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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

2016 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 on 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. Tissue analysis revealed a 41.6% decrease in tumor burden, as compared to the control group, with no observable toxic effects. Consistent with previous in vivo findings, PEITC had minimal effects on prostate-specific antigen (PSA), an androgen-responsive gene in tumors. Significant reduction of KI67 (11.4%), a marker of cell proliferation, was observed in the PEITC treatment group. Moreover, the reduced expression of an EGF-like module containing, mucin-like, hormone receptor-like sequence 1 (Emr1 or F4/80) (-38.8%), a macrophage marker was observed. A 49% increase in an insulin-like growth factor binding protein 3 (IGFBP3) expression, responsible for forming a complex with insulin-like growth factor (IGF) (a prostate cancer-associated proliferative hormone), was also significantly altered in tumors of mice supplemented with a 5 µmol PEITC/ g diet. In summary, dietary administration of PEITC in this xenograft model attenuated prostate cancer progression by reducing proliferation, macrophage infiltration, and inhibiting IGF-1-mediated pathways. The effect of dietary bioactive components (i.e. Polyphenols) on the modulation of a host immune response and gut function has been evaluated using pig fed diets supplemented with cocoa powder, which is a food rich in polyphenols, including flavanols, anthocyanidins, and threobromine. We investigated the effects of consuming Cocoa powder on changes in the composition of selected species of gut microbiota, the metabolomic profile of several biofluids and tissues, and the changes in gene expression on inflammatory markers in pig intestinal tissue. Three doses of Cocoa powder at comparable amounts as reported in humans were given to weaned pigs. We found that the consumption of cocoa powder modulated colonic microbial metabolites and increased the abundance of Lactobacillus species in feces and Bifidobacterium species in proximal colon contents. In addition, gene expression of the pro-inflammatory cytokine TNFa and TLR2, TLR4 and TLR9 was reduced in ileal Peyer Patches, Mesenteric lymph nodes, and proximal colon of pigs fed cocoa powder. The concentration of a few metabolites significantly changed in response to cocoa powder doses used. Notably O-methyl-epicathecin glucuronide conjugates were consistently found in serum, urine, and visceral adipose tissue. Taken together, these results suggest that consumption of physiologically relevant doses of cocoa in the diet can provide potential benefits to gut health. A follow-up experiment combining the highest doses of cocoa powder with probiotic Lactobacillus rhamnosus LGG has also been tested to validate the prebiotic effect of cocoa powder. Whole transcriptome analysis of blood and microbiome composition changes were followed after a seven-week dietary intervention in a two-factorial design experiment (Probiotic* Cocoa) with matching controls to identify biomarkers in response to dietary intervention with and without antigen challenge with parasite.

1. Identify novel biomarker for hyper-cholestrolaemia. Hyper-cholesterolaemia is a risk factor for CVD, which is a leading cause of death in industrialized societies. The biosynthetic pathways for cholesterol metabolism are well understood; however, the regulation of circulating cholesterol by diet is still not fully elucidated. Using the hamster model, ARS scientists at the Beltsville Human Nutrition Center, Beltsville, Maryland, additional molecular insights into the influence of circulating cholesterol on hepatic cholesterol metabolism pathways during hyper-cholesterolaemia was obtained. The liver cholesterol synthesis enzyme CYP51 was identified as a novel gene/biomarker responsive to circulating cholesterol, and may be developed as biomarker for assessment of metabolic effects resulted from hyper-cholestrolaemia in human.

2. Elucidate molecular mechanisms of processing-induced food contaminants fatty acid esters of 3-chloro-1, 2-propanediol (3-MCPD esters). ARS scientists at the Beltsville Human Nutrition Center, Beltsville, Maryland, investigated whether and how the JNK/p53 pathway may play a role in the nephrotoxic effect of 3-MCPD esters. Molecular and cellular analysis were conducted and provided results that JNK/p53 might play a critical role in the tubular cell apoptosis induced by MPE and other 3-MCPD fatty acid esters.

3. The consumption of Cocoa powder enhances gut health. ARS scientists at the Beltsville Human Nutrition Center, Beltsville, Maryland, found that feeding flavanol enriched cocoa powder to pigs enhanced the abundance of probiotic-type bacteria like Lactobacillus and Bifidobacterium species and modulated markers of localized intestinal immunity as well as other markers at the brain level. These results suggest that the consumption of physiologically relevant doses of cocoa in the diet can provide potential benefits to gut health in pigs and humans.

4. The consumption of Cocoa powder enhances anti-inflammatory activity. ARS scientists at the Beltsville Human Nutrition Center, Beltsville, Maryland, found that pigs fed a standard diet and given a daily measure of flavanol enriched cocoa powder expressed greater anti-inflammatory activity in whole blood cells and intestinal tissues. This may be related to increased abundance of beneficial Bifidobacterium bacterial species that increased in feces collected after a 4 week dietary intervention. These results demonstrate a link between cocoa powder in the diet and improved health benefits in the intestine and circulating blood cells.

Review Publications
Liu, M., Liu, J., Wu, Y., Gao, B., Wu, P., Shi, H., Sun, X., Huang, H., Wang, T.T., Yu, L. 2016. Preparation of five 3-MCPD fatty acid esters and the effects of their chemical structures on acute oral toxicity in Swiss mice. Journal of Food Chemistry. doi: 10.1002/jsfa.7805.
Liu, M., Huang, G., Wang, T.T., Sun, X., Yu, L. 2016. 3-MCPD 1-palmitate induced renal tubular cell apoptosis in vivo via JNK/p53 pathway. Toxicological Sciences. 151(1):181-92. doi: 10.1093/toxsci/lfw033.
Jang, S., Sun, J., Chen, P., Lakshman, S., Molokin, A., Harnly, J.M., Vinyard, B.T., Urban Jr, J.F., Davis, C.D., Solano Aguilar, G. 2016. Flavanol-enriched cocoa powder alters the intestinal microbiota, tissue and fluid metabolite profiles, and intestinal gene expression in pigs. Journal of Nutrition. 146(4):673-80. doi: 10.3945/jn.115.222968.
Wang, A., Yang, Z., Grinchuk, V., Smith, A.D., Qin, B., Lu, N., Wang, D., Wang, H., Ramalingam, T.R., Wynn, T.A., Urban Jr, J.F., Shea-Donohue, T., Zhao, A. 2015. IL-25 or IL-17E protects against high-fat diet-induced hepatic steatosis in mice dependent upon IL-13 activation of STAT6. Journal of Immunology. 195(10): 4771-4780. doi:10.4049/jimmunol.1500337.
Fricke, F.W., Song, Y., Wang, A., Smith, A.D., Grinchuk, V., Mongodin, E., Pei, C., Ma, B., Lu, N., Urban Jr, J.F., Shea-Donohue, T. 2015. Immunity-dependent reduction of segmented filamentous bacteria in mice infected with the helminthic parasite Nippostrongylus brasiliensis. BMC Microbiome. doi: 10.1186/S40168-015-0103-8.
Li, R.W., Li, W., Baldwin, R.L., Yu, P., Urban Jr, J.F. 2016. The effect of helminth infection on the microbial composition and structure of the caprine abomasal microbiome. Scientific Reports. 6:20606.
Huang, Y., Guo, L., Qiu, J., Chen, X., Hu-Li, J., Siebenlist, U., Williamson, P.R., Paul, W.E., Urban Jr, J.F., Paul, W.E. 2015. IL-25-responsive, lineage-negative, KLRG1(hi) cells are multipotential “inflammatory” type-2 innate lymphoid cells. Nature Immunology. 16(2):161-169. doi: 10.1038/ni.3078