Antioxidant Enzymes in Cancer Cells: Their Role in Photodynamic Therapy Resistance and Potential as Targets for Improved Treatment Outcomes.

Photodynamic therapy (PDT) is a selective tumor treatment that consists of a photosensitive compound-a photosensitizer (PS), oxygen, and visible light. Although each component has no cytotoxic properties, their simultaneous use initiates photodynamic reactions (PDRs) and sequentially generates reactive oxygen species (ROS) and/or free radicals as cytotoxic mediators, leading to PDT-induced cell death. Nevertheless, tumor cells develop various cytoprotective mechanisms against PDT, particularly the adaptive mechanism of antioxidant status. This review integrates an in-depth analysis of the cytoprotective mechanism of detoxifying ROS enzymes that interfere with PDT-induced cell death, including superoxide dismutase (SOD), catalase, glutathione redox cycle, and heme oxygenase-1 (HO-1). Furthermore, this review includes the use of antioxidant enzymes inhibitors as a strategy in order to diminish the antioxidant activities of tumor cells and to improve the effectiveness of PDT. Conclusively, PDT is an effective tumor treatment of which its effectiveness can be improved when combined with a specific antioxidant inhibitor.

Impact of Pineapple Juice on Expression of CYP3A4, NAT2, SULT1A1 and OATP1B1 mRNA in HepG2 Cells.

<b>Background and Objective:</b> Pineapple (<i>Ananas comosus</i>) is a popular fruit worldwide with natural antioxidant properties. This study examined how pineapple modified the expression of drug-metabolizing enzymes (CYP1A2, CYP2C9, CYP3A4, UGT1A6, NAT2 and SULT1A1) and a drug transporter (OATP1B1) in human hepatocarcinoma (HepG2) cells. <b>Materials and Methods:</b> HepG2 cells (2.5×10⁵ cells/well in a 24-well plate) were incubated with pineapple juice extract (125-1,000 µg mL¹) for 48 hrs in phenol red-free medium. Resazurin reduction, ROS, AST and ALT assays were performed. The mRNA expression of target genes was determined by RT/qPCR. <b>Results:</b> Pineapple juice slightly reduced HepG2 cell viability to 80% of the control, while ROS, AST and ALT levels were not changed. Pineapple juice did not alter the expression of CYP1A2, CYP2C9 and UGT1A6 mRNA. All tested concentrations of pineapple juice suppressed CYP3A4, NAT2 and OATP1B1 expression, while SULT1A1 expression was induced. <b>Conclusion:</b> Though pineapple juice slightly decreased the viability of HepG2 cells, cell morphology and cell function remained normal. Pineapple juice disturbed the expression of phase I (CYP3A4) and phase II (NAT2 and SULT1A1) metabolizing genes and the drug transporter OATP1B1. Therefore, the consumption of excessive amounts of pineapple juice poses a risk for drug interactions.

Plumbagin and Plumbago indica Differentially Modulated Cytochrome P450 and Transporter Profiles in BeWo and HepG2 Cells.

<b>Background and Objective:</b> The medicinal herb <i>Plumbago indica</i> (PI) and its major constituent plumbagin have reported pharmacological properties but there is a lack of information about their herb-drug interactions. The effects of methanolic (PI-MeOH) and ethanolic (PI-EtOH) crude extracts of PI and plumbagin on the expression of cytochrome P450s (<i>CYP1A2</i>, <i>CYP2E1</i> and <i>CYP3A4</i>) and transporters (<i>ABCC1</i>, <i>ABCG2</i> and <i>SLC22A11</i>) were investigated in BeWo and HepG2 cells. <b>Materials and Methods:</b> BeWo or HepG2 cells were treated with 0.5-5 µM plumbagin or 25-500 µg mL¹ of PI-MeOH or PI-EtOH for 24 hrs. Total RNA was extracted and mRNA expression of CYPs and transporters were determined using RT-qPCR. <b>Results:</b> PI and plumbagin affected mRNA expression differently in the two tested cell types. In BeWo cells, all concentrations of PI-MeOH induced <i>CYP2E1</i>, 100 and 500 µg Ml¹ PI-MeOH and PI-EtOH up-regulated <i>CYP1A2</i>, <i>CYP3A4 </i>and <i>ABCG2 </i>and 500 µg mL¹ PI-EtOH induced <i>ABCG2</i> expression. Plumbagin suppressed <i>CYP1A2</i> and induced <i>SLC22A11 </i>expression at the highest concentration, 5 µM. In HepG2 cells, 5 µM plumbagin and 500 µg Ml¹ PI-EtOH suppressed <i>CYP3A4 </i>expression and 500 µg mL¹ PI-MeOH and PI-EtOH up-regulated <i>CYP1A2</i> and <i>CYP2E1 </i>expression. <i>ABCC1</i> expression was induced by all treatments while <i>ABCG2</i> and <i>SLC22A11 </i>were induced only by 500 µg mL¹ PI-MeOH and PI-EtOH. <b>Conclusion:</b> The use of PI or plumbagin supplements in large quantities or for long periods should be carefully considered due to the risk of herbal drug interactions via modulated expression of CYPs and transporters.

Differential Impacts of Phenol Red on Benzo[a]pyrene and Dexamethasone-Modified Cytochrome P450s in Human Cancer Cells.

<b>Background and Objective:</b> Phenol red, the pH indicator in cell culture media, influences the expression of cytochrome P450s (CYPs) in cell lines. This study aimed to examine how phenol red modified CYP induction by benzo[<i>a</i>]pyrene and dexamethasone in human hepatocarcinoma (HepG2), colorectal adenocarcinoma (Caco-2) and choriocarcinoma (BeWo) cells. <b>Materials and Methods:</b> The cells (1×10⁵ cells/well in a 24-well plate) were incubated with benzo[<i>a</i>]pyrene (0.1, 1 and 10 µM) or dexamethasone (1, 5 and 10 µM) in either phenol red or phenol red-free media for 24 hrs. The mRNA expression of CYPs was determined by Real-Time Polymerase Chain Reaction (RT/qPCR). <b>Results:</b> Phenol red enhanced expression of benzo[<i>a</i>]pyrene-induced CYP1A2 inHepG2 and BeWo cells and suppressed benzo[<i>a</i>]pyrene-induced CYP2A6 expression in HepG2 and Caco-2 cells, benzo[<i>a</i>]pyrene induced CYP2B6 expression in HepG2 cells and benzo[<i>a</i>]pyrene- and dexamethasone-induced CYP3A4 expression in HepG2 and Caco-2 cells. The expression of CYP3A5 was affected differently in HepG2 and Caco-2 cell lines. Phenol red enhanced benzo[<i>a</i>]pyrene- and dexamethasone-induced CYP3A5 expression in Caco-2 cells but suppressed benzo[<i>a</i>]pyrene- and dexamethasone-induced CYP3A5 expression in HepG2 cells. <b>Conclusion:</b> Phenol red differentially influenced expression of benzo[<i>a</i>]pyrene- and dexamethasone-induced CYP1A2, CYP2A6, CYP2B6, CYP3A4 and CYP3A5 mRNAs in HepG2, Caco-2 and BeWo cells. Therefore, the inclusion of phenol red in cell culture media is of concern in studies of drug and xenobiotic metabolism via CYPs in human cell line models.

Impact of Pineapple on Mitochondrial Permeability Transition and Drug Metabolizing Genes in Caco-2 Cells.

<b>Background and Objective:</b> Pineapple (<i>Ananas comosus</i> L.) has antioxidant and other pharmacological properties. This study examined how pineapple modified mitochondrial permeability transition and expression of drug-metabolizing enzymes, i.e., CYP1A2, CYP2C9, CYP3A4, UGT1A6, NAT2 and the drug transporter OATP1B1 in human colorectal adenocarcinoma (Caco-2) cells. <b>Materials and Methods:</b> Caco-2 cells (2.5×10⁵ cells well¹ in 24-well plates) were incubated with pineapple (125 to 1,000 µg mL¹) for 48 hrs in a phenol red-free medium. Mitochondrial permeability transition, resazurin cell viability and AST and ALT levels were investigated. The mRNA expression of target genes was determined by RT/qPCR. <b>Results:</b> Pineapple significantly reduced depolarized mitochondria, slightly decreased cell viability and did not change AST and ALT levels. Pineapple did not modify the mRNA expressions of CYP1A2, CYP2C9 and CYP3A4 but markedly induced UGT1A6 expression. The highest tested concentration of pineapple (1,000 µg mL¹) significantly suppressed NAT2 and OATP1B1 expression. <b>Conclusion:</b> Although pineapple slightly decreased cell viability to ~80% of control, the morphology and functions of the cells were unaffected. Pineapple showed a beneficial effect to reduce depolarized mitochondria, which consequently decreased reactive oxygen species production. Pineapple did not modify the expression of CYPs, whilst it altered the expression of phase 2 metabolizing genes UGT1A6 and NAT2 and the transporter OATP1B1. Therefore, the consumption of large amounts of pineapple is of concern for the risk of drug interaction via alteration of UGT1A6, NAT2 and OATP1B1 expression.

Reused palm oil from frying pork or potato induced expression of cytochrome P450s and the SLCO1B1 transporter in HepG2 cells.

Deep frying degrades the oil and generates harmful products. This study evaluated effects of reused palm oil (from frying pork or potato) on expression of cytochrome P450s (CYPs), the transporter (SLCO1B1), and lipid metabolism regulators; proliferator-activated receptors (PPAR) and sterol regulatory element binding protein (SREBP). Human hepatic carcinoma cell line (HepG2) cells were incubated with oleic acid (OA), new palm oil, or reused palm oils for 24 hr. Fatty acid accumulation was examined by Nile red staining. Total RNA was extracted, followed by RT/qPCR of the target genes. Fatty acid accumulation was significantly different between the new and the reused oils. Expression of CYP1A2, CYP2C19, CYP2E1, CYP3A4, CYP4A11, and SLCO1B1 was induced by reused oils. Expression of PPAR-α was strongly increased in all treatments while SREBP-1a and SREBP-1c were suppressed. Modification of CYPs, PPAR-α, and SLCO1B1 by palm oil might increase the risk of fatty acid accumulation with associated oxidative stress. Therefore, consumption of palm oil or reused oil should be limited. PRACTICAL APPLICATIONS: Deep frying degrades the oil and generates harmful products. This study evaluated effects of reused palm oil (from frying pork or potato) on expression of cytochrome P450s (CYPs), the transporter (SLCO1B1), and lipid metabolism regulators; PPAR and SREBP in HepG2 cells. Both of the reused oils-induced profiles of all CYP and SLCO1B1, but the new oil upregulated CYP2E1, CYP3A4, and CYP4A11. PPAR-α was induced while SREBP-1a and SREBP-1c were suppressed by all treatments. Inductions of CYPs with suppression of SREBP-1a and SREBP-1c might contribute to an increased risk of fatty acid accumulation. These findings revealed the impacts of reused palm oil on metabolism via CYPs which related to oxidative stress for further study. Hence, consumption of palm oil or reused cooking oil should be of concern.

Physiology and Pathophysiology of Steroid Biosynthesis, Transport and Metabolism in the Human Placenta.

The steroid hormones progestagens, estrogens, androgens, and glucocorticoids as well as their precursor cholesterol are required for successful establishment and maintenance of pregnancy and proper development of the fetus. The human placenta forms at the interface of maternal and fetal circulation. It participates in biosynthesis and metabolism of steroids as well as their regulated exchange between maternal and fetal compartment. This review outlines the mechanisms of human placental handling of steroid compounds. Cholesterol is transported from mother to offspring involving lipoprotein receptors such as low-density lipoprotein receptor (LDLR) and scavenger receptor class B type I (SRB1) as well as ATP-binding cassette (ABC)-transporters, ABCA1 and ABCG1. Additionally, cholesterol is also a precursor for placental progesterone and estrogen synthesis. Hormone synthesis is predominantly performed by members of the cytochrome P-450 (CYP) enzyme family including CYP11A1 or CYP19A1 and hydroxysteroid dehydrogenases (HSDs) such as 3ß-HSD and 17ß-HSD. Placental estrogen synthesis requires delivery of sulfate-conjugated precursor molecules from fetal and maternal serum. Placental uptake of these precursors is mediated by members of the solute carrier (SLC) family including sodium-dependent organic anion transporter (SOAT), organic anion transporter 4 (OAT4), and organic anion transporting polypeptide 2B1 (OATP2B1). Maternal-fetal glucocorticoid transport has to be tightly regulated in order to ensure healthy fetal growth and development. For that purpose, the placenta expresses the enzymes 11ß-HSD 1 and 2 as well as the transporter ABCB1. This article also summarizes the impact of diverse compounds and diseases on the expression level and activity of the involved transporters, receptors, and metabolizing enzymes and concludes that the regulatory mechanisms changing the physiological to a pathophysiological state are barely explored. The structure and the cellular composition of the human placental barrier are introduced. While steroid production, metabolism and transport in the placental syncytiotrophoblast have been explored for decades, few information is available for the role of placental-fetal endothelial cells in these processes. With regard to placental structure and function, significant differences exist between species. To further decipher physiologic pathways and their pathologic alterations in placental steroid handling, proper model systems are mandatory.

Immune response and inflammatory pathway of ulcerative colitis.

Ulcerative colitis (UC) is an idiopathic relapsing inflammatory disease. Although the etiology of UC remains unclear, it could be characterized by inflammation of the intestinal mucosa, starting from the rectum and potentially involving the entire colon. The immune response and inflammatory pathway of UC have shown that tissue damage is driven by dynamic and complexes of cells and cytokines. Various types of cells, including antigen-presenting cells (dendritic cells and macrophages), T helper cells, regulatory T cells, and natural killer T cells, play a crucial role in UC pathogenesis by regulation, suppression, and maintenance of inflammation. Moreover, cytokine networks become an important part due to their signaling function, which is indispensable for cell communication. Pro-inflammatory cytokines [tumor necrosis factor-α, interleukin (IL)-1, IL-6, IL-9, IL-13, and IL-33] play significant roles in upregulation, while anti-inflammatory cytokines (transforming growth factor-ß, IL-10, and IL-37) play significant roles in downregulation of disease progression. The pathogenesis of UC consists of immuno-inflammatory pathways related to the multiple components of the intestine, including the epithelial barrier, commensal microflora, antigen recognition, dysregulation of immunological responses, leukocyte recruitment, and genetic factors. The understanding of immuno-inflammatory pathways of UC might lead to the development of a specific therapy and/or a novel treatment that could be more efficient.

Fenofibrate inhibits tumour intravasation by several independent mechanisms in a 3-dimensional co-culture model.

Lymph node metastasis of breast cancer is a clinical marker of poor prognosis. Yet, there exist no therapies targeting mechanisms of intravasation into lymphatics. Herein we report on an effect of the antidyslipidemic drug fenofibrate with vasoprotective activity, which attenuates breast cancer intravasation in vitro, and describe the potential mechanisms. To measure intravasation in a 3-dimensional co-culture model MDA-MB231 and MCF-7 breast cancer spheroids were placed on immortalised lymphendothelial cell (LEC) monolayers. This provokes the formation of circular chemorepellent induced defects (CCIDs) in the LEC barrier resembling entry ports for the intravasating tumour. Furthermore, the expression of adhesion molecules ICAM-1, CD31 and FAK was investigated in LECs by western blotting as well as cell-cell adhesion and NF-κB activity by respective assays. In MDA-MB231 cells the activity of CYP1A1 was measured by EROD assay. Fenofibrate inhibited CCID formation in the MDA-MB231/LEC- and MCF-7/LEC models and the activity of NF-κB, which in turn downregulated ICAM-1 in LECs and the adhesion of cancer cells to LECs. Furthermore, CD31 and the activity of FAK were inhibited. In MDA-MB231 cells, fenofibrate attenuated CYP1A1 activity. Combinations with other FDA-approved drugs, which reportedly inhibit different ion channels, attenuated CCID formation additively or synergistically. In summary, fenofibrate inhibited NF-κB and ICAM-1, and inactivated FAK, thereby attenuating tumour intravasation in vitro. A combination with other FDA-approved drugs further improved this effect. Our new concept may lead to a novel therapy for cancer patients.

Regulation of cancer-related genes - Cyp1a1, Cyp1b1, Cyp19, Nqo1 and Comt - expression in ß-naphthoflavone-treated mice by miroestrol.

OBJECTIVE: The effects of miroestrol (MR), an active phytoestrogen from Pueraria candollei var. mirifica, on expression of cancer-related genes were determined. METHODS: Seven-week-old female ICR mice (n = 5 each) were subcutaneously administered estradiol (E2, 0.5 mg/kg/day) or MR (0.5 or 5 mg/kg/day) daily for 7 days. Some were given ER or MR in combination with ß-naphthoflavone (BNF, 30 mg/kg/day) for the last 3 days. The expression of cancer-related genes including cytochrome P450 1A (Cyp1a), cytochrome P450 1B1 (Cyp1b1), aromatase P450 (Cyp19), NAD(P)H: quinone oxidoreductase 1 (Nqo1) and catechol-O-methyltransferase (Comt) were evaluated. KEY FINDINGS: In the presence of BNF, MR suppressed hepatic CYP1A1 activity and CYP1A2 activity, expression of CYP1B1 mRNA and expression of CYP1A1/2 and CYP1B1 protein. E2, by contrast, did not. MR restored expression levels of hepatic NQO1 and uterine COMT in BNF-treated mice. Furthermore, MR increased expression of uterine CYP19 to the same extent as E2. CONCLUSION: MR may be superior to E2 as it downregulates expression of CYP1. Moreover, MR normalized expression of both NQO1 and COMT, the protective enzymes, in murine liver and uteri. These results support the use of MR as an alternative supplement for menopausal women, MR having the extra benefit of reducing cancer risk.

AHR/CYP1A1 interplay triggers lymphatic barrier breaching in breast cancer spheroids by inducing 12(S)-HETE synthesis.

A causal link between overexpression of aryl hydrocarbon receptor (AHR) and its target cytochrome P450 1A1 (CYP1A1) and metastatic outgrowth of various cancer entities has been established. Nevertheless, the mechanism how AHR/CYP1A1 support metastasis formation is still little understood. In vitro we discovered a potential mechanism facilitating tumour dissemination based on the production of 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE). Utilising a three-dimensional lymph endothelial cell (LEC) monolayer & MDA-MB231 breast cancer cell spheroid co-culture model in combination with knock-down approach allowed elucidation of the molecular/biochemical basis of AHR/CYP1A1-induced tumour breaching through the LEC barrier. Enzyme immunoassay evidenced the potential of recombinant CYP1A1 to synthesise 12(S)-HETE in vitro and qPCR and Western blotting measured gene and protein expression in specific experimental settings. In detail, AHR induced CYP1A1 expression and 12(S)-HETE secretion in tumour spheroids, which caused LEC junction retraction thereby forming large discontinuities allowing transmigration of the tumour. This was enforced by the activating AHR ligand 6-formylindolo (3,3-b)carbazole (FICZ), or inhibited by the AHR antagonist 3,3'-diindolylmethane (DIM) as well as by siRNA against AHR and CYP1A1. AHR and NF-κB were negatively cross talking and therefore, the inhibition of AHR (but not CYP1A1) induced RELA, RELB, NFKB1, NFKB2 and the NF-κB target MMP1, which itself promotes tumour intravasation by a mechanism that is different from 12(S)-HETE. Conversely, the inhibition of NFKB2 induced AHR, CYP1A1 and 12(S)-HETE synthesis. The approved clinical drugs guanfacine and vinpocetine, which inhibit CYP1A1 and NF-κB, respectively, significantly inhibited LEC barrier breaching in vitro indicating an option to reduce metastatic dissemination.

Thai red rice extract provides liver protection in paracetamol-treated mice by restoring the glutathione system.

CONTEXT: The incidence of drug-induced liver disease associated with oxidant-antioxidant imbalance is increasing. Colored rice can potentially improve these hepatic disorders through antioxidative and glutathione-restoring effects. OBJECTIVES: The objective of this study is to determine the in vitro antioxidant properties of extracts from red (Hom-Dang and Hom-Kularb-Dang) and black (Hom-Dum-Sukhothai and Kum-Doi-Saket) Thai rice cultivars [Oryza sativa L. (Poaceae)] and to examine the in vivo hepatoprotective potential of Hom-Dang extract in paracetamol-treated mice. MATERIALS AND METHODS: The in vitro antioxidant properties of the extracts were determined by ABTS, [Formula: see text], [Formula: see text], metal chelating capacity, and lipid peroxidation assays. To investigate hepatoprotective effects in vivo, mice administered 60 mg/kg/d paracetamol were given Hom-Dang extract (128, 256, and 512 mg/kg/d) and/or control antioxidant N-acetyl-cysteine (NAC, 150 mg/kg/d) for 7 and 30 d. Liver health was ascertained by measuring levels of hepatic transaminases (GPT/GOT), determining the glutathione profile (GSH/GSSG ratio), and histomorphological examination of liver tissue. RESULTS: Hom-Dang extract showed the highest in vitro antioxidant potency (an IC50 value of 36.50 ± 0.46, 12.98 ± 0.23, 21.83 ± 2.58, 15.87 ± 0.30, and 86.21 ± 2.45 mg/mL for ABTS, OH(•), [Formula: see text], metal chelating, and lipid peroxidation, respectively). Mice administered paracetamol exhibited increases in GPT/GOT with decreases in GSH and GSH/GSSG ratio followed by histomorphological signs of liver injury. In the presence of the Hom-Dang extract, the GPT/GOT values were normalized, GSH production was induced, and the GSH/GSSG ratio was increased. CONCLUSION: Thai colored rice cultivars, especially the Hom-Dang variety, are promising candidates for health supplements due to their antioxidative and hepatoprotective properties.

The germacranolide sesquiterpene lactone neurolenin B of the medicinal plant Neurolaena lobata (L.) R.Br. ex Cass inhibits NPM/ALK-driven cell expansion and NF-κB-driven tumour intravasation.

BACKGROUND: The t(2;5)(p23;q35) chromosomal translocation results in the expression of the fusion protein NPM/ALK that when expressed in T-lymphocytes gives rise to anaplastic large cell lymphomas (ALCL). In search of new therapy options the dichloromethane extract of the ethnomedicinal plant Neurolaena lobata (L.) R.Br. ex Cass was shown to inhibit NPM/ALK expression. PURPOSE: Therefore, we analysed whether the active principles that were recently isolated and found to inhibit inflammatory responses specifically inhibit growth of NPM/ALK+ ALCL, leukaemia and breast cancer cells, but not of normal cells, and the intravasation through the lymphendothelial barrier. METHODS: ALCL, leukaemia and breast cancer cells, and normal peripheral blood mononuclear cells (PBMCs) were treated with isolated sesquiterpene lactones and analysed for cell cycle progression, proliferation, mitochondrial activity, apoptosis, protein and mRNA expression, NF-κB and cytochrome P450 activity, 12(S)-HETE production and lymphendothelial intravasation. RESULTS: In vitro treatment of ALCL by neurolenin B suppressed NPM/ALK, JunB and PDGF-Rß expression, inhibited the growth of ALCL cells late in M phase, and induced apoptosis via caspase 3 without compromising mitochondrial activity (as a measure of general exogenic toxicity). Moreover, neurolenin B attenuated tumour spheroid intravasation probably through inhibition of NF-κB and CYP1A1. CONCLUSION: Neurolenin B specifically decreased pro-carcinogenic NPM/ALK expression in ALK+ ALCL cells and, via the inhibition of NF-kB signalling, attenuated tumour intra/extravasation into the lymphatics. Hence, neurolenin B may open new options to treat ALCL and to manage early metastatic processes to which no other therapies exist.

Effect of tetrahydrocurcumin on the profiles of drug-metabolizing enzymes induced by a high fat and high fructose diet in mice.

Cytochrome P450 (CYP), a superfamily of hepatic monooxygenase enzymes, catalyzes biotransformation of endogenous compounds and xenobiotics. Modification of CYPs associated with metabolic diseases and continuous consumption of diet with excessive energy levels. Tetrahydrocurcumin (THC) exhibited beneficial effects in metabolic syndromes such as diabetic mellitus and dyslipidemia. The present study aimed to investigate the effects of THC and vitamin E (vitE) on the expression profiles of CYPs in the livers of mice fed with the high fat and high fructose diet. In addition to ad libitum access to commercial regular diet, the high fat and high fructose diet (HFD) group of adult male ICR mice was administered a HFD, which consisted of intragastric administration of hydrogenated soybean oil (1mL/day) and the addition of 20% fructose to the drinking water for 8weeks. During the induction period, subgroups of mice (n=5) were daily intragastrically administered with THC (100 or 200mg/kg/day) or vitE (100mg/kg/day). The expressions of CYP mRNA and protein were quantified using real-time PCR and the levels of these proteins were quantified using immunoblotting. Continuous consuming of high fat and high fructose for 8weeks significantly increased the expressions of Cyp1a1, Cyp1a2, Cyp1b1, Cyp2c29, and Cyp3a11 while THC ultimately normalized these CYPs profiles. In the control mice, most of the investigated CYPs was unchanged by THC, with the exception that the Cyp1a1, Cyp2b9, and Cyp3a11 proteins were elevated. These findings provided additional important information on the effects of THC on diet induced-metabolic dysfunctions. However, drug interactions due to the use of THC as an alternative supplement are of concern, particularly in the combinations that include a drug that is a substrate of Cyp1a1, Cyp2b9, and Cyp3a11.

Improvement of antioxidant balance in diabetes mellitus type 1 mice by glutathione supplement.

Diabetes mellitus (DM) type 1 is a chronic disease characterized by hyperglycemia and lacking of insulin. Oxidative stress participates in development and progression of DM, in which changes of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione (GSH) content were noted in DM mice. In this study, the effects of GSH supplement on anti-oxidation system in streptozotocin-induced DM type 1 Imprinting Control Region (ICR) mice were determined. The co-treatment of insulin and GSH significantly lowered the hepatic manganese superoxide dismutase (Mn-SOD), CAT, and GPx mRNA expression. Moreover, co-administration of insulin and GSH restored SOD and CAT activities to non-DM group except that of the CAT activity in the kidney. The GSH contents and GSH/GSSG ratio in the mouse livers were normalized to the normal levels by the GSH treatment and the co-administration of insulin and GSH. These observations reveal that GSH supplement potentially has the protective roles in delaying diabetic progression via the improvement of antioxidant balance.

Miroestrol, a phytoestrogen from Pueraria mirifica, improves the antioxidation state in the livers and uteri of ß-naphthoflavone-treated mice.

Oxidative stress is involved in the progression of several diseases such as diabetes, hypertension, and age-related diseases. Miroestrol (MR) is a potent phytoestrogen from the tuberous root of Pueraria mirifica, a plant used in traditional Thai medicine that is claimed to have rejuvenating effects. In this study, the effects of MR on the antioxidation system, including anti-lipid peroxidation; on the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase; and on glutathione content in the livers and uteri of ß-naphthoflavone (BNF)-treated mice were determined. BNF-treated mice are a model of procarcinogen-exposed mice. The results showed that MR improved the antioxidant activities of SOD and CAT in the livers and uteri of both normal and BNF-treated mice, while estradiol (E2) increased SOD activity in the uteri of normal mice and CAT activity in the livers of both normal and BNF-treated mice. In the liver, MR increased the levels of several forms of glutathione, whereas in the uteri E2 and MR reduced the level of lipid peroxidation by decreasing the level of malondialdehyde. Therefore, the use of MR as an alternative hormone replacement therapy might be beneficial due to its ability to improve antioxidation systems.

Effects of Pueraria mirifica and miroestrol on the antioxidation-related enzymes in ovariectomized mice.

OBJECTIVES: The influences of Pueraria candollei var. mirifica (PM), a Thai medicinal plant with long tradition of medicinal consumption among menopausal women for rejuvenation and estrogen hormone replacement, on oxidative status in ovariectomized (OVX) mice were determined. METHODS: The crude extract of PM and its active phytoestrogen, miroestrol (MR), were given to OVX mice. The effect of them on antioxidation enzymes and glutathione (GSH) levels in livers and uteri were examined in OVX mice and compared with the synthetic estradiol hormone. KEY FINDINGS: Ovariectomy significantly decreased total GSH content, reduced GSH content, and the ratio of GSH to oxidized glutathione (GSSG) in both the livers and the uteri of mice. Moreover, an ovariectomy reduced the activities of glutathione peroxidase (GPx), superoxide dismutase (SOD), and catalase (CAT). The crude extract of PM as well as MR significantly increased levels of GSH, levels of reduced GSH, and the ratio of GSH/GSSG in both the livers and the uteri, while estradiol did not. In addition, the potential of PM and MR to return the activities of GPx, SOD, and CAT to normal levels was noted. CONCLUSIONS: These observations support using PM and MR as promising alternative medicine candidates for hormone replacement therapy of estradiol because of their ability to improve GSH levels and the activities of antioxidative enzymes, especially in OVX mice.

Impact of Pueraria candollei var. mirifica and its potent phytoestrogen miroestrol on expression of bone-specific genes in ovariectomized mice.

Miroestrol (MR) is a highly active phytoestrogen isolated from tuberous root of Pueraria candollei var. mirifica (PM). Modulatory effects of PM and MR on osteoprotegerin (OPG) and receptor activator of nuclear factor kappa B ligand (RANKL) mRNAs which are bone-specific genes were investigated in ovariectomized female ICR mice. After ovariectomy, expression of OPG mRNA was suppressed but that of RANKL was induced. Estradiol benzoate (E2) recovered OPG expression to the level comparable to the sham while that of RANKL was suppressed in ovariectomized mice. PM crude extract (PME) significantly down-regulated the expression of RANKL mRNA with no change in the OPG level whereas MR elevated the expression of OPG mRNA with lowering level of RANKL mRNA, resulting in the increased OPG/RANKL ratio, and consequently lead to lowering progression of osteoporosis at molecular level. These findings revealed potential of PME and MR on bone loss prevention via increasing the ratio of OPG to RANKL (osteoformation/osteoresorption) in liver of ovariectomized mice. Therefore, using PME and MR as alternative hormone replacement therapy of E2 might be beneficial recommended due to advantageous on regulation of osteoporosis related genes.

Alteration of hepatic glutathione peroxidase and superoxide dismutase expression in streptozotocin-induced diabetic mice by berberine.

CONTEXT: Diabetes mellitus (DM), a chronic disease, has been increasing and subsequently devastates the quality of life and economic status of the patients. Oxidative stress participates in development and progression of diabetes, in which levels of glutathione peroxidase (GPx) and superoxide dismutase (SOD) were changed in diabetic mice. Berberine has been widely used as an alternative medicine and proved to be effective for treatment of DM and dyslipidemia. OBJECTIVE: Impacts of berberine on regulation of GPx and SOD messenger RNAs (mRNAs), and glutathione (GSH) content were examined in diabetic mice to clarify its antioxidative stress potential. MATERIALS AND METHODS: Noninsulin-dependent diabetes was induced in mice by a single intraperitoneal streptozotocin injection. Diabetic mice were daily treated with metformin (100 mg/kg/d) or berberine (200 mg/kg/d) for 2 weeks. The fasting blood glucose and GSH content were monitored. GPx and SOD mRNA expression were semi-quantified by reverse transcription-polymerase chain reaction. RESULTS: Berberine showed the same hypoglycemic potential as metformin, a hypoglycemic drug. Interestingly, berberine did not change levels of GPx, copper-zinc SOD (CuZn-SOD), and manganese SOD (Mn-SOD) mRNA in the normal mice but significantly recovered these levels in the diabetic mice to nearly the same levels as the normal. The GSH contents, including total GSH and reduced/oxidized GSH contents, were restored to the normal level by berberine, corresponded to GPx levels. DISCUSSION AND CONCLUSION: Berberine conveyed antioxidative effect via down- and up-regulation of GPx and CuZn-SOD expression, respectively. Therefore, use of berberine as a hypoglycemic compound for alternative treatment of DM could bring extra-beneficent consequence according to its antioxidative stress.

Different AhR binding sites of diterpenoid ligands from Andrographis paniculata caused differential CYP1A1 induction in primary culture in mouse hepatocytes.

Andrographis paniculata has been employed as a folklore remedy. Andrographolide (Andro), 14-deoxy-11,12-didehydroandrographolide (DHA), andrographiside (AS), and neoandrographolide (Neo), are major diterpenoids isolated from this plant. In the present study, influence of the four diterpenoids on CYP1A1 mRNA expression was investigated in primary cultured mouse hepatocytes. Additionally, binding of these compounds to aryl hydrocarbon receptor (AhR) was examined using molecular docking analysis to clarify mechanism of CYP1A1 induction. Andro and DHA induced CYP1A1 expression by itself, and co-treatment with a CYP1A1 inducer (BNF, beta-naphthoflavone) showed a synergistic increase of CYP1A1 expression. Andro demonstrated higher enhancing activity than DHA at every similar concentration. On the other hand, Neo suppressed BNF-induced CYP1A1 expression, but AS did not modify the induction. Results from molecular docking analysis of BNF and four diterpenoids on ligand binding domain of AhR were consistent with levels of CYP1A1 mRNA expressions. Furthermore, difference of binding sites of BNF in the presence of diterpenoids might affect the synergism or inhibition of CYP1A1 expression. These results suggest that use of A. paniculata as a health supplement should be concerned in term of herb-drugs interactions or risk of carcinogenesis, according to its ability to influence CYP1A1 expression.