Activity. Mushrooms have been shown to possess anti-tumor, anti-viral, and anti-bacterial properties. These effects suggest that mushrooms have the ability to regulate immune cell functions. However, most studies evaluated the effect of exotic mushrooms on the immune system, and not the commonly eaten white button mushrooms (WBM), which represent 90% of mushrooms consumed in the US. Thus we determined the effect of feeding WBM powder for 10 weeks on the immune response of mice. Results showed that WBM supplementation dose-dependently enhanced the activity of immune cells involved in fight against tumors and viruses (natural killer cell) as well as production of proteins with anti-tumor and anti-viral activity activity, interferon-g (IFN-g) and TNF-a. These results suggest that increased intake of white button mushrooms, the major mushroom consumed in United States may promote immunity against tumors and viruses. NP107 Human Nutrition Component 5-Health Promoting Properties of Plant and Animal Foods. LAB: VASCULAR BIOLOGY 1)Early Vitamin E Supplementation may Prevent Atherosclerosis in Mice There has been no control study conducted to date to demonstrate the preventive role of vitamin E supplementation on atherosclerosis in middle age when supplementation is initiated at an early age. Therefore we hypothesize that “supplementation with vitamin E from an early age prevents or retards the development of atherosclerosis and the risk of CHD in individuals with either Western dietary habits (high fat, high cholesterol) or reduced intake of fat and cholesterol as suggested by the American Heart Association.” We found that vitamin E supplementation from an early age in LDL receptor deficient mice, which are genetically prone to atherosclerosis, significantly protected the mice from mortality and reduced atherosclerotic lesions during 18 months of age. However, when the diet of mice contained high fat and cholesterol (Western type diet), vitamin E supplementation was not effective to prevent the development of atherosclerosis. Therefore, individuals that are genetically prone to atherosclerosis may benefit from early age vitamin E supplementation if avoid foods containing high fat and cholesterol. NP107 Human Nutrition Component 5- Health Promoting Properties of Plant and Animal Foods. 2) Antioxidant combinations, Western Diet and effect on Atherosclerosis in Mice Combined dietary lycopene, epigallocatechin gallate (EGCG) and vitamin E do not prevent atherosclerosis in mice when Western type diet is consumed. Several strong antioxidants have been identified in fruits and vegetables, such as vitamin E, catechins in green tea, and lycopene in tomatoes. In combination, antioxidants may have synergistic effects in biological systems. Therefore, in ApoE deficient mice which are genetically prone to atherosclerosis, we tested combinations of these dietary antioxidants when mice were fed with a Western diet for two months. We found that single or combined dietary antioxidants are not effective to reduce atherogenicity of Western type diet, which contains high fat and cholesterol. NP107 Human Nutrition Component 5- Health Promoting Properties of Plant and Animal Foods. 5.Significant Activities that Support Special Target Populations None. 6.Technology Transfer Number of non-peer reviewed presentations and proceedings 32 Number of newspaper articles and other presentations for non-science audiences 8 Review Publications Nie, L., Wise, M.L., Peterson, D.B., Meydani, M. 2006. Mechanism by which avenanthramide-c, a polyphenol of oats, blocks cell cycle progression in vascular smooth muscle cells. Free Radicals in Biology and Medicine. 41(5):702-8. Freeman, V.L., Flanigan, R.C., Meydani, M. 2007. Prostatic Fatty Acids and Cancer Recurrence Following Radical Prostatectomy for Early-Stage Prostate Cancer. Cancer Causes and Control. 18: 211-218. Marko, M.G., Ahmed, T., Bunnell, S.C., Wu, D., Chung, H., Huber, B.T., Meydani, S. 2007. Age-associated decline in effective immune synapse formation of CD4+ T cells is reversed by vitamin E supplementation. Journal of Immunology. 178:1443-1449. Han, S., Adolfsson, O., Lee, C., Prolla, T.A., Ordovas, J.M., Meydani, S. 2006. Age and vitamin E-induced changes in Gene Expression Profiles of T cells. Journal of Immunology. 177(12): 6052-6061. Gay, R.T., Belisle, S., Beck, M.A., Meydani, S.N. 2006. An aged host promotes the evolution of an avirulent coxsackievirus into a virulent strain. Proceedings of the National Academy of Sciences. 103(37):13825-13830. Wu, D., Han, S., Meydani, M., Meydani, S.N. 2006. Effect of concomitant consumption of fish oil and vitamin E on T cell mediated function in the elderly: a randomized double-blind trial. Journal of American College of Nutrition. 25(4):300-306. Sempertegui, F., Estrella, B., Elmieh, N., Jordan, M., Ahmed, T., Rodriguez, A., Tucker, K.L., Hamer, D.H., Reeves, P.G., Meydani, S.N. 2006. Nutritional, immunological, and health status of the elderly population living in poor neighborhoods of quito, ecuador: a preliminary report. British Journal of Nutrition. 96:845-853. Meydani, S.N., Han S.N. 2006. Nutrient Regulation of the Immune Response: The Case of Vitamin E. In: Bowman B.A., Russell, R.M., editors. Present Knowledge in Nutrition. 9th edition, Volume II. Washington, DC: International Life Sciences Institute. p. 585-603. Han, S., Meydani, S. 2006. The impact of vitamin e on immune function and its clinical implications. Expert Review of Clinical Immunology. 2(4):561-567. Wu, D., Pae, M., Ren, Z., Guo, Z., Smith, D., Meydani, S. 2007. Dietary Supplementation with White Button Mushroom Enhances Natural Killer Cell Activity in C57BL/6 Mice. Journal of Nutrition. 137(6): 1472-1477.