Development of a novel reporter gene vector for cell based angiogenic studies

Angiogenesis is a promising area of research that targets key therapeutic areas like cancer; wound healing, inflammatory diseases, etc. There is an increasing demand for screening of potential angiogenic and anti-angiogenic agents using sensitive, robust cell-based assays. We have developed a reporter vector containing cis-acting elements that respond to growth factors/angiogenic ligands for use in a cell-based luciferase reporter assay. We performed transient transfection of our reporter gene vector in MCF-7 cells to establish its application for screening of potential pro/anti-angiogenic agents. Reporter gene transactivation studies with different concentrations of fetal bovine serum clearly indicated that the vector is functionally responsive to the angiogenic signals mediated by serum growth factors. We also used endostatin to inhibit transactivation and prove responsiveness to the anti-angiogenic agent. This vector is a promising tool for studying angiogenesis using cell-based reporter gene assays.     

Reactivity of tris(acetylacetonato) iron(III) with tridentate [ONO] donor Schiff base as an access to newer mixed-ligand iron(III) complexes

Two new mixed-ligand iron(III) complexes, [Fe(L(n))(acac)(C(2)H(5)OH)] incorporating coordinated ethanol from the reaction solvent were accessed from the reaction of [Fe(acac)(3)] with [ONO] donor dibasic tridentate unsymmetrical Schiff base ligands derived from condensation of 2-hydroxy-1-napthaldehyde with 2-aminophenol (H(2)L(1)) or 2-aminobenzoic acid (H(2)L(2)). The thermal study (TGA-DTA) provided evidence for weakly bound ethanol which is readily substituted by neutral N-donor molecule imidazole, benzimidazole or pyridine to produce an array of newer complexes, [Fe(L(n))(acac)X] (n=1, 2; X=Im, Bim, Py). The compounds were characterized by elemental analyses, FT-IR, UV-vis, solution electrical conductivity, FAB mass, (1)H and (13)C NMR spectroscopy. Room temperature magnetic susceptibility measurements (μ(eff)～5.8 B.M.) are consistent with spin-free octahedral iron(III) complexes. Cyclic voltammetry of ethanol complexes revealed a quasi-reversible one electron redox response (ΔE(p)>100 mV) for the Fe(III)/Fe(II) couple. Low half wave redox potential (E(1/2)) values suggested easy redox susceptibility. The ground state geometries of the ethanol and imidazole complexes have been ascertained to be distorted octahedral by density functional theory using DMol3 program at BLYP/DNP level.     

Structural analysis of biologically active peptides and recombinant proteins and their modified counterparts by mass spectrometry

The structural characterization of the Escherichia coli-expressed human interferon alpha-2b (rh-IFN alpha-2b) was carried out by employing the fast atom bombardment (FAB) and plasma desorption (PD) mapping methods. The mass spectral data of the rh-IFN alpha-2b and the trypsin-generated peptide mixture allowed rapid and facile confirmation of the cDNA-derived sequence and determination of the existing disulfide pattern in the protein molecule. The same PD/FAB mapping approach was successfully employed in the structural determination of the iodination reaction product of rh-IFN alpha-2b and the potent vasoconstrictor peptide endothelin.     

The effect of viscosity on the waves due to a moving oscillatory surface pressure

Summary It is known that when a pressure distributionp ₀(x) exp (i t moves with a constant velocityu ₀ parallel to on the surface (y=0) of an inviscid liquid, (i) the steady-state wave amplitude does not die out at large distances from the source for (u ₀/g1/4, and(ii) the solution becomes singular for u ₀/g=1/4.Here is shown that the consideration of even a slight viscosity of the liquid leads to a wave decay with distance for all (u ₀/g), the solution remaining non-singular for (u ₀/g=1/4. Furthermore, for a finitely distributedp ₀(x), the wave amplitude is always non-zero, unlike in the case of an inviscid fluid, where no energy is transmitted for an infinite number of values of a parameter involving ,u ₀ and the length of the pressure strip.

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Zusammenfassung Es ist bekannt, daß, wenn eine periodische Druckverteilungp ₀(x) exp (i t) sich mit einer konstanten Geschwindigkeitu ₀ inx-Richtung über die Oberflächey=0 einer reibungslosen Flüssigkeit bewegt, daß dann (1) die Amplitude des stationären Zustandes in großen Entfernungen von der Quelle nicht abklingt für (u ₀/g)1/4, und (2) die Lösung wird singulär für (u ₀/g)=1/4.Hier wird gezeigt, daß die Berücksichtigung einer geringen Zähigkeit in der Flüssigkeit zu einem Abklingen der Wellen führt für alle (u ₀/g)=1/4. Zudem ist für eine Verteilung vonp ₀(x) über eine endliche Länge die Wellenamplitude immer endlich, im Gegensatz zum reibungsfreien Fall, in dem keine Energie übertragen wird für gewisse Werte eines Parameters, der ,u ₀ und die Länge des Druckstreifens enthält.

     

Synthesis and fluorescent properties of a new class of heterocycles of isoindole fused imidazoles with phenolic subunits

A new class of heterocycles of isoindole fused imidazoles with phenolic subunits has been readily synthesized by a two-step one-pot reaction. In aprotic solvent they show high fluorescent properties (Phi(F) up to 0.93), but in protic polar solvent fluorescent intensity decreases. They show green fluorescence in weak acidic medium such as acetic acid but lack emission in basic medium. The compounds can also stain human squamous epithelium cells.     

Boat-like Au nanoparticles embedded mesoporous γ-Al2O3 films: an efficient SERS substrate

A new water-dispersible nanostructure based on magnetite (Fe₃O₄) and usnic acid (UA) was prepared in a well-shaped spherical form by a precipitation method. Nanoparticles were well individualized and homogeneous in size. The presence of Fe₃O₄@UA was confirmed by transmission electron microscopy, Fourier transform-infrared spectroscopy, and X-ray diffraction. The UA was entrapped in the magnetic nanoparticles during preparation and the amount of entrapped UA was estimated by thermogravimetric analysis. Fabricated nanostructures were tested on planktonic cells growth (minimal inhibitory concentration assay) and biofilm development on Gram-positive Staphylococcus aureus (S. aureus), Enterococcus faecalis (E. faecalis) and Gram-negative Escherichia coli (E. coli), Pseudomonas aeruginosa (P. aeruginosa) reference strains. Concerning the influence of Fe₃O₄@UA on the planktonic bacterial cells, the functionalized magnetic nanoparticles exhibited a significantly improved antimicrobial activity against E. faecalis and E. coli, as compared with the Fe₃O₄ control. The UA incorporated into the magnetic nanoparticles exhibited a very significant inhibitory effect on the biofilm formed by the S. aureus and E. faecalis, on a wide range of concentrations, while in case of the Gram-negative microbial strains, the UA-loaded nanoparticles inhibited the E. coli biofilm development, only at high concentrations, while for P. aeruginosa biofilms, no inhibitory effect was observed. The obtained results demonstrate that the new water-dispersible Fe₃O₄@UA nanosystem, combining the advantages of the intrinsic antimicrobial features of the UA with the higher surface to volume ratio provided by the magnetic nanocarrier dispersible in water, exhibits efficient antimicrobial activity against planktonic and adherent cells, especially on Gram-positive strains.     

Modulated photophysics of 3-pyrazolyl-2-pyrazoline derivative entrapped in micellar assembly

The photophysical behavior of 3-pyrazolyl-2-pyrazoline derivative (PZ), a newly synthesized biologically active compound has been studied in micellar solutions of anionic sodium dodecyl sulfate (SDS), cationic cetyl trimethylammonium bromide (CTAB) and nonionic p- tert-octylphenoxy polyoxyethanol (Triton X-100, TX-100) micelle using steady state and time-resolved fluorescence spectroscopy technique. Influence of the micelles on the photophysics of PZ has also been investigated using different approaches. The location of the fluorophore PZ in the micelle has been identified by cetyl pyridinium chloride (CpCl) induced fluorescence quenching and micropolarity surrounding that fluorophore in micellar solution. The effect of urea on the steady state fluorescence and relaxation dynamics of the micelle bound probe has also been observed. The results have been interpreted in terms of the model that urea displaces water molecules from the micellar interface and the consequent destabilization leads to the expulsion of the probe molecules from the interfacial region. An attempt has been made to determine probe sensing microviscosities for these micellar microenvironments in the light of average reorientation times of the probe PZ.     

Posaconazole (Noxafil, SCH 56592), a new azole antifungal drug, was a discovery based on the isolation and mass spectral characterization of a circulating metabolite of an earlier lead (SCH 51048)

Posaconazole (SCH 56592) is a novel triazole antifungal drug that is marketed in Europe and the United States under the trade name 'Noxafil' for prophylaxis against invasive fungal infections. SCH 56592 was discovered as a possible active metabolite of SCH 51048, an earlier lead. Initial studies have shown that serum concentrations determined by a microbiological assay were higher than those determined by HPLC from animals dosed with SCH 51048. Subsequently, several animals species were dosed with (3)H-SCH 51048 and the serum was analyzed for total radioactivity, SCH 51048 concentration and antifungal activity. The antifungal activity was higher than that expected based on SCH 51048 serum concentrations, confirming the presence of active metabolite(s). Metabolite profiling of serum samples at selected time intervals pinpointed the peak that was suspected to be the active metabolite. Consequently, (3)H-SCH 51048 was administered to a large group of mice, the serum was harvested and the metabolite was isolated by extraction and semipreparative HPLC. LC-MS/MS analysis suggested that the active metabolite is a secondary alcohol with the hydroxyl group in the aliphatic side chain of SCH 51048. All corresponding monohydroxylated diastereomeric mixtures were synthesized and characterized. The HPLC retention time and LC-MS/MS spectra of the diastereomeric secondary alcohols of SCH 51048 were similar to those of the isolated active metabolite. Finally, all corresponding individual monohydroxylated diasteriomers were synthesized and evaluated for in vitro and in vivo antifungal potencies, as well as pharmacokinetics. SCH 56592 emerged as the candidate with the best overall profile.