Rods, tori, and honeycombs: the directed self-assembly of microtissues with prescribed microscale geometries.

It is thought that, due to energy and surface area:volume minimization, the spheroid is the terminal structure of cellular self-assembly. We investigated whether self-assembly could be directed to generate complex-shaped structures. Using micromolded, nonadhesive agarose hydrogels seeded with rat hepatoma (H35s), human fibroblasts (NHFs), or their mix (1:1), we show that cells can self-assemble rods, tori, and honeycombs.

Dynamics of the self-assembly of complex cellular aggregates on micromolded nonadhesive hydrogels.

The process by which cells self-assemble to form three-dimensional (3D) structures is central to morphogenesis and development of living tissues and hence is of growing interest to the field of tissue engineering. Using rapid prototyping technology we made micromolded nonadhesive hydrogels to study the dynamics of self-assembly in a low-shear environment with simple spherical geometries as well as more complex geometries such as a toroid. Aggregate size, shape, and composition were easily controlled; aggregates were easily retrieved; and the dynamics of the assembly process were readily observed by time-lapse microscopy.

Studies on the effect of Amadoriase from Aspergillus fumigatus on peptide and protein glycation in vitro.

Amadoriase I is a fructosyl amine oxidase from Aspergillus fumigatus that catalyzes the oxidation of Amadori products (APs) producing glucosone, H2O2, and the corresponding free amine. All the enzymes of this family discovered so far only deglycate small molecular weight products and are inactive toward large molecular weight substrates, such as glycated BSA or ribonuclease A. Therefore, they cannot be used to reverse protein glycation occurring in diabetes or in foods.

HIV-1 Nef impairs the dynamic of DC/NK crosstalk: different outcome of CD56(dim) and CD56(bright) NK cell subsets.

Dendritic cells (DCs) and natural killer (NK) cells are essential components of the innate immunity and play a critical role in the first phase of host defense against infection. Interactions between DCs and NK cells have been demonstrated in a variety of settings, with evidence emerging of complex bidirectional crosstalk between the two cell types. The accessory HIV-1 Nef protein is a crucial determinant for viral replication and pathogenesis.

An easy-to-run method for routine analysis of eumelanin and pheomelanin in pigmented tissues.

A procedure for analysis of melanin-pigmented tissues based on alkaline hydrogen peroxide degradation coupled with high-performance liquid chromatography (HPLC) ultraviolet determination of pyrrole-2,3,5-tricarboxylic acid (PTCA) for eumelanin and 6-(2-amino-2-carboxyethyl)-2-carboxy-4-hydroxybenzothiazole (BTCA) and 1,3-thiazole-2,4,5-tricarboxylic acid for pheomelanin was recently developed. Despite advantages related to the degradation conditions and sample handling, a decrease of the reproducibility and resolution was observed after several chromatographic runs. We report herein an improved chromatographic methodology for simultaneous determination of PTCA and BTCA as representative markers of eumelanin and pheomelanin, respectively, based on the use of an octadecylsilane column with polar end-capping with 1% formic acid (pH 2.

Liquid chromatography/electrospray ionization tandem mass spectrometry profiling of compounds from the infusion of Byrsonima fagifolia Niedenzu.

A rapid analytical approach suitable to achieve a comprehensive characterization of the compounds present in the infusion prepared from the leaves of Byrsonima fagifolia Niedenzu (Malpighiaceae), a Brazilian plant used as an infusion to treat gastric disorders, was developed. The method was based on high-performance liquid chromatography coupled to electrospray negative ionisation multistage ion trap mass spectrometry (HPLC/ESI-ITMSn). The main ions in the ESI-ITMS spectra were attributed to a quinic acid core containing from one to five galloyl units.

The first 5,6-dihydroxyindole tetramer by oxidation of 5,5',6,6'-tetrahydroxy- 2,4'-biindolyl and an unexpected issue of positional reactivity en route to eumelanin-related polymers.

[structure: see text]. The first tetramer of the eumelanin precursor 5,6-dihydroxyindole has been obtained, as the acetyl derivative, by peroxidase/H2O2-induced oxidative coupling of 5,5',6,6'-tetrahydroxy-2,4'-biindolyl (2) in the presence of Zn2+ ions. The tetramer, 5,5',5'',5''',6,6',6'',6'''-octaacetoxy-2,4':2',3'':2'',4'''-tetraindolyl (acetylated 7), incorporates an unprecedented 2,3'-biindolyl substructure suggestive of a different positional reactivity of the 5,6-dihydroxyindole system when framed into a dimeric scaffold.

Chemical, pulse radiolysis and density functional studies of a new, labile 5,6-indolequinone and its semiquinone.

The chemical and spectroscopic characterization of 5,6-indolequinones and their semiquinones, key transient intermediates in the oxidative conversion of 5,6-dihydroxyindoles to eumelanin biopolymers, is a most challenging task. In the present paper, we report the characterization of a novel, relatively long-lived 5,6-indolequinone along with its semiquinone using an integrated chemical, pulse radiolytic, and computational approach. The quinone was obtained by oxidation of 5,6-dihydroxy-3-iodoindole (1a) with o-chloranil in cold ethyl acetate or aqueous buffer: it displayed electronic absorption bands around 400 and 600 nm, was reduced to 1a with Na2S2O4, and reacted with o-phenylenediamine to give small amounts of 3-iodo-1H-pyrrolo[2,3-b]phenazine (2).

The chemical basis of the antinitrosating action of polyphenolic cancer chemopreventive agents.

A regular intake of polyphenolic agents widely found in fruits and vegetables is believed to decrease the incidence of certain forms of cancer, due in part to their ability to act as antinitrosating agents capable of lowering the impact of toxic nitrosation processes and carcinogenic nitrosamine formation within the acidic environment of the stomach. As a result, the study of the interactions between reactive nitrogen species and phenolic antioxidants has emerged as an area of great promise for delineating innovative strategies in cancer chemoprevention. The burst of interest in (poly)phenolic cancer chemopreventive agents of dietary origin is exemplified by the exponential growth of scientific literature on green tea catechins, as well as on hydroxycinnamates, hydroxytyrosol, flavonoids and other phenolic compounds of the Mediterranean diet, currently regarded as a cultural model for dietary improvement.

Short-lived quinonoid species from 5,6-dihydroxyindole dimers en route to eumelanin polymers: integrated chemical, pulse radiolytic, and quantum mechanical investigation.

The transient species formed by oxidation of three dimers of 5,6-dihydroxyindole (1), a major building block of the natural biopolymer eumelanin, have been investigated. Pulse radiolytic oxidation of 5,5',6,6'-tetrahydroxy-2,4'-biindolyl (3) and 5,5',6,6'-tetrahydroxy-2,7'-biindolyl (4) led to semiquinones absorbing around 450 nm, which decayed with second-order kinetics (2k=2.8x10(9) and 1.

Treatment of cereal products with a tailored preparation of trichoderma enzymes increases the amount of soluble dietary fiber.

Nutritionists recommend increasing the intake of soluble dietary fiber (SDF), which is very low in most cereal-based products. Conversion of insoluble DF (IDF) into SDF can be achieved by chemical treatments, but this affects the sensorial properties of the products. In this study, the possibility of getting a substantial increase of SDF from cereal products using a tailored preparation of Trichoderma enzymes is reported.

Dopaquinone redox exchange with dihydroxyindole and dihydroxyindole carboxylic acid.

A pulse radiolytic investigation has been conducted to establish whether a redox reaction takes place between dopaquinone and 5,6-dihydroxyindole (DHI) and its 2-carboxylic acid (DHICA) and to measure the rate constants of the interactions. To obviate possible confounding reactions, such as nucleophilic addition, the method employed to generate dopaquinone used the dibromide radical anion acting on dopa to form the semiquinone which rapidly disproportionates to dopaquinone. In the presence of DHI the corresponding indole-5,6-quinone (and/or tautomers) was also formed directly but, by judicious selection of suitable relative concentrations of initial reactants, we were able to detect the formation of additional indolequinone from the redox exchange reaction of DHI with dopaquinone which exhibited a linear dependency on the concentration of DHI.

Practical one-pot conversion of 17beta-estradiol to 10beta-hydroxy- (p-quinol) and 10beta-chloro-17beta-hydroxyestra-1,4-dien-3-one.

An efficient one-pot procedure for the preparation of 10beta,17beta-dihydroxyestra-1,4-dien-3-one (p-quinol, 1, 75%) is reported, involving oxidation of 17beta-estradiol with potassium permanganate. Similar treatment of 17beta-estradiol with sodium chlorite led to 10beta-chloro-17beta-hydroxyestra-1,4-dien-3-one (2) in 44% yield along with smaller amounts 4-chloro-10beta,17beta-dihydroxyestra-1,4-dien-3-one (3), 2,10beta-dichloro-17beta-hydroxyestra-1,4-dien-3-one (4), and 4,10beta-dichloro-17beta-hydroxyestra-1,4-dien-3-one (5).

Acid-promoted reaction of the stilbene antioxidant resveratrol with nitrite ions: mild phenolic oxidation at the 4'-hydroxystiryl sector triggering nitration, dimerization, and aldehyde-forming routes.

In 0.1 M phosphate buffer, pH 3.0, and at 37 degrees C, resveratrol ((E)-3,4',5-trihydroxystilbene, 1a), an antioxidant and cancer chemopreventive phytoalexin, reacted smoothly at 25 microM or 1 mM concentration with excess nitrite ions (NO2(-)) to give a complex pattern of products, including two novel regioisomeric alpha-nitro (3a) and 3'-nitro (4) derivatives along with some (E)-3,4',5-trihydroxy-2,3'-dinitrostilbene (5), four oxidative breakdown products, 4-hydroxybenzaldehyde, 4-hydroxy-3-nitrobenzaldehyde, 3,5-dihydroxyphenylnitromethane, and 3,5-dihydroxybenzaldehyde, two dimers, the resveratrol (E)-dehydrodimer 6 and restrytisol B (7), and the partially cleaved dimer 2.

Glyoxal formation by Fenton-induced degradation of carbohydrates and related compounds.

In this paper, we provide a systematic analysis of glyoxal (1) formation from a range of monosaccharides and related compounds, to determine their potential role as sources of this alpha-oxoaldehyde in vivo. Substrates were reacted with the Fenton reagent (Fe(2+)/EDTA/H(2)O(2)) and the mixtures were analyzed by HPLC using the 6-hydroxy-2,4,5-triaminopyrimidine fluorimetric assay. The rank order of hexoses and their derivatives as glyoxal sources was found to be fructose > glucose = mannose = galactose > glucose-6-phosphate > mannitol.

Free radical oxidation of 15-(S)-hydroxyeicosatetraenoic acid with the Fenton reagent: characterization of an epoxy-alcohol and cytotoxic 4-hydroxy-2E-nonenal from the heptatrienyl radical pathway.

The oxidation of (5Z,8Z,11Z,13E,15S)-15-hydroxy-5,8,11,13-eicosatetraenoic acid (15-(S)-HETE, 1a) with the Fenton reagent (Fe2+/EDTA/H2O2) was investigated. In phosphate buffer, pH 7.4, the reaction proceeded with 75% substrate consumption after 1 h to give a mixture of products, one of which was identified as (2E,4S)-4-hydroxy-2-nonenal (3a, 18% yield).

The catecholic antioxidant piceatannol is an effective nitrosation inhibitor via an unusual double bond nitration.

Piceatannol (1) was found to be more effective than caffeic acid, an established antinitrosating agent, in inhibiting N-nitrosation of 2,3-diaminonaphthalene. Product analysis of the reaction mixture of 1 (20 microM) with nitrite ions (80 microM) at pH 3.0 and at 37 degrees C showed conversion to a single major nitration product, (E)-3,3',4,5'-tetrahydroxy-beta-nitrostilbene (2) (68% yield).

Vacuum-assisted core biopsy (Mammotome) for the diagnosis of non-palpable breast lesions: four-year experience in an Italian center.

Aims And Background: Microinvasive biopsy techniques are increasingly employed in the diagnostic assessment of non-palpable breast lesions (NPBLs). This study reports the four-year experience of an Italian surgical center in the evaluation of the diagnostic effectiveness versus adverse effects of vacuum-assisted core biopsy (Mammotome).

Methods: 314 NPBLs with a dubious ultrasound and/or radiographic appearance were subjected to mammography-guided (86) or ultrasound-guided (228) Mammotome biopsy.

Reactions of D-glucose with phenolic amino acids: further insights into the competition between Maillard and Pictet-Spengler condensation pathways.

The reactions of 5-S-cysteinyldopa, L-alpha-methyldopa and DL-m-tyrosine with D-glucose were investigated at 90 degrees C in phosphate buffer at pH ranging from 5.0 to 9.0.

Glycation of lysine-containing dipeptides.

Protein glycation through Maillard reaction (MR) is a fundamental reaction both in foods and in the human body. The first step of the reaction is the formation of Amadori product (AP) that is converted into intermediate and advanced MR products during reaction development. Although the MR is not an enzymatic reaction, a certain degree of specificity in the glycation site has been observed.

Tyrosinase-catalyzed oxidation of 17beta-estradiol: structure elucidation of the products formed beyond catechol estrogen quinones.

This paper reports a systematic characterization of the products formed by oxidation of 17beta-estradiol (1) with tyrosinase/O2 at low concentrations of physiological relevance. With the substrate at 1-10 nM concentration, the main reaction products included, beside the catechol estrogens 2-hydroxyestradiol (2) and 4-hydroxyestradiol (3), 6-oxo-2-hydroxyestradiol (4), 9,11-dehydro-2-hydroxyestradiol (6), 6,7-dehydro-2-hydroxyestradiol (7), and 9,11-dehydro-4-hydroxyestradiol (10). At higher estradiol concentrations, e.

Apples increase nitric oxide production by human saliva at the acidic pH of the stomach: a new biological function for polyphenols with a catechol group?

Dietary inorganic nitrate is secreted in saliva and reduced to nitrite by bacterial flora. At the acidic pH of the stomach nitrite is present as nitrous acid in equilibrium with nitric oxide (*NO), and other nitrogen oxides with nitrating and nitrosating activity. *NO in the stomach exerts several beneficial effects, but nitrosating/nitrating species have been implicated as a possible cause of epithelial neoplasia at the gastroesophageal junction.

Oxidative chemistry of 2-nitro and 4-nitroestradiol: Dichotomous behavior of radical intermediates and novel potential routes for oxyfunctionalization and B-ring fission of steroidal scaffolds.

Oxidation of 4-nitro-17beta-estradiol (1) with the peroxidase/H(2)O(2) system gave the symmetric C(2)-linked dimer (3) through phenoxy radical coupling. Similar oxidation of 2-nitro-17beta-estradiol (2), in which the nitro group is coplanar with the aromatic ring, yielded 9alpha- and 9beta-hydroxy-2-nitro-17beta-estradiol (4a,b), (17beta)-2-nitroestra-1(10),2,4,9(11)-tetraene-3,17-diol (5), and (12alpha,17beta)-2-nitroestra-1(10),2,4,9(11)-tetraene-3,12,17-triol (6). With higher concentrations of H(2)O(2), the novel secoestra-1(10),2,4-trien-9-one derivative 7 was obtained from 2.

Autosomal dominant hereditary spastic paraplegia: DHPLC-based mutation analysis of SPG4 reveals eleven novel mutations.

We set up a new denaturing high-performance liquid chromatography (DHPLC)-based protocol to screen patients with autosomal dominant hereditary spastic paraplegia (AD-HSP) for mutations in SPG4. Six patients had a complicated form and 49 a pure HSP phenotype. We also analyzed 19 unrelated patients presenting with an HSP phenotype (pure in 17 and complicated in two subjects) but no clear family history, as such patients may be cases of dominant inheritance with low penetrance.

The acid-promoted reaction of the green tea polyphenol epigallocatechin gallate with nitrite ions.

Exposure of 400 microM (-)-epigallocatechin-3-O-gallate (EGCG), the main polyphenolic constituent of green tea, to equimolar concentrations of nitrite ions in 0.5 M HCl at 37 degrees C resulted in the formation of a distinct pattern of products that were trapped as phenazine derivatives by treatment with o-phenylenediamine. Repeated chromatographic fractionation eventually allowed isolation of four main species, which were identified by 2D NMR and MS analysis as 1b, derived from EGCG quinone 1a, the isomeric oximes 2b,c, arising from nitrosation of EGCG on the pyrogallol B-ring, and the dioxime 4b in which the A-ring was doubly substituted.