Neuropeptide Y at the cellular level – Studies on PreproNPY L7P Polymorphism and the Mitochondrial Form of NPY

Neuropeptide Y (NPY) is a widely expressed neurotransmitter in the central and peripheral nervous systems. Thymidine 1128 to cytocine substitution in the signal sequence of the preproNPY results in a single amino acid change where leucine is changed to proline. This L7P change leads to a conformational change of the signal sequence which can have an effect on the intracellular processing of NPY. The L7P polymorphism was originally associated with higher total and LDL cholesterol levels in obese subjects. It has also been associated with several other physiological and pathophysiological responses such as atherosclerosis and T2 diabetes. However, the changes on the cellular level due to the preproNPY signal sequence L7P polymorphism were not known. The aims of the current thesis were to study the effects of the [p.L7]+[p.L7] and the [p.L7]+[p.P7] genotypes in primary cultured and genotyped human umbilical vein endothelial cells (HUVEC), in neuroblastoma (SK-N-BE(2)) cells and in fibroblast (CHO-K1) cells. Also, the putative effects of the L7P polymorphism on proliferation, apoptosis and LDL and nitric oxide metabolism were investigated. In the course of the studies a fragment of NPY targeted to mitochondria was found. With the putative mitochondrial NPY fragment the aim was to study the translational preferences and the mobility of the protein. The intracellular distribution of NPY between the [p.L7]+[p.L7] and the [p.L7]+[p.P7] genotypes was found to be different. NPY immunoreactivity was prominent in the [p.L7]+[p.P7] cells while the proNPY immunoreactivity was prominent in the [p.L7]+[p.L7] genotype cells. In the proliferation experiments there was a difference in the [p.L7]+[p.L7] genotype cells between early and late passage (aged) cells; the proliferation was raised in the aged cells. NPY increased the growth of the cells with the [p.L7]+[p.P7] genotype. Apoptosis did not seem to differ between the genotypes, but in the aged cells with the [p.L7]+[p.L7] genotype, LDL uptake was found to be elevated. Furthermore, the genotype seemed to have a strong effect on the nitric oxide metabolism. The results indicated that the mobility of NPY protein inside the cells was increased within the P7 containing constructs. The existence of the mitochondria targeted NPY fragment was verified, and translational preferences were proved to be due to the origin of the cells. Cell of neuronal origin preferred the translation of mature NPY (NPY1-36) when compared to the non neuronal cells that translated both, NPY and the mitochondrial fragment of NPY. The mobility of the mitochondrial fragment was found to be minimal. The functionality of the mitochondrial NPY fragment remains to be investigated. L7P polymorphism in the preproNPY causes a series of intracellular changes. These changes may contribute to the state of cellular senescence, vascular tone and lead to endothelial dysfunction and even to increased susceptibility to diseases, like atherosclerosis and T2 diabetes.

Molecular Interactions of Prodiginines with the BH3 Domain of Anti-Apoptotic Bcl-2 Family Members.

Prodigiosin and obatoclax, members of the prodiginines family, are small molecules with anti-cancer properties that are currently under preclinical and clinical trials. The molecular target(s) of these agents, however, is an open question. Combining experimental and computational techniques we find that prodigiosin binds to the BH3 domain in some BCL-2 protein families, which play an important role in the apoptotic programmed cell death. In particular, our results indicate a large affinity of prodigiosin for MCL-1, an anti-apoptotic member of the BCL-2 family. In melanoma cells, we demonstrate that prodigiosin activates the mitochondrial apoptotic pathway by disrupting MCL-1/BAK complexes. Computer simulations with the PELE software allow the description of the induced fit process, obtaining a detailed atomic view of the molecular interactions. These results provide new data to understand the mechanism of action of these molecules, and assist in the development of more specific inhibitors of anti-apoptotic BCL-2 proteins.

Building complexity in O2-binding copper complexes : site-selective metalation and intermolecular O2-binding at dicopper and heterometallic complexes derived from an unsymmetric ligand

A novel unsymmetric dinucleating ligand (LN3N4) combining a tridentate and a tetradentate binding sites linked through a m-xylyl spacer was synthesized as ligand scaffold for preparing homo- and dimetallic complexes, where the two metal ions are bound in two different coordination environments. Site-selective binding of different metal ions is demonstrated. LN3N4 is able to discriminate between CuI and a complementary metal (M′ = CuI, ZnII, FeII, CuII, or GaIII) so that pure heterodimetallic complexes with a general formula [CuIM′(LN3N4)]n+ are synthesized. Reaction of the dicopper(I) complex [CuI 2(LN3N4)]2+ with O2 leads to the formation of two different copper-dioxygen (Cu2O2) intermolecular species (O and TP) between two copper atoms located in the same site from different complex molecules. Taking advantage of this feature, reaction of the heterodimetallic complexes [CuM′(LN3N4)]n+ with O2 at low temperature is used as a tool to determine the final position of the CuI center in the system because only one of the two Cu2O2 species is formed

Mitochondrial fatty acid oxidation in obesity

Significance: Current lifestyles with high-energy diets and little exercise are triggering an alarming growth in obesity. Excess of adiposity is leading to severe increases in associated pathologies, such as insulin resistance, type 2 diabetes, atherosclerosis, cancer, arthritis, asthma, and hypertension. This, together with the lack of efficient obesity drugs, is the driving force behind much research. Recent Advances: Traditional anti-obesity strategies focused on reducing food intake and increasing physical activity. However, recent results suggest that enhancing cellular energy expenditure may be an attractive alternative therapy. Critical Issues: This review evaluates recent discoveries regarding mitochondrial fatty acid oxidation (FAO) and its potential as a therapy for obesity. We focus on the still controversial beneficial effects of increased FAO in liver and muscle, recent studies on how to potentiate adipose tissue energy expenditure, and the different hypotheses involving FAO and the reactive oxygen species production in the hypothalamic control of food intake. Future Directions: The present review aims to provide an overview of novel anti-obesity strategies that target mitochondrial FAO and that will definitively be of high interest in the future research to fight against obesity-related disorders.

Synthesis, structure and physicochemical properties of zinc and copper complexes based on sulfonamides containing 8-aminoquinoline ligands

Sulfonamides obtained by reaction of 8-aminoquinoline with 4-nitrobenzenesulfonylchloride and 2,4,6-triisopropylbenzenesulfonyl chloride were used to synthesize coordination compounds with CuII and ZnII with a ML2 composition. Determination of the crystal structures of the resulting zinc and copper complexes by X-ray diffraction show a distorted tetrahedral environment for the [Cu(qnbsa)2], [Cu(qibsa)2] and [Zn(qibsa)2] complexes in which the sulfonamide group acts as a bidentate ligand through the nitrogen atoms from the sulfonamidate and quinoline groups. The complex [Zn(qnbsa)2] crystallizes with a water molecule from the solvent and the Zn is five-coordinated and shows a bipyramidal-trigonal geometry. The electrochemical and electronic spectroscopy properties of the copper complexes are also discussed.

Improvement of genistein content in solid genistein/-cyclodextrin complexes β

Genistein:β-cyclodextrin complexes with high drug loading (19.22%) were prepared by freeze-drying and characterized by differential scanning calorimetry and hydrogen nuclear magnetic resonance spectroscopy. The spatial configuration of the complex was proposed by means of 2D-NOESY experiment combined with molecular modeling. According to the results obtained, the interaction of genistein with β -cyclodextrin in a 1:1 complex is supposed to occur mainly through the insertion of the guest A-ring in cyclodextrin cavity, without rule out the possibility of inclusion through the B-ring, as previously reported in the literature.

Spectroscopic characterization of schiff base-copper complexes immobilized in smectite clays

Herein, the immobilization of some Schiff base-copper(II) complexes in smectite clays is described as a strategy for the heterogenization of homogeneous catalysts. The obtained materials were characterized by spectroscopic techniques, mostly UV/Vis, EPR, XANES and luminescence spectroscopy. SWy-2 and synthetic Laponite clays were used for the immobilization of two different complexes that have previously shown catalytic activity in the dismutation of superoxide radicals, and disproportionation of hydrogen peroxide. The obtained results indicated the occurrence of an intriguing intramolecular redox process involving copper and the imine ligand at the surface of the clays. These studies are supported by computational calculations.

Activity and sulfur resistance of Rh(I) and Pd(II) complexes

Two complexes of Rh(I) and Pd(II) with chloride and tridecylamine ligands were obtained and characterized by Elementary Analysis and by XPS and FTIR spectroscopies. Complexes anchored on γ-Al2O3 were tested in the styrene semi-hydrogenation reaction carried out in the absence or presence of a sulfur poison. Although both low loaded catalysts were highly selective, the Pd(II) complex was three times more active than the Rh(I) complex. The rhodium complex was more sulfur resistant but less active than the palladium complex. Differences in conversion and sulfur resistance between both complexes could be related to electronic and/or geometric effects.

Improving the solubility of the antichagasic drug benznidazole through formation of inclusion complexes with cyclodextrins

This study describes unpublished research on improving the solubility of benznidazole by the formation of an inclusion complex. The cyclodextrins selected were αCD, βCD, γCD, HPβCD, RMβCD and SBβCD. All complexes were obtained in solution, presenting 1:1 stoichiometry according to the phase solubility diagram. The highest association constants were obtained with RMβCD and SBβCD, being selected for attainment of solid state complexes. These were characterized using XRD, SEM and dissolution test. The data obtained suggest the formation of complexes and indicate that these may provide a promising alternative way of developing solid doses of drug with suitable biopharmaceutical properties.

Photochemical characteristics of diclofenac and its photodegradation of inclusion complexes with β-cyclodextrins

Diclofenac is one of most frequently detected compounds in the water cycle. In this work, the effect of initial concentration, liquid inclusion complexes with β-Cyclodextrins (β-CDs) on the photodegradation of diclofenac were studied. Six phototransformation products were detected by HPLC chromatograms. UV-absorption spectra of diclofenac and phototransformation products were determined. One of the phototransformation products was identified. The degradation followed pseudo-first-order kinetics. The experiment showed that irradiation of diclofenac in the presence of β-CDs increase photodegradation rate and determined the optimal molar ratio of diclofenac to β-CDs as 1:2. The reduced photohaemolytic activity of diclofenac in the presence of β-CDs may be attributed to the sequestering and stabilizing of the radical intermediates and /or photoproducts by complexation.

Synthesis and characterization of Fe(III)-piperazine-derived complexes encapsulated in zeolite Y

Zeolite-encapsulated complexes have been widely applied in hydrocarbon oxidation catalysis. The "ship-in-a-bottle" encapsulation of iron(III) complexes containing piperazine and piperazine-derivative ligands in zeolite-Y is described. The flexible ligand methodology was employed and the efficiency and reproducibility of the procedure was investigated. The catalysts were characterized employing several techniques and the results indicate the presence of coordinated and uncoordinated iron(III) ions inside and outside the zeolitic cage.

Ni(II), Cu(II), AND Zn(II) COMPLEXES DERIVED FROM A NEW SCHIFF BASE 2-((Z)-(3-METHYLPYRIDIN-2- YLEIMINO)METHYL)PHENOL AND SYNTHESIS OF NANO SIZED METAL OXIDE PARTICLES FROM THESE COMPOUNDS

Synthesis, spectral identification, and magnetic properties of three complexes of Ni(II), Cu(II), and Zn(II) are described. All three compounds have the general formula [M(L)2(H2O)2], where L = deprotonated phenol in the Schiff base 2-((z)-(3-methylpyridin-2-yleimino)methyl)phenol. The three complexes were synthesized in a one-step synthesis and characterized by elemental analysis, Fourier transform infrared spectroscopy, electronic spectra, X-ray diffraction (XRD), and room temperature magnetic moments. The Cu(II) and Ni(II) complexes exhibited room temperature magnetic moments of 1.85 B.M. per copper atom and 2.96 B.M. per nickel atom. The X-band electron spin resonance spectra of a Cu(II) sample in dimethylformamide frozen at 77 K (liquid nitrogen temperature) showed a typical ΔMS = ± 1 transition. The complexes ([M(L)2(H2O)2]) were investigated by the cyclic voltammetry technique, which provided information regarding the electrochemical mechanism of redox behavior of the compounds. Thermal decomposition of the complexes at 750 ºC resulted in the formation of metal oxide nanoparticles. XRD analyses indicated that the nanoparticles had a high degree of crystallinity. The average sizes of the nanoparticles were found to be approximately 54.3, 30.1, and 44.4 nm for NiO, CuO, and ZnO, respectively.

Thermoanalytical study of the complexes of 4-dimetilaminocinnamylidenepiruvate with manganese (II), cobalt (II), nickel (II), cupper (II), zinc (II) and lead (II), in the solid state

Solid state compounds M-4-DMCP, where 4-DMCP is 4-dimethylaminocinnamylidenepyruvate and M represents Mn (II), Co (II), Ni (II), Cu (II), Zn (II) and Pb (II) were prepared. These compounds were studied by thermoanalitycal techniques: thermogravimetry (TG), derivative thermogravimetry (DTG), differential scanning calorimetry (DSC), X-ray diffraction powder patterns and complexometric titration with EDTA. From the results obtained by the complexometric titration with EDTA, TG, DTG and DSC curves, was possible to establish the hydration degree, stoichiometry and thermal stability of the prepared compounds.

Thermal degradation behaviour of some polydithiooxamide metal complexes

The thermal decomposition behavior of the Fe(II), Co(II), Ni(II) and Zn(II) complexes of polydithiooxamide has been investigated by thermogravimetric analysis (TGA) at a heating rate of 20°C min-1 under nitrogen. The Coats-Redfern integral method is used to evaluate the kinetic parameters for the successive steps in the decomposition sequence observed in the TGA curves. The processes of thermal decomposition taking place in the four complexes are studied comparatively as the TGA curves indicate the difference in the thermal decomposition behavior of these complexes. The thermal stabilities of these complexes are discussed in terms of repulsion among electron pairs in the valence shell of the central ion and electronegativity effects.

Thermal behaviour of TEGMMA copolymers obtained by photopolymerization using iron complexes

Copolymers of methyl methacrylate (MMA) and triethyleneglycol dimethacrylate (TEGDMA) obtained by photoinitiated polymerization using Fe(III) complexes were submitted to thermogravimetry (TGA) under dynamic air atmosphere and N2, and differential scanning calorimetric analysis (DSC). Thermal events were observed only between 90 - 110 ºC. Glass transitions were observed at ca. 100 ºC, followed by an exothermic peak at 170 ºC. The exothermic peak was assigned to a thermal curing process due to the presence of unreacted vinyl groups of the monomers. DSC revealed to be a useful tool to evaluate the curing completeness in this kind of material, using small amounts of sample in relatively short time.