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ARS Home » Northeast Area » Beltsville, Maryland (BHNRC) » Beltsville Human Nutrition Research Center » Diet, Genomics and Immunology Laboratory » Research » Research Project #426494

Research Project: Regulatory Mechanisms Induced by Health-Promoting Bioactive Food Components on Sex Steroid Hormone-Dependent Pathways, Cancer Cell-Stromal Cell Interactions, and the Gut Microbiome

Location: Diet, Genomics and Immunology Laboratory

2015 Annual Report

The overall goal of the project is to elucidate the molecular and cellular mechanisms that respond to selected food components to reduce the risk of chronic diseases such as cancer and obesity, with a focus on immune modulation in relation to obesity. A secondary aim is to further develop the utility of a porcine model to test the effect of probiotics and prebiotics on health maintenance through modulation of the gut microbiome and metabolome. Objective 1. Validate protective effects of bioactive food components such as glyceollins, indoles, and isothiocyanates on development of prostate cancer, and elucidate the regulation of sex steroid hormone-dependent pathways and cancer cell-stromal cell interactions as mechanisms of action by these bioactive food components. (NP107; C3, PS3B, C4, PS 4B) Objective 2. Study, in a swine model or other models as approriate, diet and gut microbiome interactions, focusing on the role of Lactobacillus, Bifidobacterium, and Bacteroides species, for the prevention of obesity and obesity-related metabolic syndrome. (NP107; C3, PS3B, C4, PS 4B)

For Objective 1, a complementary cell culture and tumor xenograft model will be used to test and identify efficacies of bioactive compounds from the diet and elucidate the mechanisms of how these bioactive food components act. The research is expected to identify mechanisms where food components alter biological processes such as proliferation, apoptosis, cell cycles, intracellular cell signaling, inflammation, metastasis, and post-transcriptional message regulation from both cell culture and whole animal studies. Genes involved in pathways mediated by the sex steroid hormones estrogen and androgen, orphan receptors, and cytokine-mediated pathways will be characterized. The conditions that modulate these pathways will include the use of plants with different phytochemical composition to delineate the role of specific compounds along with related food matrix effects. For Objective 2, a juvenile porcine animal model as a surrogate model for humans will be used to validate the effect of selected prebiotics and probiotics on the modulation of the host immune and metabolic responses to an obesogenic diet. The research will use a whole nutrigenomic approach where transcriptomics, metabolomic and metagenomic changes are integrated to identify biomarkers associated with health and disease that can be used as targets for nutritional interventions. Data generated from these studies is expected to reveal mechanisms of action of prebiotic and probiotic products added to the diet.

Progress Report
We continue our efforts to identify active components in the diet that may contribute to the protective effects of chronic diseases including cancer, obesity, and inflammation. We tested the hypothesis that the cruciferous-derived compound phenethylisothiocyanate (PEITC) may protect against prostate cancer development by using a tumor cell xenograft model of prostate cancer development. Nude mice were fed with or without PEITC for two weeks, then the animals were injected in their flanks with the human prostate cancer cell LNCaP. Animals were on the experimental diet for 8 weeks after tumor cell injection. Tumor size, food intake, and animal weight were measured. Tumor volumes in PEITC fed-animals were significantly lower than in control diet animals. These data support a cancer protective effect exerted by PEITC. Molecular analysis using a molecular marker for cell proliferation, androgen-dependent pathway (including IGFBP-3), angiogenesis, and immune responses (including IL-6) were determined in the tumor samples. Analysis of the results are in progress to elucidate mechanisms of action. We examined the effect of probiotics on the early onset of dietary-induced obesity and its consequences in a juvenile pig model. Replacement gilts have been designated for breeding and pig litters are expected in late winter to initiate the experiments. The intestinal microbiota is affected by changes in dietary patterns that influence selective bacterial groups and production of metabolites which contribute to define host health status. An experiment to follow the long-term changes in microbiota and metabolite composition of Ossabaw pigs fed a high calorie atherogenic diet or Heart Healthy diet has been started to determine the impact of the different diets on nutrient metabolism and on inflammatory biomarkers. A Cocoa powder rich in flavanols was identified and chemically analyzed for testing in a porcine model. In vivo testing has been completed to determine the effects of eating cocoa-derived flavanols on the composition of the gut microbiota and associated metabolic profiles. Twenty-four feeder pigs were supplemented with 0, 2.5, 10, or 20 grams of flavanol-enriched cocoa powder/day for 27 days. Chromatographic analysis demonstrated that O-methyl-(epi)catechin-glucuronide conjugates were detected in a dose-dependent manner in urine, serum, liver, and adipose tissue of pigs fed cocoa powder. Products of microbial metabolism were also detected in most of the tissues and bio-fluids. Pigs fed 20 grams of cocoa/day had significantly increased abundance of Lactobacillus species in the feces and distal colon contents. Moreover, consumption of cocoa powder reduced the gene expression of tumor necrosis factor (TNF)-a and toll-like receptors (TLR) -2, -4, and -9 in intestinal tissues of cocoa powder fed pigs. These results suggested a prebiotic effect of cocoa derived flavanols on beneficial bacteria as their relative abundance was increased and a modulating effect on inflammation was observed in the intestine. The higher dose of cocoa powder containing 20 grams of flavanol was further tested in a separate experiment in conjunction with feeding Lactobacillus rhamnosus (LGG) to evaluate a direct effect on LGG. Thirty-two pigs were fed a diet supplemented with cocoa powder, LGG, or a combination of cocoa powder and LGG to evaluate synergistic/symbiotic effects. The feeding component of the study was completed and analysis to determine the prebiotic effect of cocoa-derived flavanols is in progress.

1. New model for colonic digestion. Food-derived phytochemicals, many known for their health beneficial effects, often exist in conjugated forms containing sugar moieties such as glucose or rhamnose in foods. The uptake of these compounds requires colonic bacterial cleavage of sugar moieties. An assay was developed by ARS researchers in the Beltsville Human Nutrition Research Center, Beltsville, Maryland, that uses commercially available hesperidinase to mimic colonic digestion and to allow for accurate testing of the biological properties of food extracts by basic and translational scientist.

2. Method to purify biologically active soy-derived peptide. A novel and simplified method was developed to purify biologically-active soy-derived peptide by ARS researchers in the Beltsville Human Nutrition Research Center, Beltsville, Maryland. The procedure facilitates production of soy peptide lunasin and allows for further testing of its health promoting bioactivities by basic as well as translational scientists.

3. Lipid metabolism and role in inflammation. The effective function of the macrophage is dependent upon fatty acid oxidation and in particular triacylglycerol substrates that are scavenged from the blood by a receptor called CD36 and further metabolized by an enzyme called lysosomal acid lipase. ARS researchers in the Beltsville Human Nutrition Research Center, Beltsville, Maryland found that this process elevates oxidative phosphorylation, enhances spare respiratory capacity, and prolongs the survival and expression of genes that maintain the function of these cells. Inhibition of this metabolic process blocks the protective responses to worm infection. These findings demonstrate a critical role for lipid metabolism in the activation of certain immune and inflammatory cells, and is important to nutritionists that study how dietary components can improve health.

4. Health benefits can be induced by dietary induced modulation of microbiota. The pig is a suitable model to study the effect of dietary bioactives like cocoa derived flavanols in tissues not easily accessible in humans. ARS researchers in the Beltsville Human Nutrition Research Center, Beltsville, Maryland, found that consumption of cocoa-derived flavanols contributes to gut health by enhancing the abundance of Lactobacillus species and modulating intestinal immunity by ameliorating expression of inflammatory markers and Toll like receptors in a porcine model. These results provide basic and translation scientific information on molecular mechanisms underlying the health promoting effects of diet-derived flavanols.

Review Publications
Krishnan, H.B., Wang, T.T. 2015. An effective and simple procedure to isolate abundant quantities of biologically active chemopreventive lunasin-protease inhibitor concentrate (LPIC) from soybean. Food Chemistry. 177:120-126.
Huang, S.C., Everts, B., Ivanova, Y., O'Sullivan, D., Nascimento, M., Smith, A.M., Beatty, W., Love-Gregory, L., Lam, W.Y., O'Neil, C.M., Yan, C., Du, H., Abumrad, N.A., Urban Jr, J.F., Artyomov, M.N., Pearce, E.L., Pearce, E.J. 2014. Cell-intrinsic lysosomal lipolysis is essential for alternative activation of macrophages. Nature Immunology. 15(9):846-55. doi:10.1038/ni.2956.
Sun, J., Monagas, M., Jang, S., Molokin, A., Harnly, J.M., Urban, J.F., Aguilar, G., Chen, P. 2014. High fat diet leads to changes in metabolite patterns in pig plasma, fecal, and urine samples detected by a ultra-high performance liquid chromatography tandem with high resolution mass spectrometry metabolomic study. Food Chemistry. 15:173:171-178.
Yu, L., Huang, H., Yu, L.L., Wang, T.T. 2014. Utility of hesperidinase for food function research: enzymatic digestion of botanical extracts alters cellular antioxidant capacities and anti-inflammatory properties. Journal of Agricultural and Food Chemistry. 62(34):8640-8647.