Combinatory effects of Dipterocarpus alatus twig emulgel: Wound-restoring, antibacterial, and anti-inflammatory activities against methicillin-resistant Staphylococcus aureus-infected mouse superficial wounds.

Dipterocarpus alatus has been used for the treatment of infectious skin diseases and ulcerative wounds in Thai traditional medicine. A major pathogen in human superficial skin infections is methicillin-resistant Staphylococcus aureus (MRSA). This study determined the wound healing, antibacterial, and anti-inflammatory activities of D. alatus twig emulgel against MRSA-infected mouse superficial skin wounds. Ethyl acetate-methanol crude extract of D. alatus twig was incorporated into emulgel at concentrations of 20 and 40 mg/g (D20 and D40) and its activity was compared to tetracycline emulgel (160 µg/g, Tetra). MRSA-infected superficial wounds demonstrated decreased skin barrier strength, increased transepidermal water loss (TEWL), and mast cell accumulation. Expression of toll-like receptor 2 (TLR-2), NF-κß, TNFα, IL-1ß, IL-6 and IL-10 genes were induced after MRSA infection. Daily application of 100 µL of D20 or D40 for 9 days restored skin barrier strength and TEWL while reducing mast cell and MRSA numbers compared to the non-treated group (MRSA-NT). The wounds treated with D20 and D40 were entirely healed on day 9. Expression of TLR-2 and cytokine-related genes NF-κß, TNFα, IL-1ß, IL-6 and IL-10 were normalized by treatment with either D20 or D40. Therefore, emulgel containing 20 to 40 mg/g ethyl acetate-methanol crude D. alatus twig extract is a good candidate for development as a topical formulation for MRSA-infected ulcerated wounds.

Garcinia mangostana and α-Mangostin Revive Ulcerative Colitis-Modified Hepatic Cytochrome P450 Profiles in Mice.

<b>Background and Objective:</b> Ulcerative colitis (UC) is inflammation of the large intestine with ulceration but can also cause extraintestinal manifestations (EIM) by damaging surrounding organs such as the liver. <i>Garcinia mangostana</i> (GM) pericarp and α-mangostin (MGS) have been reported to have anti-inflammatory activity. This study evaluated the effects of GM pericarp extract and MGS on the expression of hepatic cytochrome P450 (CYP) enzymes as an EIM of UC. <b>Materials and Methods:</b> Male ICR mice were orally administered GM pericarp extract (40, 200 and 1000 mg/kg/day), MGS (30 mg/kg/day) or sulfasalazine (SUL) (100 mg/kg/day) daily for 7 days. On days 4-7, UC was induced by dextran sulfate sodium (DSS 40 kDa, 6 g/kg/day). Profiles of CYP mRNA expression were determined by RT/qPCR. Alkoxyresorufin <i>O</i>-dealkylation (including ethoxy-, methoxy-, pentoxy- and benzyloxy-resorufin), aniline hydroxylation and erythromycin <i>N</i>-demethylation CYP responsive activities were also examined. <b>Results:</b> The DSS-induced UC mice showed suppressed expression<i> </i>of <i>Cyp1a1</i>, <i>Cyp1a2</i>, <i>Cyp2b9/10</i>, <i>Cyp2e1</i>, <i>Cyp2c29</i>, <i>Cyp2d9</i>, <i>Cyp3a11</i> and <i>Cyp3a13</i> mRNAs. The GM pericarp extract and MGS restored expression of all investigated CYPs and their responsive enzyme activities in DSS-induced UC mice to levels comparable to the control and parallel to the effects of the anti-inflammatory control SUL. <b>Conclusion:</b> The GM is a promising therapy to restore UC-modified hepatic CYP profiles.

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.

Dill Shows Potential for Herb-Drug Interactions via Up-Regulation of CYP1A2, CYP2C19, SULT1A1, NAT2 and ABCB1 in Caco-2 Cells.

<b>Background and Objective:</b> Dill<i> </i>(<i>Anethum graveolens</i> L.) has the potential to develop as a new alternative medicine due to its pharmacological activities. However, studies into its safety regarding herb-drug interactions have been neglected. This study investigated the risk of dill-induced herb-drug interactions (HDI) by examining its effect on the expression of phase I and II drug-metabolizing enzyme and transporter genes in Caco-2 cells. <b>Materials and Methods:</b> Caco-2 cells (5×10⁵ cells/well) were treated with 10 µM ketoconazole, 20 µM rifampicin or dill extract (60-240 µg mL¹) for 72 hrs. Cell viability was assessed using the resazurin assay and reactive oxygen species (ROS) content was determined with 2 ,7 -dichlorofluorescein diacetate. Aspartate (AST) and alanine aminotransferase (ALT) levels were measured using L-aspartate and L-alanine with α-ketoglutarate as substrate. Expression of phase I (<i>CYP1A2</i>, <i>CYP2C19</i>, <i>CYP2D6</i>, <i>CYP2E1 </i>and <i>CYP3A4</i>) and II (<i>UGT1A6</i>,<i> SULT1A1</i>,<i> NAT1</i>,<i> NAT2 </i>and<i> GSTA1/2</i>) metabolizing genes and transporters (<i>ABCB1</i>,<i> ABCC2</i>,<i> ABCG2 </i>and <i>SLCO1B1</i>) were determined by RT/qPCR. <b>Results:</b> All tested concentrations of dill did not affect cell viability or AST and ALT levels. The highest concentration of dill extract (240 µg mL¹) significantly lowered the ROS level. Expression of <i>CYP1A2</i>, <i>CYP2C19</i>, <i>SULT1A1</i>, <i>NAT2 </i>and <i>ABCB1 </i>mRNA was significantly up-regulated by dill extract. <b>Conclusion:</b> Dill extract did not directly damage Caco-2 cells but prolonged use of dill may increase the risk of HDI via the up-regulation of the drug-metabolizing genes <i>CYP1A2</i>, <i>CYP2C19</i>, <i>SULT1A1</i>, <i>NAT2 </i>and the transporter <i>ABCB1</i>.

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.

Permeation, stability and acute dermal irritation of miroestrol and deoxymiroestrol from Pueraria candollei var. mirifica crude extract loaded transdermal gels.

In this study, permeation behaviors and chemical stability of miroestrol and deoxymiroestrol from Pueraria candollei var. mirifica (PM), Thai traditional medicine, crude extract containing transdermal gels were firstly evaluated. Three different PM extract containing gels were formulated, including hydroalcoholic and microemulsion gels using carbomer, and silicone gel using silicone elastomer. In vitro permeation through porcine ear skin demonstrated that the flux and 24 h cumulative permeation of miroestrol and deoxymiroestrol were in the order of hydroalcoholic > silicone > microemulsion gels. Hydroalcoholic gel provided the highest partition coefficient from gel onto skin, and thus the skin permeability coefficient. After 24 h permeation, no miroestrol and deoxymiroestrol remained deposited in the skin. Accelerated study using heating-cooling revealed insignificant difference between the remaining percentages of miroestrol and deoxymiroestrol in aqueous and non-aqueous based gels. Long-term stability study showed that miroestrol contents remained constant for 90 d and 30 d under 5 ± 3 °C and 30 ± 2 °C, 75 ± 5%RH, respectively; whereas the percentage of deoxymiroestrol decreased significantly after 30 d storage, irrespective of storage conditions. Acute dermal irritation test on New Zealand White rabbits showed that PM hydroalcoholic gels were non-irritant, with no signs of erythema or oedema.[Figure: see text].

Protective Role of Vanillic Acid against Diethylnitrosamine- and 1,2-Dimethylhydrazine-Induced Hepatocarcinogenesis in Rats.

This study aimed to evaluate the cancer chemopreventive activity of vanillic acid (VA) in diethylnitrosamine- and 1,2-dimethylhydrazine-induced liver and colon carcinogenesis in rats. VA did not induce the formation of hepatic glutathione S-transferase placental form (GST-P) positive foci and colonic aberrant crypt foci, demonstrating no carcinogenic activity. VA (75 mg kg-1 body weight) could significantly reduce the number and areas of hepatic GST-P positive foci when administered before carcinogen injections, but no such effect was seen when it was administered after carcinogen injection. No protection was seen in the colon when VA was treated before or after carcinogen injection. Immunohistochemical studies demonstrated the decreased expression of proliferating cell nuclear antigen and the induction of apoptosis. Mechanistic studies showed that VA significantly induced the expression of GSTA-5 and Nrf-2 genes, which are associated with the detoxification system. Likewise, the antiproliferative effect was noticed by the reduction of Cyclin D1 expression. The apoptotic activity may be due to the upregulation of Caspase-3 and Bad levels and downregulation of the Bcl-2 level. These data suggest that VA exhibited significant protection against diethylnitrosamine- and 1,2-dimethylhydrazine-induced hepatocarcinogenesis, which might be related to the induction of the detoxifying enzyme, the reduction of proliferation and the induction of apoptosis.

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.

Ethanolic Garcinia mangostana extract and α-mangostin improve dextran sulfate sodium-induced ulcerative colitis via the suppression of inflammatory and oxidative responses in ICR mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Ulcerative colitis (UC) is an inflammatory disorder of the colon. Garcinia mangostana Linn. (GM) has been traditionally used for its anti-inflammatory and antioxidant activities. AIM OF THE STUDY: The effects of GM and its bioactive constituent α-mangostin on dextran sulfate sodium (DSS)-induced UC in mice were investigated. MATERIALS AND METHODS: Adult ICR mice (n = 63) were pretreated with ethanolic GM extract at 40, 200, and 1000 mg/kg/day (GM40, GM200, and GM1000), α-mangostin at 30 mg/kg/day, or sulfasalazine at 100 mg/kg/day (SA) for 7 consecutive days. On days 4-7, UC was induced in the mice by the oral administration of DSS (40 kDa, 6 g/kg/day), while control mice received distilled water. The UC disease activity index (DAI) and histological changes were recorded. The activities of myeloperoxidase, catalase, and superoxide dismutase, and the levels of reactive oxygen species (ROS), nitric oxide (NO), and malondialdehyde (MDA) were determined. The mRNA expression of inflammatory related genes including proinflammatory cytokine Tnf-α, Toll-like receptor (Tlr-2), adhesion molecules (Icam-1 and Vcam-1), and monocyte chemoattractant protein (Mcp-1) were evaluated. RESULTS: Treatment with GM or α-mangostin decreased the UC DAI and protected against colon shortening and spleen and kidney enlargement. GM and α-mangostin prevented histological damage, reduced mast cell infiltration in the colon, and decreased myeloperoxidase activity. GM and α-mangostin increased catalase and superoxide dismutase activity and decreased ROS, NO, and MDA production. GM downregulated mRNA expression of Tnf-α, Tlr-2, Icam-1, Vcam-1, and Mcp-1. CONCLUSIONS: GM and α-mangostin attenuated the severity of DSS-induced UC via anti-inflammatory and antioxidant effects. Therefore, GM is a promising candidate for development into a novel therapeutic agent for UC.

The effect of green tea catechins on breast cancer resistance protein activity and intestinal efflux of aflatoxin B1 via breast cancer resistance protein in Caco-2 cells.

Aflatoxin B1 (AFB1), a mycotoxin produced by Aspergillus spp., was proved as one of the major causes of human hepatocellular carcinoma (HCC) when chronically consumed. An efflux of AFB1 was reported to be associated with breast cancer resistance protein (BCRP) whose activity could also be modulated by green tea catechins. The purpose of this study was, therefore, to examine the impacts of green tea catechins on BCRP activity in Caco-2 cells by H33342 (bis-benzamide, BCRP substrate) accumulation and AFB1 efflux. Results showed a significant decrease (p < 0.05) of AFB1 in the efflux ratio following the incubation with Ko143, a specific BCRP inhibitor, and sodium fluoride, confirming the association of BCRP in AFB1 efflux transport across the cells. Pre-incubation with green tea and gallate catechins (ECG and EGCG) significantly reduced the efflux ratio of AFB1 (p < 0.05) and significantly increased the intracellular H33342 substrate (p < 0.05) in Caco-2 cells, clearly indicating the inhibitory effects of green tea and gallate catechins on BCRP function. Further study on H33342 accumulation revealed a dose-dependent increment of intracellular H33342 when co-administered with increasing concentrations of AFB1. This result implied a possible role of AFB1 as a BCRP competitive inhibitor. The findings from this study concluded the roles of BCRP as an efflux transporter for AFB1 and could be modulated by the exposure of green tea catechins. Owing to a reduction of its efflux, an inhibitory effect of BCRP when pre-exposed with green tea catechins could be crucial for AFB1 cellular accumulation.

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.

Anti-inflammatory effect of Garcinia mangostana Linn. pericarp extract in methicillin-resistant Staphylococcus aureus-induced superficial skin infection in mice.

Garcinia mangostana (mangosteen) pericarp has antibacterial effects; however, information regarding its anti-inflammatory activity in vivo is limited. The anti-inflammatory effect of G. mangostana pericarp extract against methicillin-resistant Staphylococcus aureus (MRSA)-induced superficial skin infection was investigated in mice using a tape stripping model. G. mangostana pericarp ethanolic extract (GME) and its constituent, α-mangostin, were topically administered to mice with MRSA-induced superficial skin infection. MRSA-infected wounds treated with GME were completely healed on the 10th day of the study and the number of MRSA-colonies decreased from the first day of the study, whereas α-mangostin-treated wounds never completely healed with higher numbers of MRSA colonies. The epidermis of GME-treated wounds had nearly completely regenerated, with no inflammatory cell infiltration. In contrast, α-mangostin-treated wounds exhibited neutrophil infiltration and accumulation of mast cells. MRSA-infected wounds without treatment showed high expression of TNF-α, IL-6, IL-1ß, and TLR-2 genes. In contrast, GME decreased mRNA levels, restoring expression of those genes to normal levels. Notably, α-mangostin did not down-regulate the expression of pro-inflammatory cytokines to the same extent as GME. Hence, GME is a promising alternative MRSA treatment because of its antibacterial, anti-inflammatory, and wound healing effects.

Augmentation of diethylnitrosamine-induced early stages of rat hepatocarcinogenesis by 1,2-dimethylhydrazine.

Diethylnitrosamine (DEN) and 1,2-dimethylhydrazine (DMH) are classical carcinogens used in experimental rodent carcinogenesis. However, the interaction effects of these carcinogens on biochemical and molecular changes during carcinogenesis have not been investigated. Therefore, the effect of DEN and DMH co-administration on preneoplastic lesion formation and its molecular mechanism in rats were determined. Triple intraperitoneal administrations of DEN were made before, during or after double subcutaneous injections of DMH. At week 8 of the experiment, the preneoplastic hepatic glutathione-S-transferase placental form (GST-P) positive foci and colonic aberrant crypt foci (ACF) were analyzed. The combined treatment of these carcinogens increased toxicity to rats. Administration of DMH alone did not induce hepatic GST-P positive foci, while co-treatment with DMH enhanced hepatic GST-P positive foci formation. However, DEN did not influence the size or number of colonic ACF. The treatment with DMH alone induced CYP2E1 and P450 reductase, demonstrating that DMH enhanced DEN metabolism in DEN- and DMH-treated rats. These findings were related to increases in hepatic O6-methylguanine DNA adducts and hepatotoxicity, which are associated with the induction of cell proliferation and liver cancer development. DEN-induced early stages of rat hepatocarcinogenesis were synergistically promoted by DMH via metabolic enzyme induction leading to enhanced DNA mutation and hepatocarcinogenicity.

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.

Metabolism of Curcumin in Human Breast Cancer Cells: Impact of Sulfation on Cytotoxicity.

Curcumin is a natural polyphenol with promising anticancer properties that undergoes pronounced metabolism in humans. In order to determine whether metabolism of curcumin also occurs in tumor cells and whether biotransformation has any impact on cytotoxicity, metabolism experiments were conducted with hormone-dependent ZR-75-1 and hormone-independent MDA-MB-231 human breast cancer cells. By using HPLC-ESI-Qq-TOF-MS, it was possible to identify one main metabolite, namely curcumin sulfate, in both cell lines. Its concentration in the cytoplasm and culture medium was 1.6- to 1.7-fold higher in ZR-75-1 cells than in MDA-MB-231 cells, concomitant with a 2-fold higher IC50 value in the ZR-75-1 cell line (14 µM compared to 7.3 µM). The net result of sulfation seems to lower the intracellular concentration of curcumin, thereby decreasing its growth inhibitory activity. Interestingly, for the first time, we also found the formation of a curcumin dimer in the cytoplasm but not in the cellular medium of both cell lines. Compared to curcumin sulfate, however, its maximal intracellular concentrations were up to 4-fold lower, indicating only a minor contribution to the overall curcumin clearance. In conclusion, our data elucidated the metabolism of curcumin in breast cancer cells, which must be considered in humans following oral uptake of dietary curcumin as a chemopreventive agent.

In vivo antibacterial activity of Garcinia mangostana pericarp extract against methicillin-resistant Staphylococcus aureus in a mouse superficial skin infection model.

CONTEXT: Garcinia mangostana Linn. (Guttiferae) (GM) pericarp has been shown to exhibit good in vitro antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA); however, there is currently no available information regarding its in vivo antibacterial activity. OBJECTIVE: To examine in vivo antibacterial activity of G. mangostana extract against MRSA. MATERIALS AND METHODS: GM pericarp was extracted by ethanol (GM-EtOH) and methanol (GM-MeOH). The crude extracts were examined for in vitro antibacterial activity against MRSA using broth microdilution assay. The in vivo antibacterial activity of 10% GM-EtOH against MRSA was determined in a tape stripping mouse model of superficial skin infection for 9 days by evaluating transepidermal water loss (TEWL) and performing colony counts from cultured swabs. RESULTS: GM-EtOH showed greater in vitro activity against MRSA than GM-MeOH in broth microdilution assay with minimum inhibitory concentration 17 versus 20 µg/mL and minimum bactericidal concentration 30 versus 35 µg/mL, respectively. The GM-EtOH (13.20 ± 0.49%) contained α-mangostin more than the GM-MeOH (9.83 ± 0.30%). In the tape stripping mouse model, 10% GM-EtOH reduced the number of MRSA colonies (0-1) recovered from infected wounds (>100 colonies) on the first day of treatment, restored TEWL to normal levels on the fourth day, and had completely healed the wounds by day 9. CONCLUSION: GM-EtOH showed promising in vivo antibacterial activity against MRSA in a superficial skin infection model in mice. It is of interest to develop a topical formulation of GM-EtOH to further study its potential as a novel antibacterial agent.

Imbalance of the antioxidative system by plumbagin and Plumbago indica L. extract induces hepatotoxicity in mice.

BACKGROUND/AIM: Plumbago indica (PI) L. and its active constituent, plumbagin, has been traditionally claimed for several pharmacological activities; however, there is little information regarding their toxicity. The present study aims to examine the effects of plumbagin and PI extract (PI) on hepatic histomorphology and antioxidative system in mice. MATERIALS AND METHODS: Adult male intelligent character recognition mice were intragastrically administered plumbagin (1, 5, and 15 mg/kg/day) or PI (20, 200, and 1,000 mg/kg/day) consecutively for 14 days. Hepatic histomorphology was examined. Plasma alanine transaminase (ALT) and aspartate transaminase (AST) levels, hepatic lipid peroxidation, superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) activities, and the ratio of reduced to oxidized glutathione (GSH/GSSG) were determined. RESULTS: Plumbagin and PI concentration-dependently induced hepatic injury based on histopathological changes via imbalance of antioxidative system. Plumbagin and PI significantly increased plasma ALT and AST levels, hepatic lipid peroxidation, and GPx activity but significantly decreased hepatic SOD and CAT activities. The GSH/GSSG ratio was significantly reduced by plumbagin. CONCLUSION: Plumbagin and PI caused hepatotoxic effects in the mice by unbalancing of the redox defense system. Therefore, plumbagin and PI-containing supplements should be used cautiously, especially when consumed in high quantities or for long periods.

A High-Fat, High-Fructose Diet Induces Antioxidant Imbalance and Increases the Risk and Progression of Nonalcoholic Fatty Liver Disease in Mice.

Excessive fat liver is an important manifestation of nonalcoholic fatty liver disease (NAFLD), associated with obesity, insulin resistance, and oxidative stress. In the present study, the effects of a high-fat, high-fructose diet (HFFD) on mRNA levels and activities of the antioxidant enzymes, including superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx), were determined in mouse livers and brains. The histomorphology of the livers was examined and the state of nonenzymatic reducing system was evaluated by measuring the glutathione system and the lipid peroxidation. Histopathology of the liver showed that fat accumulation and inflammation depended on the period of the HFFD-consumption. The levels of mRNA and enzymatic activities of SOD, CAT, and GPx were raised, followed by the increases in malondialdehyde levels in livers and brains of the HFFD mice. The oxidized GSSG content was increased while the total GSH and the reduced GSH were decreased, resulting in the increase in the GSH/GSSG ratio in both livers and brains of the HFFD mice. These observations suggested that liver damage and oxidative stress in the significant organs were generated by continuous HFFD-consumption. Imbalance of antioxidant condition induced by long-term HFFD-consumption might increase the risk and progression of NAFLD.