The effect of 2,4,6-trimethyl-N-(meta-3-trifluoromethyl-phenyl)- benzenesulfonamide (m-3M3FBS), a presumed phospholipase C activator, on cytosolic free Ca2+ concentrations ([Ca2+]i) in Madin Darby canine kidney (MDCK) cells and OC2 human oral cancer cells was unclear. This study explored whether m-3M3FBS changed basal [Ca2+]i levels in suspended MDCK and OC2 cells by using fura-2 as a Ca2+-sensitive fluorescent dye. m-3M3FBS at concentrations between 0.1-20 £gM increased [Ca2+]i in a concentration-dependent manner in MDCK cells, however in OC2 cells, m-3M3FBS at concentrations between 10-60 £gM increased [Ca2+]i in a concentration-dependent manner. The Ca2+ signals were reduced partly by removing extracellular Ca2+ in the two cell types. m-3M3FBS-induced Ca2+ influx was inhibited by the store-operated Ca2+ channel blockers nifedipine, econazole and SK&F96365, and by the phospholipase A2 inhibitor aristolochic acid. In Ca2+-free medium, m-3M3FBS pretreatment abolished the [Ca2+]i rise induced by the endoplasmic reticulum Ca2+ pump inhibitors thapsigargin, cyclopiazonic acid or 2,5-di-tert-butylhydroquinone (BHQ). Conversely, pretreatment with thapsigargin, cyclopiazonic acid or BHQ partly reduced m-3M3FBS-induced [Ca2+]i rise. Inhibition of phospholipase C with U73122 did not alter m-3M3FBS-induced [Ca2+]i rise. Collectively, in MDCK and OC2 cells, m-3M3FBS induced [Ca2+]i rises by causing phospholipase C-independent Ca2+ release from the endoplasmic reticulum and Ca2+ influx via store-operated Ca2+ channels and other unidentified Ca2+ channels. Additionally, 5-100 £gM of m-3M3FBS killed cells in a concentration-dependent manner in OC2 cells. The cytotoxic effect of m-3M3FBS was not reversed by prechelating cytosolic Ca2+ with 1,2-bis(2-aminophenoxy)ethane- N,N,N',N'-tetraacetic acid (BAPTA). Propidium iodide staining data suggest that m-3M3FBS (20 or 50 £gM) induced apoptosis in a Ca2+-independent manner. We were also interested in knowing whether BAPTA suppressed cell death during oxidative stress in MDCK cells. BAPTA loading altered tBHP (tert-butyl hydroperoxide) and H2O2-induced cell death in a concentration-dependent manner. This suggests that the cell death induced by tBHP and H2O2 appears to be Ca2+-dependent in MDCK cells. The tBHP and H2O2-induced cell death was not suppressed by 2 £gM U73122 (PLC inhibitor), 50 £gM zVAD-fmk (caspase inhibitor), 2 £gM cyclosporin A (a potent inhibitor of the MPTP), 20 £gM PD98059 (ERK inhibitor) or 2 £gM SP600125 (JNK inhibitor). This suggests that the tBHP and H2O2-induced MDCK cells death was not via the PLC, MPTP, caspase, ERK or JNK pathways. Propidium iodide staining, caspase-3 activity assay and cell morphology data suggest that tBHP and H2O2-induced cell death was necrosis, not via apoptosis, and the cell death appears to be caspase-independent and Ca2+-dependent. The effect of the antidepressant paroxetine on [Ca2+]i in OC2 human oral cancer cells is unclear. This study also explored whether paroxetine changed basal [Ca2+]i levels in suspended OC2 cells by using fura-2 as a Ca2+-sensitive fluorescent dye. Paroxetine at concentrations between 100-1000 £gM increased [Ca2+]i in a concentration-dependent manner. The Ca2+ signal was reduced by 50% by removing extracellular Ca2+. Paroxetine-induced Ca2+ influx was inhibited by the store-operated Ca2+ channel blockers nifedipine, econazole and SK&F96365, the phospholipase A2 inhibitor aristolochic acid, and protein kinase C modulators. In Ca2+-free medium, pretreatment with the endoplasmic reticulum Ca2+ pump inhibitor thapsigargin abolished paroxetine¡Vinduced [Ca2+]i rise. Inhibition of PLC with U73122 did not alter paroxetine-induced [Ca2+]i rise. Paroxetine at 10-50 £gM induced cell death in a concentration-dependent manner. The death was not reversed when cytosolic Ca2+ was chelated with BAPTA. Propidium iodide staining suggests that apoptosis played a role in the death. Collectively, in OC2 cells, paroxetine induced [Ca2+]i rise by causing PLC-independent Ca2+ release from the endoplasmic reticulum and Ca2+ influx via store-operated Ca2+ channels in a manner regulated by protein kinase C and phospholipase A2. Paroxetine also induced cell death in a Ca2+-independent manner.

Submitted by José Henrique Lazaro de Carvalho (jose.carvalho@fc.unesp.br) on 2018-04-26T14:52:46Z No. of bitstreams: 1 merged.pdf: 3543516 bytes, checksum: 94a6da42fee42b5469644d670ca0dddc (MD5) / Approved for entry into archive by Minervina Teixeira Lopes null (vina_lopes@bauru.unesp.br) on 2018-04-26T17:33:49Z (GMT) No. of bitstreams: 1 carvalho_jhl_me_bauru.pdf: 3133645 bytes, checksum: 07b206851bb163d2aab65e55f0340aea (MD5) / Made available in DSpace on 2018-04-26T17:33:49Z (GMT). No. of bitstreams: 1 carvalho_jhl_me_bauru.pdf: 3133645 bytes, checksum: 07b206851bb163d2aab65e55f0340aea (MD5) Previous issue date: 2018-02-26 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Os hidrocarbonetos se constituem na função mais simples da química orgânica. Mesmo sendo formados apenas por carbono e hidrogênio, esta classe é uma das mais diversas, onde se destacam os hidrocarbonetos alquil aromáticos (HAA) e os hidrocarbonetos aromáticos policíclicos (HAP). Por possuírem baixa reatividade e serem lipossolúveis, os HAA e HAP tendem a se acumular na natureza e nos tecidos vivos podendo levar a mutações genéticas e ao câncer. Funcionalizações oxigenadas nestes hidrocarbonetos ampliam suas aplicações industriais e auxiliam na sua remediação ambiental. A oxidação radicalar utilizando radicais hidroxila via TBHP (t-butil-hidroperóxido) é uma rota promissora devido à ausência de resíduos metálicos nocivos no processo. Paralelamente, o uso de metais do bloco-d, como o nióbio, como catalisadores heterogêneos, que são conhecidos por apresentarem boa estabilidade térmica, porosidade, maior superfície de contato e podendo ser recuperado e reutilizado, vem sendo descrito como bons catalisadores para a reação, permitindo rendimentos maiores e condições reacionais mais brandas. O nióbio tem despertado o interesse de muitos grupos de pesquisa devido às suas propriedades e aplicabilidades na metalurgia, supercondutores e supercapacitores, e na síntese orgânica. Este trabalho focou no estudo da aplicação do óxido de nióbio como catalisador heterogêneo em reações de oxidação de HAA e HAP usando o TBHP como oxidante. Diferentes variáveis reacionais como o solvente, a concentração e morfologia do catalisador, temperatura, dentre outras, foram investigadas a fim de otimizar a reação para a melhor conversão no derivado oxigenado. A melhor condição obtida foi utilizando Nb2O5 amorfo em 10% em mol de reagente, em acetonitrila e em refluxo com mais de 40 % de conversão em derivado oxigenado para qualquer substrato de hidrocarboneto estudado. / Hydrocarbons are the simplest function of organic chemistry. Although formed only by carbon and hydrogen, this class is one of the most diverse, in which the alkyl aromatic hydrocarbons (AAHs) and the polycyclic aromatic hydrocarbons (PAHs) stand out. These substances are mostly fossil raw materials widely used in industry in diverse applications. Because they have low reactivity and are liposoluble, AAHs and PAHs tend to accumulate in nature and in living tissues, leading to genetic mutations and cancer. Oxygenated functionalities in these hydrocarbons broaden their industrial applications and assist in their environmental remediation. Radical oxidation using hydroxyl radicals via TBHP (t-butylhydroperoxide) is a promising route due to the absence of noxious metal residues in the process. At the same time, the use of d-block metals, such as niobium, as heterogeneous catalysts, which are known to have good thermal stability, porosity, larger contact surface and be able to be recovered and reused, have been described as good catalysts for the reaction, allowing for higher yields and softer reaction conditions. Niobium has attracted the interest of many research groups because of its properties and applications in metallurgy, superconductors and supercapacitors, and in organic synthesis. This work focused on the study of the application of niobium oxide as a heterogeneous catalyst in oxidation reactions of AAHs and HAP using TBHP as oxidant. Many reactional variables like the solvent, the catalyst concentration and morphology, as well as temperature have been investigated in order to optimize the reaction for the better conversion to oxygen derivatives. The best obtained condition was using 10 % in mol amorphous Nb2O5 in acetonitrile under reflux with a conversion yield of more than 40 % for any studied hydrocarbon substrate.

>Magister Scientiae - MSc / In this thesis we describe the synthesis of several carbonyl complexes of molybdenum and tungsten, compounds (Cl-ClO). The compounds Cl- C4 are zero valent carbonyl complexes containing N-base ligands prepared by following a common synthetic procedure. Compounds Cl and C2 were metal pentacarbonyl of 3-(1-methylpyrrolidin-2-yl) pyridine while C3 and C4 are metal tetracarbonyl complexes of 3, 5- dimethylpyrazole, (M=Mo, W). The compounds C5-C10 are divalent metal carbonyl complexes. Compounds CS and C6 were 3,5-dimethylpyrazole dibromotricarbonyl metal complexes prepared from the dibromotetracarbonyl metal dimers at room temperature while the compounds C7 and C8 were cyclopentadienyl halogenoaryltricarbonyl complexes prepared from the cyclopentadienyl metal dimers. Compounds C9 and ClO were prepared from cyclopentadienyl metal dimers by reacting the [CpM(C0)3r anion with CCl4 to obtain [CpM(C0)3Cl] and further reacted with 3-(1- methylpyrrolidin-2-yl) pyridine. All the compounds, Cl-ClO, were characterized by the standard analytical techniques such as FTIR, 1H, 13C NMR; and UV-Vis spectroscopy. Compound C4 was characterized by X-ray crystallography. The structure is depicted as having a distorted octahedral geometry around the metal centre. The compounds Cl-ClO were then tested towards the epoxidation of selected cyclic and straight chain alkenes. The substrates used were cis-cyclooctene (Cyg), 1-octene (C8) cyclohexene (Cy6), 1-hexene (C6) and styrene (Sty). The epoxidation reactions were carried out at a temperature of 55 °C using tertbutylhydroperoxide (TBHP) as the oxidant and dichloroethane (DCE) as the solvent. The metal carbonyl complexes were pre-activated by first reacting them with the oxidant TBHP to obtain the metal-oxo complexes which are the active compounds for epoxidation reactions. The products were analyzed using GC techniques. The compounds, Cl-ClO showed a promising activity towards epoxidation reactions owing to the high conversions obtained by these compounds. For example, conversions of 81% (1-octene), 90% (cis-cyclooctene) were obtained by compound C5, 87% (cis-cyclooctene-compound C3, 95% (cis-cyclooctene-compound · C7) and 69% (ciscyclooctene- compound C4) for an average period of 24 h. The divalent metal carbonyl complexes showed a higher activity but with poor selectivity towards the expected epoxides compared to the zero valent metal carbonyl complexes.

Yes / We investigated the protective role of myricetin bulk and nanoforms, against reactive oxygen species (ROS)-induced oxidative stress caused by hydrogen peroxide and tertiary-butyl hydro peroxide in lymphocytes in vitro from healthy individuals and those from pre-cancerous patients suffering with monoclonal gammopathy of undetermined significance (MGUS). The change in intracellular reactive oxygen species was measured once cells were treated with myricetin bulk forms and nanoforms with and without either hydrogen peroxide or tertiary-butyl hydro peroxide co-supplementation. The direct and indirect antioxidant activity of myricetin was spectrofluometrically measured using the fluorescent dye 2',7'-dichlorofluorescin diacetate and using the Comet assay, respectively. Hydrogen peroxide (50 µM) and tertiary-butyl hydro peroxide (300 µM) induced a higher level of reactive oxygen species-related DNA damage and strand breaks. Addition of myricetin nanoform (20 µM) and bulk (10 µM) form could, however, significantly prevent hydrogen peroxide- and tertiary-butyl hydro peroxide-induced oxidative imbalances and the nanoform was more effective. Glutathione levels were also quantified using a non-fluorescent dye. Results suggest that myricetin treatment had no significant effect on the cellular antioxidant enzyme, glutathione. The current study also investigates the effect of myricetin on the induction of double-strand breaks by staining the gamma-H2AX foci immunocytochemically. It was observed that myricetin does not induce double-strand breaks at basal levels rather demonstrated a protective effect.

Morphine is a natural opioid which is used in medicine due to his potent analgesic and sedative effects. In the forefront of scientific interest is a chronic usage of opioids which can lead to a development of drug addiction. Morphine role in oxidative stress was described in last years. It was revealed its protective potencial by many studies. However, some studies described its pro-oxidative effect. The aim of this study was to determinate effect of chronic morphine on cell survival after oxidative stress caused by H202 analog - tBHP in the SH-SY5Y neuroblastoma cell line. The results verified morphine protective effect against oxidative stress. The highest protective effect of morphine was achieved in a concetration of 10 µM. It was desribed that morphine can induce activation of mu-opioid (MOR) and Toll-like 4 (TLR4) receptors signalling pathway on molecular level. The aim of this thesis was to evaluate the role of MOR a TLR4 in protective effect of morphine against oxidative stress by two methods. Firstly, it was used tests of oxidative stress on cell viability. The obtained results demonstrated majority role of TLR4 and minory role of MOR. Afterwards, we assesed changes in the expression of MOR a TLR4 after chronic morphine by SDS-PAGE electrophoresis. Results of these experiments did not...