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Publications (4)4.2 Total impact

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    ABSTRACT: Nuclear factor-erythroid 2-related factor 2 (Nrf2) plays a pivotal role in maintaining cellular redox homeostasis and eliminating reactive toxic species. Nrf2 is epigenetically suppressed due to CpG hypermethylation in prostate tumors from the transgenic adenocarcinoma of the mouse prostate (TRAMP) model. We previously showed that dietary feeding of a γ-tocopherol-rich mixture of tocopherols (γ-TmT) suppressed prostate tumorigenesis in TRAMP mice associated with higher Nrf2 protein expression. We hypothesized that γ-TmT may maintain Nrf2 through epigenetic inhibition of promoter CpG methylation. In this study, 8-wk-old male TRAMP mice were fed 0.1% γ-TmT or a control diet for 16 wk. The methylation in the Nrf2 promoter was inhibited in the prostate of the γ-TmT group compared with the control group. Protein expressions of DNA methyltransferase (DNMT), including DNMT1, DNMT3A, and DNMT3B, were lower in the prostate of the γ-TmT group than in the controls. TRAMP-C1 cells were treated with 30 μmol/L of γ-TmT or blank medium for 5 d. The methylation in the Nrf2 promoter was inhibited in the γ-TmT-treated cells compared with the untreated cells at d 5, and mRNA and protein expressions of Nrf2 and NAD(P)H:quinone oxidoreductase 1 were higher. Interestingly, only DNMT3B was inhibited in the γ-TmT-treated cells compared with the untreated cells. In the aggregate, our findings demonstrate that γ-TmT could inhibit CpG methylation in the Nrf2 promoter in the prostate of TRAMP mice and in TRAMP-C1 cells, which might lead to higher Nrf2 expression and potentially contribute to the prevention of prostate tumorigenesis in this TRAMP model.
    Journal of Nutrition 03/2012; · 4.20 Impact Factor
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    ABSTRACT: PURPOSE: This study is to investigate the pharmacokinetics in rat plasma and the pharmacodynamic response of phase II drug metabolizing enzyme genes in rat lymphocytes following intravenous administration of sulforaphane, a phytochemical showing cancer chemopreventive effects in various animal cancer models. METHODS: Single intravenous dose (25 mg/Kg) of sulforaphane was administered to four groups each comprising four male Sprague-Dawley JVC rats. Blood samples (~ 0.3 mL) from each rat were drawn and pooled at 0, 0.033, 0.083, 0.25, 0.5, 0.75, 1, 2, 3, 4, 6, 8, 12, 18, 24, and 48 hours. Plasma were obtained and analyzed using a validated LC/MS/MS method. Lymphocytes were obtained using Ficoll-Paque centrifuge medium. RNAs were extracted, converted to cDNA and quantitative real-time PCR for the relative expression of phase II DME and Nrf2-driven genes, UGT1A1, UGT1A6, GSTM1, GSTT1, NQO1, HO-1, GPx1, Keap1, Maf, Nrf2 and SOD, were performed against those at time zero hour. RESULTS: SFN and its major metabolites, SFN-NAC and SFN-GSH from each plasma sample were assayed followed by WinNonlin Non-compartment analysis. SFN declined with a t1/2 of 1.62 hours, whereas SFN-NAC and SFN-GSH were formed rapidly and then declined with a terminal t1/2 of 5.01 and 8.82 hours. Rat lymphocyte mRNA expression levels show no changes for GSTM1, Keap1, SOD, or UGT1A6. Moderate increase (2-5 folds increase over the control time zero) was seen with HO-1, Nrf2, NQO1, and UGT1A1, and significant increase (> 5 folds) for GSTT1, GPx1, and Maf. Nrf2 and Maf reached their maximal expression level at ~ 1 hour. While the other phase II genes reached their maximal expression at ~ 1-2 hours, GPx1 and HO-1 reached maximal expression at ~ 0.75 hour. CONCLUSION: For the first time, this study simultaneously linked plasma concentrations and lymphocyte mRNA expression levels. The results from this study clearly show that the plasma SFN concentrations would drive the gene expression of phase II DME and Nrf2-mediated genes in rat lymphocytes and provide a framework for future clinical studies.
    American Association of Pharmaceutical Scientists (AAPS) Annual Meeting and Exposition, New Orleans, Louisiana, USA; 11/2010
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    C L Saw, Y Huang, A T Kong
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    ABSTRACT: Purpose. Inflammatory response plays a very important role in normal physiology and pathology such as cancers. As chronic inflammations are associated with malignancies, it is important to prevent the inflammation before neoplasm formation, promotion and/or progression. One way of doing so is by the use of chemopreventive agents such as curcumin (CUR). Polyunsaturated fatty acids (PUFA) such as docosahexaenoic acid (DHA) or eicosapentaenoic acid (EPA) are potentially potent anti-inflammatory agents by decreasing the production of inflammatory eicosanoids, cytokines, and reactive oxygen species. The present study aims to examine whether CUR with DHA or EPA have enhanced chemopreventive effects in RAW 264.7 cells. Methods. The cell toxicity profiles of single or combination treatments were identified. The cells were first treated with various chemopreventive agents, 1 h later the cells were challenged with inflammatory stimulus lipopolysaccharides (LPS). Western blots and qRT-PCR were used to examine the expression levels of inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2) and hemeoxygenase-1 (HO-1). The expression levels of 5-lipoxygenase (5-LOX) and cytosolic phospholipase A2 (cPLA2) were also quantified by qRT-PCR. Results. Of all the treatments, with and without LPS, besides the highest concentration of compounds, i.e. CUR 80 μM, DHA 100 μM and EPA 100 μM, all treatments were non-toxic as compared to non-treated cells. Non-toxic concentrations for single and combination compounds were further examined at 6 h, 12 h and 24 h. The NO suppression effects were most prominent at 24 h. All combinations for CUR and DHA or EPA at lower than CUR 5 μM and 12.5 μM of DHA or EPA were found to have synergistic effects in suppressing NO. Interestingly, it was found that CUR 2.5 μM and DHA or EPA of 0.78125 μM could suppress synergistically the LPS induced-prostaglandin E2 level. The combinations were also found to suppress iNOS, COX-2, 5-LOX and cPLA2 but induce HO-1. Conclusion. The study clearly shows the synergistic anti-inflammatory as well as anti-oxidative stress effects of CUR and PUFA.
    American Association of Pharmaceutical Scientists (AAPS) Annual Meeting and Exposition, Los Angeles, CA, USA; 11/2009
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    ABSTRACT: Purpose. To investigate the role of Nrf2 in suppressing LPS-mediated inflammation in ex vivo macrophages by polyunsaturated fatty acids (PUFA) such as DHA and EPA. Methods. nrf2 +/+ wild-type and nrf2 -/-knock-out mice weighing 20 to 25 grams were injected with thioglycollate broth intraperitoneally. Mice were sacrificed three days later and macrophage cells were collected by injecting 5 mL of phosphate buffer solution (pH 7.4) and collecting the rinsing solution. The cells were plated in Dulbecco's Modified Eagle's Medium supplemented with 10% heat-inactivated fetal bovine serum in cell culture plates with 1×10 6 cells/mL. After two hours of incubation, the cells were treated with 1 µg/mL of lipopolysaccharides (LPS) in the presence or absence of 25, or 50 µM of DHA or EPA. Total RNA were isolated using Qiagen RNeasy Macro Kit six hours later. The RNA were then transcribed into cDNA using Invitrogen SuperScript III First Strand Synthesis System, and agorose gel electrophoreses were performed after PCR. Protein samples were collected using RIPA buffer after 18 hours since LPS treatment, and Western Blot was performed to analyze protein expression levels. Results. Both mRNA and protein analyses show that LPS potently induced cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) in the macrophages collected from nrf2 +/+ wild-type mice, and DHA and EPA potently inhibited LPS-induced COX-2 and iNOS but significantly increased hemoxygenase (HO-1) expression. EPA was found to be more potent than DHA in the inhibition. LPS also induced COX-2 and iNOS in the macrophages collected from nrf2 -/-knock-out mice, however, DHA and EPA were not suppressing COX-2 and iNOS as compared to those from nrf2 +/+ macrophages. As found in nrf2 +/+ macrophages, HO-1 in the macrophages collected from nrf2 -/-mice were also induced by DHA and EPA. Conclusion. It appears that Nrf2 is involved in the biological functions of DHA/EPA in their suppression of LPS-induced inflammation in mouse peritoneal macrophages. The in vivo protective effects of fish oil/PUFAs (DHA/EPA) against acute inflammation under the influence of Nrf2 genotypes would require further investigation.
    American Association of Pharmaceutical Scientists (AAPS) Annual Meeting and Exposition, Los Angeles, CA, USA.; 11/2009