Progress 01/01/13 to 12/31/16
Outputs
Target Audience:General public, academic and scientific communities and health-care providers Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?
Nothing Reported
How have the results been disseminated to communities of interest?
Nothing Reported
What do you plan to do during the next reporting period to accomplish the goals?
Nothing Reported
Impacts
What was accomplished under these goals?
1. Understand the mechanism of action of synthetic PPARβ/δ ligands and their potential as a treatment for pancreatic cancer. Peroxisome proliferator-activated receptor β/δ (PPARβ/δ) is a member of the nuclear receptor superfamily and a ligand-activated transcription factor that is involved in the regulation of the inflammatory response via activation of anti-inflammatory target genes and ligand-induced disassociation with the transcriptional repressor B-cell lymphoma 6 (BCL6). Chronic pancreatitis is considered to be a significant etiological factor for pancreatic cancer development, and a better understanding of the underlying mechanisms of the transition between inflammation and carcinogenesis would help further elucidate chemopreventative options. The aim of this study was to determine the role of PPARβ/δ and BCL6 in human pancreatic cancer of ductal origin, as well as the therapeutic potential of PPARβ/δ agonist, GW501516. Over-expression of PPARβ/δ inhibited basal and TNFα-induced Nfkb luciferase activity. GW501516-activated PPARβ/δ suppressed TNFα-induced Nfkb reporter activity. RNAi knockdown of Pparb attenuated the GW501516 effect on Nfkb luciferase, while knockdown of Bcl6 enhanced TNFα-induced Nfkb activity. PPARβ/δ activation induced expression of several anti-inflammatory genes in a dose-dependent manner, and GW501516 inhibited Mcp1 promoter-driven luciferase in a BCL6-dependent manner. Several pro-inflammatory genes were suppressed in a BCL6-dependent manner. Conditioned media from GW501516-treated pancreatic cancer cells suppressed pro-inflammatory expression in THP-1 macrophages as well as reduced invasiveness across a basement membrane. These results demonstrate that PPARβ/δ and BCL6 regulate anti-inflammatory signaling in human pancreatic cancer cells by inhibiting NFκB and pro-inflammatory gene expression, and via induction of anti-inflammatory target genes. Activation of PPARβ/δ may be a useful target in pancreatic cancer therapeutics. 2. Determine how dietary bioactive molecules or pollutants affect the process of reverse cholesterol transport (RCT) and inflammation, important steps in atherosclerosis and cardiovascular disease. The nuclear receptor Farnesoid X Receptor (FXR) functions as a bile acid sensor that impacts the expression of genes involved in cholesterol, triglyceride and bile acid production and increases cholesterol efflux in macrophages. The purpose of these studies was to determine if the effect of FXR on cholesterol efflux is mediated by microRNAs (miRNAs), several of which affect cholesterol homeostasis in a variety of cell types. MicroRNA-708 expression was increased by FXR ligands in THP-1-derived macrophages and Huh7 cells, but its target genes and effects on cholesterol transport have not been explored. Transfection with miR-708-5p repressed CD38 mRNA and protein levels and attenuated expression of the pro-inflammatory genes NFκB and IL-1β in both cell types. Targeted knockdown of CD38 augmented Sirt-1 transcripts levels, reduced foam cell formation and enhanced cholesterol efflux. Our results suggest that miR-708-5p is an important FXR-regulated gene and its regulation of CD38 enhances reverse cholesterol transport. Thus, miR-708-5p represents a novel therapeutic target for the enhancement of reverse cholesterol transport, reduction of inflammation and treatment of atherosclerosis.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2016
Citation:
Vanden Heuvel, J. P., Bullenkamp, J., and Reproducibility Project: Cancer Biology Registered report: Systematic identification of genomic markers of drug sensitivity in cancer cells, Elife 5:e13620
- Type:
Journal Articles
Status:
Published
Year Published:
2016
Citation:
Hannon, D. B., Thompson, J. T., Khoo, C., Juturu, V. and Vanden Heuvel, J. P. (2016), Effects of cranberry extracts on gene expression in THP-1 cells. Food Sci Nutr. doi:10.1002/fsn3.374
- Type:
Journal Articles
Status:
Published
Year Published:
2016
Citation:
Hinds, Jr., T.D., Burns, K.A., Hosick, P.A., McBeth, L., Nestor-Kalinoski, A., Drummond, H.A., AlAmodi, A.A., Hankins, M.W., Vanden Heuvel, J.P., Stec, D.E. Biliverdin Reductase A Attenuates Hepatic Steatosis by Inhibition of glycogen synthase kinase beta Phosphorylation of Serine 73 of peroxisome proliferator-activated receptor-alpha J. Biol Chem.
- Type:
Journal Articles
Status:
Published
Year Published:
2016
Citation:
Smith, R.W., Coleman, J.D., Thompson, J.T., and Vanden Heuvel, J.P. (2016) Therapeutic potential of GW501516 and the role of Peroxisome Proliferator-Activated Receptor Beta/delta and B-Cell Lymphoma 6 in inflammatory signaling in human pancreatic cancer cells. Biochem Biophys Reports, In press.
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Progress 10/01/14 to 09/30/15
Outputs
Target Audience:General public, academic and scientific communities, and health care providers. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?Under this project, graduate students were trained. How have the results been disseminated to communities of interest?The information has been disseminated through the journal article listed under "Products." What do you plan to do during the next reporting period to accomplish the goals?The specific milestones for all the studies listed above pertain to timely publication and dissemination of information to the scientific community, as well as the public at-large.
Impacts
What was accomplished under these goals?
1. Understand mechanisms of action of synthetic PPARbeta/delta ligands and their potential as a treatment for pancreatic cancer: Previous studies have shown that PPARbeta/delta inhibit inflammatory signaling as well as metastasis of pancreatic cancer cells in cell culture. The expression of PPARbeta/delta, BCL6, and several of their target genes were examined in human pancreatic ductal adenocarcinoma (PDAC) relative to matched normal pancreas tissue, and in a mouse model that mimics the human progression of disease (Pdx1-Cre;LSL-KrasG12D). PPARbeta/delta mRNA was overexpressed in human tumor tissue compared to normal tissue, while BCL6 remained unaltered. Inflammatory genes that are controlled by the PPARbeta/delta-BCL6 pathway were increased in the tumor tissue, including the important BCL6 target genes and pancreatic cancer prognostic indicators MCP-1 and MMP9. Similar observations were made in the KrasG12D mice, where pancreas of mice carrying this constitutively active oncogene had higher expression of PPARbeta/delta mRNA, unaltered BCL6 expression, and increased MCP-1 and MMP9 mRNA. These in vivo associations were further examined in the Mia Paca-2 cell line. Forced PPARbeta/delta increased proliferation of pancreatic cancer cells, while this was ameliorated by GW501516. Over expression of PPARbeta/delta also increased basal and TNFα-induced MCP1 activity, which was diminished by the transfection of BCL6 or treatment with GW501516. Taken together, these data strongly support a sequestering of BCL6 by PPARbeta/delta that occurs during the progression of PDAC, when dysregulation results in overexpression of PPARbeta/delta. 2. Determine how dietary bioactive molecules or pollutants affect the process of reverse cholesterol transport (RCT) and inflammation, important steps in atherosclerosis, and cardiovascular disease (CVD): The nuclear receptor Farnesoid X Receptor (FXR) is a bile acid sensor that has gained considerable interest as a therapeutic target for the treatment of CVD. Agonists of FXR increase cholesterol efflux and impact gene expression in macrophage-derived foam cells (MDFC), liver, and intestine. The expression of genes involved in cholesterol, triglyceride (TG), and bile acid production, as well as the expression of microRNAs (miRs) is affected by FXR. MicroRNAs are non-coding transcripts that regulate the expression of key metabolic genes, although their role in FXR-dependent cholesterol efflux was not known previously. In parallel studies, we explored the miRs whose levels were affected by the FXR ligand GW4064 in MDFC and, utilizing a novel high-throughput screening approach, we examined the effects of various miRs on cholesterol efflux. Of the several miRs that were regulated by GW4064 and in turn influenced cholesterol efflux in vitro, miR-708 was chosen for further study. Whether the induction of miR-708 by GW4064 was causally related to enhanced cholesterol efflux was examined in several in vitro studies. Following miR-708 transfection and subsequent treatment with oxLDL and GW4064 in MDFC, there was a significant decrease in nuclear factor kappa B (NFκB)and interleukin 1 beta (Il-1β) gene expression,concomitant with an increase in cholesterol efflux. Ectopic expression of miR-708 repressed CD38 mRNA and protein levels in the in vitro models listed above. CD38 is a multifunctional enzyme that utilizes NAD+ in the synthesis of cyclic ADP-ribose, and is involved in immune function and calcium signaling. Targeted knockdown of CD38-augmented expression of the metabolism regulator gene, Sirtuin-1 (Sirt-1) and enhanced cholesterol efflux. Our results suggest miR-708 affects cholesterol efflux, at least in part via its regulation of CD38, and is a beneficial microRNA in regards to its role in atherosclerosis and the reverse cholesterol transport pathway.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2015
Citation:
Vanden Heuvel, J. P., Thompson, J. T., Albrecht, P., Mandetta, D., Kamerow, H., Ford, J. P. Differential nucleobase protection against 5-fluorouracil toxicity for squamous and columnar cells: Implication for tissue function and oncogenesis. Invest New Drugs. 2015 Oct;33(5):1003-11.
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Progress 10/01/13 to 09/30/14
Outputs
Target Audience: General public, academic and scientific communities and health-care providers. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided? There are two graduate students working on this project. How have the results been disseminated to communities of interest? The information has been disseminated through the manuscripts listed above. What do you plan to do during the next reporting period to accomplish the goals? The specific milestones for all the studies listed above pertain to timely publication and dissemination of information to the scientific community as well as the public at-large. The following as specific benchmarks to be achieved. 1). Three scientific publications per year in peer-reviewed journals. 2). Actively training undergraduate and PhD students with a goal of maintaining a program of approximately ten individuals. 3). Obtain extramural funding to sustain the previous milestones.
Impacts
What was accomplished under these goals?
Understand the mechanism of action of PPARβ/δ ligands, including omega-3 PUFAs, in the treatment of pancreatic cancer; The focus of the current studies on PPARb/d ligands is to better understand the etiology and treatment of pancreaticcancer. The nuclear receptor Peroxisome Proliferator-Activated Receptor β/δ (PPARb/d) regulates expression of genes involved in inflammation, metabolism, and cancer directly or in concert with the transcriptional repressor B-Cell Lymphoma 6 (BCL6) in the pancreas. Ligand-dependent activation of PPARb/d causes the dissociation of BCL6 from the complex, which in turn suppresses genes involved in inflammation and invasion. In this study, we have examined the mRNA expression of several cancer biomarkers, as well as PPARβ/δ and BCL6 target genes in human pancreatic tissue of matched normal and tumor biopsies, and the LSL-Kras(G12D); Pdx-1-Cre mouse model of pancreatic cancer. This mouse strain contains a latent point mutant allele of Kras2 in which expression leads to pancreatic tumors in heterozygous mice. Pancreatic PPARb/d expression was higher in tumors than in surrounding tissue in humans and was augmented in Kras(G12D) expressing mice compared to non-recombinant counterparts. There was no change in BCL6 expression. BCL6 target genes, MCP1 and NFκB1 involved in inflammation, and VCAM1 in cell adhesion showed significant increases in both human tumors and in pancreas from Kras(G12D)-expressing mice. MMP9, another BCL6 target responsible for regulation of cell migration and metastasis, also showed elevated expression from normal tissue in humans. Increased I expression of PPARb/d in tumors and in preneoplastic lesions may sequester the transcriptional repressor BCL6, and hence, play an important role in cancer development and progression, and suggest means for treating pancreatic cancer. Determine how dietary bioactive molecules or pollutants affect the process of reverse cholesterol transport (RCT) and inflammation, important steps in atherosclerosis and cardiovascular disease: Cholesterol metabolism and efflux from peripheral cells, such as macrophages and foam cells, is a critical part of reverse cholesterol transport (RCT), a metabolic process that has implications for atherosclerosis and cardiovascular diseases. There have been several conflicting studies that suggest environmental and occupational exposure to Perfluorooctanoate (PFOA) and Perfluorooctanesulfonate (PFOS) affects RCT and cholesterol metabolism. However, the data from animal studies and examination of potential mechanisms of action raise questions as to whether there is a direct causality of PFOA and PFOS exposure and hypercholesterolemia. In these studies, we have directly examined effects of PFOA and PFOS on cholesterol efflux and in genes involved in this process. PFOA and PFOS are agonists for nuclear receptors that increase cholesterol efflux, such as the peroxisome proliferator-activated receptors (PPARs). Importantly, in THP-1 differentiated into macrophage-derived foam cells, PFOA and PFOS increased cholesterol efflux to either HDL or apoA. Comprehensive gene expression analysis by microarray is being pursued to determine the key genes being regulated by these perfluorinated compounds in foam cells that impact RCT. These studies also will assist in the identification of genes that are sensitive to PFOA and PFOS exposure and that can be utilized in biomonitoring studies. Taken together, these studies suggest that PFOA and PFOS exposure affects cholesterol efflux, but not in a manner consistent with a negative health impact.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Vanden Heuvel, J. P. 2013. Comment on "Associations between PFOA, PFOS and changes in the expression of genes involved in cholesterol metabolism in humans" by Fletcher et al., Environment International 57-58:2-10.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Roussell, M. A., Hill, A. M., Gaugler, T. L., West, S. G., Ulbrecht, J. S., Vanden Heuvel, J. P., Gillies, P. J. and Kris-Etherton, P. M. 2014. Effects of a DASH-like diet containing lean beef on vascular health. J Hum Hypertens 28:600-605.
- Type:
Journal Articles
Status:
Published
Year Published:
2014
Citation:
Harris Jackson, K., West, S. G., Vanden Heuvel, J. P., Jonnalagadda, S. S., Ross, A. B., Hill, A. M., Grieger, J. A., Lemieux, S. K. and Kris-Etherton, P. M. 2014. Effects of whole and refined grains in a weight-loss diet on markers of metabolic syndrome in individuals with increased waist circumference: a randomized controlled-feeding trial. Am J Clin Nutr 100, 577-586.
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Progress 01/01/13 to 09/30/13
Outputs
Target Audience: General public, academic and scientific communities and health-care providers. Changes/Problems:
Nothing Reported
What opportunities for training and professional development has the project provided?
Nothing Reported
How have the results been disseminated to communities of interest? The information has been disseminated through the manuscripts and book chapter listed in published products section. What do you plan to do during the next reporting period to accomplish the goals? The specific milestones for all the studies listed above pertain to timely publication and dissemination of information to the scientific community as well as the public at-large. The following as specific benchmarks to be achieved: 1) Three scientific publications per year in peer-reviewed journals; 2) Actively training undergraduate and PhD students with a goal of maintaining a program of approximately ten individuals; 3) Obtain extramural funding to sustain the previous milestones.
Impacts
What was accomplished under these goals?
1. Understand the mechanism of action of Omega-3 PUFAs in the prevention of breast and pancreatic cancer; the focus will be on the nuclear receptors involved in regulation of key cancer prevention pathways as well as the Omega-3 PUFA-derived metabolites responsible for this response. Among individual components of fish oil, DHA and EPA and, in certain instances, the corresponding ethylesters were examined. In previous studies, the chemopreventive efficacy of dietary DHA was remarkable in suppressing MNU-induced mammary carcinogenesis in the rat; it was significantly more effective than EPA or the mixture of EPA + DHA. Although progress has been made in understanding the mechanisms that can account for the chemopreventive effects of n-3 FA, it remains largely undefined whether DHA or one of its metabolites govern these effects. We have shown that DHA activates PPARs and RXRs and that certain active DHA metabolites (ADMs) are more efficacious activators. However, while acknowledging the existence of multiple targets for DHA and ADMs, PPARγ and not the RXRs has emerged as a potential target for breast cancer protection and thus was examined in detail. Human breast cancer cells (MCF7) were treated with several concentrations of DHA or ADMs and cell growth and PPARg activity were assessed. The results showed that 4-OH-DHA and 4-OXO-DHA significantly reduced cell number more effectively than DHA. DHA, 4-OH and 4-OXO DHA were all efficacious PPARg activators with the ADM being more effective that the parent molecule. In a follow-up study several ADMs were compared for their PPARγ agonistic activity. The activity follows the order 4-OXO-DHA > 14-OH- DHA ~ 20-OH-DHA > 4-OH-DHA > 17-OH-DHA. The remarkable activity of 4-OXO-DHA stimulated a follow-up collaborative study to further examine its growth inhibitory activity in several human breast cancer cell lines (both luminal and basal breast cancer cells). The results further demonstrate that DHA and 4-OXO-DHA are inhibitors of cell growth; 4-OXO-DHA was more effective in basal cells (MDA-MB-231, MDA-MB-468) and the HER-2/neu positive SKBR-3 than luminal type cells (BT-474, MCF-7). In addition, DHA inhibited cell growth in a dose-dependent manner in rat mammary adenocarcinoma LA7 cells; compared to human breast cancer cells, LA7 rat adenocarcinoma cells are highly sensitive to DHA. Taken together, these studies have illustrated that metabolites of DHA may be key mediators of the prevention of breast cancer. 2. Determine how omega-3 PUFAs and other dietary bioactive molecules, affect the process of reverse cholesterol transport (RCT) and inflammation, important steps in the prevention of atherosclerosis and cardiovascular disease. Atherosclerosis is the major cause of morbidity and mortality in the United States, with coronary heart disease and stroke being its two most common expressions. The reverse cholesterol transport system (RCT) involves cholesterol efflux, an important marker and mediator of macrophage cholesterol homeostasis. Cholesterol efflux from peripheral tissues and cells, such as macrophage-derived foam cells (MDFC) in atherosclerotic plaque, is an initial and critical step in RCT. Previous studies from our lab have shown that omega-3 fatty acids, in particular ALA, enhanced cholesterol efflux from MDFC, at least in part through an FXR-dependent process. Through collaborative clinical nutrition studies, we have shown that diets high in ALA and other bioactive molecules can enhance the efflux of cholesterol from MDFC to serum, ex vivo. We are currently exploring the mechanism of this protective effect and are focusing on the role of microRNAs (miRs). MicroRNAs are 21–22-nucleotide (nt), non-coding small RNAs produced within the genomic loci of cells that regulate the degradation and translation of targeted mRNAs. Using a high-throughput method developed on our lab, we have shown several miRs are able to affect FXR-dependent cholesterol efflux from MDFC. By understanding the mechanism by which these miRs affect RCT, and how they are affected by omega-3 PUFAs and FXR agonists, we can better develop diagnostic and intervention strategies for atherosclerosis.
Publications
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Coleman J. D., J. T. Thompson, R. W. Smith, B. Prokopczyk, and J. P. Vanden Heuvel. 2013. Role of Peroxisome Proliferator-Activated Receptor Beta/Delta and B-Cell Lymphoma-6 in Regulation of Genes Involved in Metastasis and Migration in Pancreatic Cancer Cells. PPAR Res 2013:121956.
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Paul K. B., J. T. Thompson, S. O. Simmons, J. P. Vanden Heuvel, and K. M. Crofton. 2013. Evidence for triclosan-induced activation of human and rodent xenobiotic nuclear receptors. Toxicol In Vitro 27:2049-2060.
- Type:
Journal Articles
Status:
Published
Year Published:
2013
Citation:
Eser P. O., J. P. Vanden Heuvel, J. Araujo, and J. T. Thompson. 2013. Marine-and plant-derived ?-3 fatty acids differentially regulate prostate cancer cell proliferation. Molecular and Clinical Oncology 1:444-452.
- Type:
Book Chapters
Status:
Awaiting Publication
Year Published:
2014
Citation:
Gillies P. J., and J. P. Vanden Heuvel. 2014. Nutrigenomic Approaches to Unraveling the Physiological Effects of Complex Foods, p. 105-115. In Nutrigenomics and Nutrigenetics in Functional Foods and Personalized Nutrition. CRC Press.
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