Location: Jean Mayer Human Nutrition Research Center On Aging
2020 Annual Report
Objectives
Vitamins and Carcinogenesis Lab
Objective 1: Define the cellular pathways by which obesity, obesigenic diets, and the intake of the 1-carbon nutrients modulate the risk of developing cancers of the colorectum and other common cancers in both animal models and human samples, and exploit these mechanistic insights in order to devise targeted means of mitigating cancer risk.
• Sub-objective 1A: Determine whether the pro-inflammatory/pro-carcinogenic NF'B pathway plays the predominant role in mediating the obesity-promoted increased risk of colorectal carcinogenesis.
• Sub-objective 1B: Determine whether supplemental levels of dietary vitamin B6 provide additional suppression of obesity-promoted tumorigenesis and colonic inflammation when combined with curcumin + salsalate, beyond that provided by the two latter agents alone.
Objective 2: Examine how modifications in the microbiome alter biochemical and molecular processes that lead to colorectal cancer, and explore how intentional manipulations of the microbiome, or its products, can be exploited for cancer prevention.
Objective 3: In both genetic and chemically-induced rodent models of colorectal carcinogenesis examine whether select alternative protein sources (e.g. insect-based foodstuffs) suppress pro-carcinogenic pathways and tumorigenesis compared to soy protein and other dietary sources of protein more common in the American diet.
Nutrition and Cancer Biology Lab
Objective 1: Investigate mechanistically the anti-inflammatory and anti-carcinogenic effect of phytochemical-rich whole food approaches, and purified phytochemicals as well as their derivatives, in preventing inflammation-promoted (e.g., induced by a high-sugar diet, diabetes, and aging) cancer development.
Objective 2: Determine the ability of phytochemical-rich whole foods and dietary phytochemicals to prevent cancer development in liver and colon by targeting multiple signaling pathways (e.g. membrane and nuclear receptors) and inter-organ crosstalk (among liver, pancreas, mesenteric adipose tissue, and gut microbiome).
Approach
Vitamins and Carcinogenesis Lab
We will identify novel strategies by which colorectal cancer (CRC), and other cancers that commonly afflict elderly Americans, can be prevented. Our aim is to lessen the risk that accompanies cancer-promoting features typifying the U.S. diet, such as its obesigenic character and emphasis on processed animal meat. Using a combination of in vitro experiments and animal models we identify biochemical and molecular pathways that mediate the effects of specific nutrients or dietary patterns on carcinogenesis. We then identify means of modulating those pathways to mitigate cancer risk. We will examine how the inflammatory state created by obesity and high-fat diets activates procancerous pathways in the colon. We are exploring the use of pharmacologic, nutritional, and microbial agents to block those pathways. The third objective is an exploratory aim, designed to generate preliminary data. We will examine whether substituting protein-rich powder derived from roasted crickets attenuates the enhanced risk of CRC that accompanies the habitual consumption of processed meats which are a prominent source of protein in the American diet. This strategy has the added value of promoting food sustainability. Our research will provide novel avenues for reducing the societal burden of common age-related cancers.
Nutrition and Cancer Biology Lab
We will conduct animal studies to investigate how one dietary phytochemical, xanthophyll beta-cryptoxanthin (BCX), inhibits metabolic syndrome, nonalcoholic fatty liver disease and liver cancer (hepatocellular carcinoma) development in the liver. Of particular interest is understanding how BCX prevents the development of hepatocellular carcinoma in rodents consuming a diet high in refined carbohydrates (HRCD). We will examine the protective effects of intact BCX, independent of its metabolites, regulating key cell signaling pathways in both young and old animals. We will examine multiple organs (liver, pancreas, adipose tissue, and gut) as well as how these organs communicate, while noting gender differences. Specifically, we will use genetically-altered carotenoid cleavage enzyme (beta-carotene 15,15’-oxygenase and beta-carotene 9’,10’-oxygenase) double knockout mice strains to determine whether HRCD-induced liver metabolic syndrome and tumorigenesis can be prevented by intact BCX itself or sweet red pepper extract (SRPE)-rich in BCX. We will treat mice (male and female) with a single injection of a hepatic carcinogen, diethylnitrosamine (DEN), followed by continued exposure to HRCD with or without BCX (or SRPE) intervention. We will examine the effects of dietary BCX intervention against fatty liver, inflammation, fibrosis, and in livers. We will investigate the protective effects of xanthophyll BCX against HRCD-promoted HCC in both young and old mice respectively. We will determine if the BCX protective action process a common mechnism or pathway, such as intestinal permeability/gap junction/adipose/liver axis, salvage pathway of NAD+ biosynthesis enzyme, and circadian transcription factors, and thereby reducing aging/metabolic syndrome-associated liver cancer development.
Progress Report
VITAMINS AND CARCINOGENESIS LAB: Last year the investigators reported they had completed a study in an animal model of obesity-promoted colon cancer that demonstrated that the combination of supplemental curcumin and vitamin B6 suppressed tumorigenesis by 60-80%, and was far superior to either agent alone. Over the course of the past year, the investigators examined the biochemical and molecular basis of this synergy and identified three cellular avenues through which curcumin and B6 might interact to produce this synergy. These observations are immediately relevant to Objective #1.
The investigators extended their studies of the gut bacterium, P. distasonis, which they have previously shown to suppress obesity-promoted colon carcinogenesis in mouse models. They first showed that the proteinaceous fraction of the organism’s cell wall was sufficient to produce the anti-inflammatory and anti-tumorigenic activities. They completed another animal study that demonstrates that the anti-tumorigenic effect also exists in non-obese animals, and that it acts, in part, by reducing intestinal permeability. These observations are immediately relevant to Objective #2.
The investigators worked with a rodent diet company to design mouse diets that could be used to explore whether a diet whose protein is sourced from cricket powder is less tumorigenic than one whose protein is sourced from processed pork, soy flour or milk solids. The diets were then formulated and the nutrient composition of the diets were analyzed by an independent firm to ensure that diet components that were potential confounding effect modifiers were normalized. This work is immediately relevant to Objective #3.
NUTRITION AND CANCER BIOLOGY LAB: Despite the consistent association between a higher intake of the provitamin A carotenoid beta-cryptoxanthin (BCX, abundant in sweet red pepper) and a lower risk of fatty liver diseases among obese people, potential mechanisms supporting BCX as a dietary protective agent are needed. In support of Objective 1 and 2, we have carried out three major animal studies.
In the first study, we examined whether sweet red pepper rich in BCX inhibits highly refined carbohydrate/sugar diet (HRCD)-induced metabolic syndrome and nonalcoholic fatty liver disease (NAFLD). We also tested if inhibition of HRCD-induced NAFLD by BCX concentrate from sweet red pepper extract (SRPE) is dependent or independent of carotenoid cleavage enzymes (beta-carotene-15, 15’-oxygenase (BCO1) and beta-carotene-9, 10’-oxygenase (BCO2)) in mice. BCO1-/-/BCO2-/- double knock out (DKO) mice and wild type (WT) mice were randomly fed either an HRCD or HRCD with BCX concentrate (10 mg/kg diet, equivalent to the on daily human consumption of 3-4 ounces of sweet red peppers) for 6 months. Results showed that BCX concentrate feeding significantly reduced fatty liver severity and total cholesterol levels, in both WT and DKO mice compared to their respective HRCD counterparts. Hepatic levels of BCX, but not vitamin A (retinol and retinyl palmitate), were significantly higher (33-fold) in the DKO mice than in the WT mice. In WT mice, BCX-mitigated steatosis was significantly associated with increased hepatic lipid ß-oxidation (PPARa, ACOX1) and cholesterol efflux (ABCG5), and with suppressed lipogenesis (ACC) in the mesenteric adipose tissue. In DKO mice, the protection of BCX was significantly (P < 0.05) associated with decreased lipogenesis (FAS, ACC1, and ACC2), cholesterol synthesis (HMGCoA-R) and increased cholesterol catabolism (CYP7A1). BCX feeding increased hepatic mRNA expressions of farnesoid X receptor and decreased expressions of inflammatory cytokines (IL-6 and IL-4) in the mesenteric adipose tissue, which were associated with significantly increased sirtuin 1 levels in the liver and mesenteric adipose tissue (P < 0.05). The present study provides compelling evidence that BCX prevents HRCD-induced NAFLD through different mechanisms, depending on the presence or absence of BCO1/BCO2.
In the second study, we determined whether BCX concentrate from SRPE could inhibit carcinogen diethylnitrosamine (DEN)-initiated, HRCD-promoted hepatocellular carcinoma (HCC) development, dependent or independent of BCO1/BCO2 activity. Two-week-old male wild-type (WT) and BCO1-/-/BCO2-/- double knock-out (DKO) mice were given a single intraperitoneal injection of DEN (25 mg/kg body weight) to initiate hepatic carcinogenesis. At six weeks of age, all animals were fed HRCD (66.5 % of energy from carbohydrate) with or without BCX for 6 months. Results showed that BCX feeding increased hepatic vitamin A levels in WT mice, but not in DKO mice that showed a significant accumulation of hepatic BCX. Compared to their respective HRCD littermates, both WT and DKO fed BCX had significantly lower HCC multiplicity (58-60%), average tumor size (21-24%), and total tumor volume (51-58%), and the steatosis scores. The chemopreventive effects of BCX were associated with increased p53 protein acetylation and gluconeogenesis markers (phosphoenolpyruvate carboxykinase, glucose 6-phosphatase) and decreased protein levels of a glycolysis marker (lactate dehydrogenase) and of the hypoxia-inducible factor-1a and its downstream targets, matrix metalloproteinase 2/9 in tumors. This study demonstrated that BCX concentrate feeding alleviates HRCD-promoted HCC progression by modulating the acetylation of p53, hypoxic tumor microenvironment and glucose metabolism, independent of BCO1 and BCO2.
In the third study, we investigated the role of SIRT1 deacetylase activity in DEN-initiated, HRCD-promoted HCC in mice and unraveled the underlying mechanisms. Male and female SIRT1 homozygous mice deleted the catalytic activity (sirt1Y/Y) from C57/BL6J background and respective wild type (WT) mice were injected intraperitoneally with a liver-specific carcinogen, diethylnitrosamine (DEN), and fed a high-refined carbohydrate diet (HRCD, 66.5 % calories from carbohydrate consisting of sucrose and maltodextrin) for 6 months. Results showed that in both male and female mice, there were no significant differences in body weight, liver weight between sirt1Y/Y and its corresponding WT mice. In female mice, 37.5 % of WT mice (3/8) developed tumors in the liver, but none of sirt1Y/Y mice developed tumors in the liver (0/6). In male mice, all WT (8/8) and sirt1Y/Y mice (6/6) developed liver tumors. However, male sirt1Y/Y mice exhibited significant decreased tumor numbers and tumor volume in the liver compared to the respective WT mice. The reduction in liver tumor number and tumor volume was associated with significant increased protein levels of acetylated-p53 and its downstream target, p27 in the livers of male sirt1Y/Y compared with WT mice. Moreover, male sirt1Y/Y mice showed significant decreased protein levels of cyclin D1 and anti-apoptotic marker Bcl-2, as well as fatty acid ß-oxidation markers (CPT1, UCP1, UCP3) as compared with its corresponding WT mice. The study suggests that SIRT1 is a potential tumor promoter in DEN-initiated, high-refined carbohydrate-diet-promoted HCC by reducing acetylated-p53, which can contribute to the reduction of fatty acid ß-oxidation. We are conducting further investigations to explore the potential mechanism for sex-specific difference on HCC development.
Accomplishments
1. VITAMINS AND CARCINOGENESIS LAB: Certain gut bacteria reduces risk of colon cancer. Colon cancer continues to be the third most common cause of cancer death in U.S. adults. Cost-effective and non-invasive means of prevention are still lacking. ARS-funded researchers in Boston, Massachusetts, built on studies of the gut bacterium, P. distasonis, which they had previously shown to suppress obesity-promoted colon cancer in mice. They found the anti-cancer effect is produced by suppressing inflammation in the colon, which reduces gut leak in mice.
2. NUTRITION AND CANCER BIOLOGY LAB: Sweet red pepper pigment reduces fatty liver cancer development. Excessive intake of refined starch and sugars may lead to nonalcoholic fatty liver disease and liver cancer development. ARS-funded researchers in Boston, Massachusetts, showed that feeding mice beta-cryptoxanthin, a pigment abundant in red sweet peppers, that naturally converts to vitamin A reduced the effects of highly-refined starch or sugars which promotes fatty liver and liver cancer development. The protective effects of beta-cryptoxanthin were increased tumor suppression and decreased breakdown of glucose in liver tumors. This study helps to explain the significant protective benefit of a high intake of sweet red pepper on fatty liver disease and liver cancer.
Review Publications
Lim, J., Wang, X. 2020. Mechanistic understanding of beta-cryptoxanthin and lycopene in cancer prevention in animal models. BBA Molecular and Cell Biology of Lipids. https://doi.org/10.1016/j.bbalip.2020.158652.
Mason, J.B., Booth, S.L. 2019. Vitamins, trace minerals, and other micronutrients. In: Goldman, L., Schafer, A.I., editors. Goldman-Cecil Medicine. 26th edition. Maryland Heights, MO: Elsevier. p.73-79.
Koh, G., Kane, A.V., Wu, X., Crott, J.W. 2020. Parabacteroides distasonis attenuates tumorigenesis, modulates inflammatory markers and promotes intestinal barrier integrity in azoxymethane-treated A/J mice. Carcinogenesis. https://doi.org/10.1093/carcin/bgaa018.
Mustra Rakic, J., Wang, X. 2020. Role of lycopene in smoke-promoted chronic obstructive pulmonary disease and lung carcinogenesis. Archives of Biochemistry and Biophysics. https://doi.org/10.1016/j.abb.2020.108439.
Lim, J., Liu, C., Hu, K., Smith, D.E., Wu, D., Lamon-Fava, S., Ausman, L.M., Wang, X. 2019. Xanthophyll beta-cryptoxanthin inhibits highly refined carbohydrate diet-promoted hepatocellular carcinoma progression in mice. Molecular Nutrition and Food Research. 64(3):e1900949. https://doi.org/10.1002/mnfr.201900949.
