Rosana Crespo

Increased mortality of Acanthoscelides obtectus by alkane-grown Beauveria bassiana

The effect of alkane-growth induction of theentomopathogenic fungus Beauveria bassiana(Balsamo) Vuillemin (Deuteromycotina:Hyphomycetes), on the ability to kill the beanweevil Acanthoscelides obtectus Say(Coleoptera: Bruchidae), was tested. Adultinsects were sprayed with an 0.01% Tween 20aqueous suspension of 4 × 106 conidia/ml.The performance of fungi grown in complete agarmedium containing glucose as carbon source(FS0) was compared to that of alkane-grownfungi (FS1) with n-hexadecane as the onlycarbon source. Mortality increased (p< 0.05) from 22 ± 4.5% to 44 ±11.4% at day 7, and from 26 ± 5.5% to 60± 7.1% 14 days after treatment withFS0 or FS1 respectively. The insectepicuticular hydrocarbons were analysed bycapillary gas chromatography (CGC); majorcomponents were saturated hydrocarbons, 27 to29 carbons in length. A variety ofmethyl-branched isomers of C27 were theprevailing structures, and nC27 was the majorstraight chain component. Whole insecthydrocarbons were qualitatively identical tothose of the epicuticular surface. Oleic,linoleic and palmitic acids accounted foralmost 88% of the fungal fatty acids,irrespective of the carbon source used forgrowth; however, the unsaturated/saturatedratio diminished markedly from 4.32(FS0) to 2.47 (FS1). These resultsindicate that alkane supplementation of culturemedia might be a tool to improve the virulenceof some mycoinsecticides.

Geraniol and simvastatin show a synergistic effect on a human hepatocarcinoma cell line.

Simvastatin is a competitive inhibitor of 3‐hydroxymethylglutaryl coenzyme A reductase activity, whereas geraniol is a monoterpene with multiple pharmacologic effects on mevalonate metabolism. Both of them inhibit growth and proliferation of many cell lines. The present study was designed to determine the action of geraniol, in combination with simvastatin, by assessing their effects in vitro on human hepatocarcinoma cell line (Hep G2). The treatment of Hep G2 cells with concentrations of simvastatin or geraniol that did not inhibit cell proliferation (5 µmol·l‐1 of simvastatin and 50 µmol·l‐1 of geraniol) resulted in a significant inhibition of cell proliferation. We also examined the effect of simvastatin, geraniol and the combination of both on the biosynthesis of lipids from [14C]‐acetate. Our results demonstrate that the combination of simvastatin and geraniol synergistically inhibited cholesterol biosynthesis and proliferation of Hep G2 cell line, contributing to a better understanding of the action of a component of essential oils targeting a complex metabolic pathway, which would improve the use of drugs or their combination in the fight against cancer and/or cardiovascular diseases. Copyright

Synergistic antiproliferative and anticholesterogenic effects of linalool, 1,8-cineole, and simvastatin on human cell lines.

Monoterpenes are naturally occurring plant hydrocarbons with multiple effects on the mevalonate pathway (MP), while statins competitively inhibit hydroxymethylglutarylcoenzyme-A reductase (HMGCR), the rate-limiting enzyme in the MP. Monoterpenes and statins proved capable of inhibiting both proliferation and cholesterogenesis. In the present study we assess the in vitro antiproliferative and anticholesterogenic effects of two monoterpenes: linalool and 1,8-cineole-either alone, in combination with each other, or combined individually with simvastatin-on liver-derived (HepG2) and extrahepatic (A549) cell lines. The three compounds alone inhibited cell proliferation in a dose-dependent fashion, while their pairwise combination produced synergistic antiproliferative effects in both cell lines. Incorporation experiments with [(14)C]acetate revealed that linalool and 1,8-cineole inhibited the MP, probably at different points, resulting in a reduction in cholesterogenesis and an accumulation of other MP intermediates and products. Linalool or 1,8-cineole, either together or individually with simvastatin, synergistically inhibited cholesterol synthesis. At low concentrations both monoterpenes inhibited steps specifically involved in cholesterol synthesis, whereas at higher concentrations HMGCR levels became down-regulated. Added exogenous mevalonate failed to reverse the inhibition of proliferation exerted by linalool and 1,8-cineole, suggesting that HMGCR inhibition alone is not responsible for the antiproliferative activity of those agents. This work demonstrates that monoterpenes in combination with each other, or individually in combination with simvastatin synergistically inhibits proliferation and cholesterogenesis in the human cell lines investigated, thus contributing to a clearer understanding of the action of essential-oil components, and their combination with the statins, in the targeting of specific points within a complex metabolic pathway.

Modulation by geraniol of gene expression involved in lipid metabolism leading to a reduction of serum-cholesterol and triglyceride levels.

BACKGROUND Geraniol (G) is a natural isoprenoid present in the essential oils of several aromatic plants, with various biochemical and pharmacologic properties. Nevertheless, the mechanisms of action of G on cellular metabolism are largely unknown. HYPOTHESIS/PURPOSE We propose that G could be a potential agent for the treatment of hyperlipidemia that could contribute to the prevention of cardiovascular disease. The aim of the present study was to advance our understanding of its mechanism of action on cholesterol and TG metabolism. STUDY DESIGN/METHODS NIH mice received supplemented diets containing 25, 50, and 75 mmol G/kg chow. After a 3-week treatment, serum total-cholesterol and triglyceride levels were measured by commercial kits and lipid biosynthesis determined by the [(14)C] acetate incorporated into fatty acids plus nonsaponifiable and total hepatic lipids of the mice. The activity of the mRNA encoding HMGCR-the rate-limiting step in cholesterol biosynthesis-along with the enzyme levels and catalysis were assessed by real-time RT-PCR, Western blotting, and HMG-CoA-conversion assays, respectively. In-silico analysis of several genes involved in lipid metabolism and regulated by G in cultured cells was also performed. Finally, the mRNA levels encoded by the genes for the low-density-lipoprotein receptor (LDLR), the sterol-regulatory-element-binding transcription factor (SREBF2), the very-low-density-lipoprotein receptor (VLDLR), and the acetyl-CoA carboxylase (ACACA) were determined by real-time RT-PCR. RESULTS Plasma total-cholesterol and triglyceride levels plus hepatic fatty-acid, total-lipid, and nonsaponifiable-lipid biosynthesis were significantly reduced by feeding with G. Even though an up-regulation of the mRNA encoding HMGCR occurred in the G treated mouse livers, the protein levels and specific activity of the enzyme were both inhibited. G also enhanced the mRNAs encoding the LDL and VLDL receptors and reduced ACACA mRNA, without altering the transcription of the mRNA encoding the SREBF2. CONCLUSIONS The following mechanisms may have mediated the decrease in plasma lipids levels in mice: a down-regulation of hepatocyte-cholesterol synthesis occurred as a result of decreased HMGCR protein levels and catalytic activity; the levels of LDLR mRNA became elevated, thus suggesting an increase in the uptake of serum LDL, especially by the liver; and TG synthesis became reduced very likely because of a decrease in fatty-acid synthesis.

Linalool induces cell cycle arrest and apoptosis in HepG2 cells through oxidative stress generation and modulation of Ras/MAPK and Akt/mTOR pathways

Aims: Linalool is a plant‐derived monoterpene with anticancer activity, however its mechanisms of action remain poorly understood. The aim of this work was to elucidate the anticancer mechanisms of action of linalool in hepatocellular carcinoma (HCC) HepG2 cells. Main methods: Cell viability and proliferation were determined by WST‐1 assay and BrdU incorporation, respectively. Cell cycle analysis was assessed through flow cytometry (FC) and western blot (WB). Apoptosis was determined by caspase‐3 activity, TUNEL assay and WB. Reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) were analyzed by FC and fluorescence microscopy. Expression of Ras, MAPKs (ERK, JNK and p38) and Akt/mTOR pathways were evaluated by WB. Key findings: Linalool (0–2.5 mM) dose‐dependently inhibited cell proliferation by inducing G0/G1 cell cycle arrest, through Cdk4 and cyclin A downregulation, p21 and p27 upregulation, and apoptosis, characterized by MMP loss, caspase‐3 activation, PARP cleavage and DNA fragmentation. Low concentrations of linalool (1.0 mM) reduced membrane‐bound Ras and Akt activity whereas higher amounts (2.0 mM) triggered mTOR inhibition and ROS generation, in correlation with MAPKs activation and Akt phosphorylation. ROS scavenger N‐acetyl‐L‐cysteine partially rescued HepG2 cell growth and prevented MPP depolarization, ERK and JNK activation. Moreover, specific ERK and Akt phosphorylation inhibitors potentiated linalool anti‐cancer activity, pointing Akt and ERK activation as pro‐survival mechanisms in response to higher concentrations of linalool. Significance: This report reveals that linalool induces G0/G1 arrest and apoptosis in HepG2 cells involving Ras, MAPKs and Akt/mTOR pathways and suggests that linalool is a promising anticancer agent for HCC therapy. Graphical abstract: Figure. No caption available.

Citrus reticulata peel oil inhibits non-small cell lung cancer cell proliferation in culture and implanted in nude mice

Non-small cell lung cancer (NSCLC) accounts for most cases of lung cancer. The peel oil of mandarin Citrus reticulata Blanco cv. Dancy (MPO) is a natural source of essential oils and carotenoids. Volatile and non-volatile lipid compounds were characterized by chromatographic methods. We demonstrate that MPO causes a dose-dependent growth inhibition of NSCLC model cells (A549) in culture and tumour growth in vivo of the same cells implanted in nude mice fed with MPO-supplemented diets. MPO induced cell cycle arrest mainly at the G0/G1 phase and reduced the amount of membrane-bound Ras protein along with apoptosis induction. No toxicological effect was found in liver parameters analysed in treated mice and histopathological analyses of their organs did not show any morphological changes. In conclusion, the data suggest that MPO possesses significant antitumor activity without causing systemic toxicity, proposing it as a dietary supplement that may be helpful in cancer prevention.

Cytotoxic effects of essential oils from four Lippia alba chemotypes in human liver and lung cancer cell lines

Abstract Essential oils (EOs) from aromatic plants contain molecules that can interfere with diseases such as cancer and are considered attractive because of their widespread use, good bioavailability, low toxicity and affordable cost. EOs from Lippia alba (LaEOs) manifest intraspecific chemical differences in its composition – defined as chemotypes – and is notable for the chemical diversity of their volatile secondary metabolites. We evaluated LaEOs chemotypes cytotoxicity on human cancer culture cells and investigated the mechanisms involved in tagetenone (ta) chemotype cytotoxicity. It exhibited selective cytotoxicity against HepG2 and A549 cells. The mechanism involved cell cycle arrest and apoptosis induction. Tagetenone chemotype (LaEOta) treatment caused 3-hydroxy-3-methylglutaryl-coenzyme A reductase decrease and profound cholesterogenesis inhibition with farnesyl pyrophosphate redirection towards other end products, such as ubiquinone. This work contributes to a clearer understanding of mechanisms of action of LaEOta, thus suggesting that the use of that tagetenone chemotype could provide significant health benefits as a chemopreventive and/or chemotherapeutic agent.