Utrophin Up-Regulation by an Artificial Transcription Factor in Transgenic Mice

Duchenne Muscular Dystrophy (DMD) is a severe muscle degenerative disease, due to absence of dystrophin. There is currently no effective treatment for DMD. Our aim is to up-regulate the expression level of the dystrophin related gene utrophin in DMD, complementing in this way the lack of dystrophin functions. To this end we designed and engineered several synthetic zinc finger based transcription factors. In particular, we have previously shown that the artificial three zinc finger protein named Jazz, fused with the appropriate effector domain, is able to drive the transcription of a test gene from the utrophin promoter “A”. Here we report on the characterization of Vp16-Jazz-transgenic mice that specifically over-express the utrophin gene at the muscular level. A Chromatin Immunoprecipitation assay (ChIP) demonstrated the effective access/binding of the Jazz protein to active chromatin in mouse muscle and Vp16-Jazz was shown to be able to up-regulate endogenous utrophin gene expression by immunohistochemistry, western blot analyses and real-time PCR. To our knowledge, this is the first example of a transgenic mouse expressing an artificial gene coding for a zinc finger based transcription factor. The achievement of Vp16-Jazz transgenic mice validates the strategy of transcriptional targeting of endogenous genes and could represent an exclusive animal model for use in drug discovery and therapeutics

Disruption of PTH Receptor 1 in T Cells Protects against PTH-Induced Bone Loss

Hyperparathyroidism in humans and continuous parathyroid hormone (cPTH) treatment in mice cause bone loss by regulating the production of RANKL and OPG by stromal cells (SCs) and osteoblasts (OBs). Recently, it has been reported that T cells are required for cPTH to induce bone loss as the binding of the T cell costimulatory molecule CD40L to SC receptor CD40 augments SC sensitivity to cPTH. However it is unknown whether direct PTH stimulation of T cells is required for cPTH to induce bone loss, and whether T cells contribute to the bone catabolic activity of PTH with mechanisms other than induction of CD40 signaling in SCs.Here we show that silencing of PTH receptor 1 (PPR) in T cells blocks the bone loss and the osteoclastic expansion induced by cPTH, thus demonstrating that PPR signaling in T cells is central for PTH-induced reduction of bone mass. Mechanistic studies revealed that PTH activation of the T cell PPR stimulates T cell production of the osteoclastogenic cytokine tumor necrosis factor alpha (TNF). Attesting to the relevance of this effect, disruption of T cell TNF production prevents PTH-induced bone loss. We also show that a novel mechanism by which TNF mediates PTH induced osteoclast formation is upregulation of CD40 expression in SCs, which increases their RANKL/OPG production ratio.These findings demonstrate that PPR signaling in T cells plays an essential role in PTH induced bone loss by promoting T cell production of TNF. A previously unknown effect of TNF is to increase SC expression of CD40, which in turn increases SC osteoclastogenic activity by upregulating their RANKL/OPG production ratio. PPR-dependent stimulation of TNF production by T cells and the resulting TNF regulation of CD40 signaling in SCs are potential new therapeutic targets for the bone loss of hyperparathyroidism

Chemical, spectroscopic characterization and antibacterial activities in vitro of a novel gold(I)-ibuprofen complex

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)A novel gold(I) complex with ibuprofen was synthesized and characterized by chemical and spectroscopic measurements. Elemental analysis led to the composition AuC(14)H(18)O(2)N. Infrared, (1)H and (13)C NMR data suggest coordination of the ligand to Au(I) through the oxygen atom of the carboxylic group in a monodentate form. An antibiotic sensitive profile indicated antibacterial activity in vitro of the complex against Gram-negative (E. coli and P. aeruginosa) and Gram-positive (S. aureus) microorganisms. (C) 2011 Elsevier B.V. All rights reserved.145738740Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)FAPESP [2010/00667-7, 2006/55367-2

Studies of the tautomeric equilibrium of 1,3-thiazolidine-2-thione: Theoretical and experimental approaches

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)The tautomeric equilibrium of the thione/thiol forms of 1,3-thiazolidine-2-thione was studied by nuclear magnetic resonance, infrared and ultraviolet-visible spectroscopies. Density functional theory was used to support the experimental data and indicates the predominance of the thione tautomer in the solid state, being in agreement with previously reported crystallographic data. In solution, the tautomeric equilibrium was evaluated using H-1 NMR at different temperatures in four deuterated solvents acetonitrile, dimethylsulfoxide, chloroform and methanol. The equilibrium constants, K = (thiol)/(thione), and free Gibbs energies were obtained by integration of N bonded hydrogen signals at each temperature for each solvent, excluding methanol. The endothermic tautomerization is entropy-driven and the combined effect of solvent and temperature can be used to achieve almost 50% thiol concentrations in solution. The nature of the electronic transitions was investigated theoretically and the assignment of the bands was made using time-dependent DFT as well as the influence of solvent on the energy of the most important bands of the spectra. (C) 2012 Elsevier B.V. All rights reserved.4086268Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Characterization, and crystal structure of thiophenyl-2-methylidene-2-aminophenol

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)The Schiff base thiophenyl-2-methylidene-2-aminophenol (ImineOH) is obtained from a stoichiometric mixture of 2-thiophenecarboxaldehyde and 2-aminophenol in ethanol under reflux at 90A degrees C. Its crystal structure is determined by single crystal X-ray diffraction. ImineOH packs in an orthorhombic unit cell in the Pbca space group with the unit cell parameters a = 16.942(4) , b = 13.4395(11) , and c = 17.5857(12) , V = 4004.1(10) (3), Z = 16. Strong hydrogen bonds are present in the ImineOH structure. Apart from the X-ray study, ImineOH was characterized by elemental analysis (CHN-S) and FT-IR (4000 cm(-1) to 400 cm(-1)), UV-Vis and C-13, H-1, and N-15 NMR spectroscopic measurements.541269273Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPEMIG [CEX-APQ-00173/09]CNPq [472468/2010-3

A simple method to synthesize fluorescent modified gold nanoparticles using tryptamine as the reducing and capping agent

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)A simple method to synthesize fluorescent modified gold nanoparticles using tryptamine as the reducing and capping agent is described. The presented method produces gold nanoparticles with 36.65 +/- 5.30 nm average size. The modified gold nanoparticles were characterized by elemental and thermal analyses, dynamic light scattering, transmission electron microscopy, zeta potential, X-ray powder diffraction and spectroscopic techniques, such as electronic spectroscopy in ultraviolet-visible and fluorescence excitation-emission. In addition, modified gold nanoparticles were analyzed by solid state N-15 nuclear magnetic resonance spectroscopy, which confirmed the coordination of tryptamine on the gold nanoparticles surface. A prominent characteristic observed is the fluorescence of tryptamine which was not quenched after the coordination to gold nanoparticles. The results presented in this paper confirm the modification of gold nanoparticles by tryptamine and suggest potential use of such nanoparticles as labeling dye in biological systems. (C) 2013 Elsevier B.V. All rights reserved.1856165Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)LNNano (Laboratorio Nacional de Nanotecnologia, Campinas-SP) Brazilian AgencyFundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [Proc. 2012/08230-2

Chemical, spectroscopic characterization, and in vitro antibacterial studies of a new gold(I) complex with N-acetyl-L-cysteine

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)A new gold(I) complex with N-acetyl-L-cysteine was synthesized and characterized by chemical and spectroscopic techniques. The elemental and thermal analyses of the solid compound fit to the composition AuC5H8NO3S center dot 0.75H2O. Solid-state 13C-nuclear magnetic resonance (SSNMR) and infrared (IR) analyses indicate the coordination of the ligand to Au(I) through sulfur. The insolubility of the complex in both polar and non-polar solvents supports a polymeric structure. The antibacterial activity of the complex was evaluated by antibiogram assays using the disc diffusion method. The compound showed effective antibacterial activity against Staphylococcus aureus (Gram positive) and Escherichia coli (Gram negative) bacterial cells.63813901397Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [2008/54290-1, 2008/54291-8, 2006/55367-2

Silver(I) and gold(I) complexes with penicillamine: Synthesis, spectroscopic characterization and biological studies

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Two new metal complexes of penicillamine (Pen), with molecular formulas [Ag(C5H10O2NS)]center dot H2O and [Au(C5H10O2NS)], were synthesized by the reaction of the ligand with, respectively, silver nitrate and potassium dicyanoaurate(I) salts in aqueous solutions. Analytical characterizations of the complexes were based on elemental (C, H, N and S), thermal (TG-DTA) and electrospray ionization mass spectrometry (ESI-MS) analyses. Solid-state C-13 nuclear magnetic resonance (SSNMR) and infrared (IR) spectroscopic measurements indicate coordination of Pen to Ag(l) or Au(I) through the sulfur atom. Studies based on density functional theory (OFT) confirmed that Pen acts as an S-donor ligand to Au(I) or Ag(I). Thermal decompositions of the silver(I) and gold(I) complexes start at temperatures near 50 degrees C, leading to the formation of Ag-0 and Au-0 at 700 degrees C and 500 degrees C, respectively. Both complexes are just slightly soluble in common solvents such as water, methanol, ethanol, acetonitrile, hexane and dimethylsulfoxide. An antibiogram assay was performed by the disc diffusion method. The AgPen complex showed antibacterial activities against Escherichia coil and Pseudomonas aeruginosa bacterial strains. (C) 2012 Elsevier Ltd. All rights reserved.341210214Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [2006/55367-2]CNPq [472468/2010-3]FAPEMIG [CEX-APQ-00713/09

Synthesis, spectroscopic characterization and molecular modeling of a tetranuclear platinum(II) complex with thiazolidine-4-carboxylic acid

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)The synthesis, spectroscopic characterization and molecular modeling of a novel tetranuclear platinum(II) complex with thiazolidine-4-carboxylic acid (THC) are described. Elemental analysis is consistent with the composition PtCl2C4H7NO2S-H2O. Infrared (IR) spectroscopic results and solid-state C-13 and N-15 nuclear magnetic resonance (NMR) data indicate coordination of the ligand to Pt(II) through the nitrogen and sulfur atoms. The square planar geometry of the platinum(II) complex is completed by chlorine atoms. Density functional theory (OFF) suggests the formation of a tetrameric cluster as the most probable structure, where each THC molecule bridges between two metal centers. The compound is insoluble in water. (C) 2012 Elsevier B.V. All rights reserved.10192126Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)FAPESP [Proc. 2006/55367-2, 2008/54290-1, 2008/54291-8]CNPq [Proc. 479415/2009-9, 472468/2010-3