Sub-tropical urban environment affecting content and composition of non-structural carbohydrates of Lolium multiflorum ssp italicum cv. Lema

This study analyzed the relationship between environmental factors, especially air pollution and climatic conditions, and non-structural carbohydrates (NSC) in plants of Lolium multiflorum exposed during 10 consecutive periods of 28 days at a polluted site (Congonhas) and at a reference site in Sao Paulo city (Brazil). After exposure, NSC composition and leaf concentrations of Al, Fe. Cu, Zn, Pb and Cd were measured. The seasonal pattern of NSC accumulation was quite similar in both sites, but plants at Congonhas showed higher concentrations of these compounds, especially fructans of low and medium degree of polymerization. Regression analysis showed that NSC in plants growing at the polluted site were explained by variations on temperature and leaf concentration of Fe (positive effect), as well as relative humidity and particulate material (negative effect). NSC in the standardized grass culture, in addition to heavy metal accumulation, may indicate stressing conditions in a sub-tropical polluted environment. (C) 2008 Elsevier Ltd. All rights reserved.

Identification of the nuclear factor kappa-beta (NF-kB) in cortical of mice Wistar using Technovit 7200 VCR (R)

Objective: this study aimed to develop a nondecalcified bone sample processing technique enabling immunohistochemical labeling of proteins by kappa-beta nuclear factor (NF-kB) utilizing the Technovit 7200 VCR (R) in adult male Wistar rats. Study Method: A 1.8 mm diameter defect was performed 0.5mm from the femur proximal joint by means of a round bur. Experimental groups were divided according to fixing solution prior to histologic processing: Group 1- ethanol 70%; Group 2-10% buffered formalin; and Group 3- Glycerol diluted in 70% ethanol at a 70/30 ratio + 10% buffered formalin. The post-surgical periods ranged from 01 to 24 hours. Control groups included a nonsurgical procedure group (NSPG) and surgical procedures where bone exposure was performed (SPBE) without drilling. Prostate carcinoma was the positive control (PC) and samples subjected to incomplete immunohistochemistry protocol were the negative control (NC). Following euthanization, all samples were kept at 4 degrees C for 7 days, and were dehydrated in a series of alcohols at -20 degrees C. The polymer embedding procedure was performed at ethanol/polymer ratios of 70%-30%, 50%-50%, 30%-70%, 100%, and 100% for 72 hours at -20 degrees C. Polymerization followed the manufacturer`s recommendation. The samples were grounded and polished to 10-15 mu m thickness, and were deacrylated. The sections were rehydrated and were submitted to the primary polyclonal antibody anti-NF-kB on a 1:75 dilution for 12 hours at room temperature. Results: Microscopy showed that the Group 2 presented positive reaction to NF-kB, diffuse reactions for NSPG and SPBE, and no reaction for the NC group. Conclusion: The results obtained support the feasibility of the developed immunohistochemistry technique.

T3 acutely increases GH mRNA translation rate and GH secretion in hypothyroid rats

Cytoskeleton controls the stability of transcripts, by mechanisms that involve mRNAs and eEF1A attachment to it. Besides, it plays a key role in protein synthesis and secretion, which seems to be impaired in somatotrophs of hypothyroid rats, whose cytoskeleton is disarranged. This study investigated the: eEF1A and GH mRNA binding to cytoskeleton plus GH mRNA translation rate and GH secretion, in sham-operated and thyroidectomized rats treated with T3 or saline, and killed 30 min thereafter. Thyroidectomy reduced: (a) pituitary F-actin content, and eEF1A plus GH mRNA binding to it; (b) GH mRNA recruitment to polysome; and (c) liver IGF-1 mRNA expression, indicating that GH mRNA stability and translation rate, as well as GH secretion were impaired. T3 acutely reversed all these changes, which points toward a nongenomic action of T3 on cytoskeleton rearrangement, which might contribute to the increase on GH mRNA translation rate and GH secretion. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Rho Signaling Pathway and Apical Constriction in the Early Lens Placode

Epithelial invagination in many model systems is driven by apical cell constriction, mediated by actin and myosin II contraction regulated by GTPase activity. Here we investigate apical constriction during chick lens placode invagination. Inhibition of actin polymerization and myosin II activity by cytochalasin D or blebbistatin prevents lens invagination. To further verify if lens placode invaginate through apical constriction, we analyzed the role of Rho-ROCK pathway. Rho GTPases expression at the apical portion of the lens placode occurs with the same dynamics as that of the cytoskeleton. Overexpression of the pan-Rho inhibitor C3 exotoxin abolished invagination and had a strong effect on apical myosin II enrichment and a mild effect on apical actin localization. In contrast, pharmacological inhibition of ROCK activity interfered significantly with apical enrichment of both actin and myosin. These results suggest that apical constriction in lens invagination involves ROCK but apical concentration of actin and myosin are regulated through different pathways upstream of ROCK. genesis 49: 368-379, 2011. (C) 2011 Wiley-Liss, Inc.

The geodiamolide H, derived from Brazilian sponge Geodia corticostylifera, regulates actin cytoskeleton, migration and invasion of breast cancer cells cultured in three-dimensional environment

We are investigating effects of the depsipeptide geodiamolide H, isolated from the Brazilian sponge Geodia corticostylifera, on cancer cell lines grown in 3D environment. As shown previously geodiamolide H disrupts actin cytoskeleton in both sea urchin eggs and breast cancer cell monolayers. We used a normal mammary epithelial cell line MCF 10A that in 3D assay results formation of polarized spheroids. We also used cell lines derived from breast tumors with different degrees of differentiation: MCF7 positive for estrogen receptor and the Hs578T, negative for hormone receptors. Cells were placed on top of Matrigel. Spheroids obtained from these cultures were treated with geodiamolide H. Control and treated samples were analyzed by light and confocal microscopy. Geodiamolide H dramatically affected the poorly differentiated and aggressive Hs578T cell line. The peptide reverted HsS78T malignant phenotype to polarized spheroid-like structures. MCF7 cells treated by geodiamolide H exhibited polarization compared to controls. Geodiamolide H induced striking phenotypic modifications in Hs578T cell line and disruption of actin cytoskeleton. We investigated effects of geodiamolide H on migration and invasion of Hs578T cells. Time-lapse microscopy showed that the peptide inhibited migration of these cells in a dose-dependent manner. Furthermore invasion assays revealed that geodiamolide H induced a 30% decrease on invasive behavior of Hs578T cells. Our results suggest that geodiamolide H inhibits migration and invasion of Hs578T cells probably through modifications in actin cytoskeleton. The fact that normal cell lines were not affected by treatment with geodiamolide H stimulates new studies towards therapeutic use for this peptide.

T3 rapidly modulates TSH beta mRNA stability and translational rate in the pituitary of hypothyroid rats

Whereas it is well known that T3 inhibits TSH beta gene transcription, its effects on TSH beta mRNA stability and translation have been poorly investigated. This study examined these possibilities, by evaluating the TSH beta transcripts poly(A) tail length, translational rate and binding to cytoskeleton, in pituitaries of thyroidectomized and sham-operated rats treated with T3 or saline, and killed 30 min thereafter. The hypothyroidism induced an increase of TSH beta transcript poly(A) tail, as well as of its content in ribosomes and attachment to cytoskeleton. The hypothyroid rats acutely treated with T3 exhibited a reduction of TSH beta mRNA poly(A) tail length and recruitment to ribosomes, indicating that this treatment decreased the stability and translation rate of TSH beta mRNA. Nevertheless, acute T3 administration to sham-operated rats provoked an increase of TSH beta transcripts binding to ribosomes. These data add new insight to an important role of T3 in rapidly regulating TSH gene expression at posttranscriptional level. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

Effects of Synthesis Conditions on the Nanostructure of Hybrid Sols Produced by the Hydrolytic Condensation of (3-Methacryloxypropyl)trimethoxysilane

Polysilsesquioxanes containing methacrylate pendant groups were prepared by the sol-gel process through hydrolysis and condensation of (3-methacryloxypropyl)trimethoxysilane (MPTS) dissolved in a methanol/methyl methacrylate (MMA) mixture. The effects of different water, MMA, and methanol contents, as well as of pH, on the nanoscopic and local structures of the system, at advanced stages of the condensation reaction, were studied by small-angle X-ray scattering (SAXS) and (29)Si nuclear magnetic resonance (NMR) spectroscopy, respectively. SAXS results indicate that the nanoscopic features of the hybrid sol could be described by a hierarchical model composed of two levels, namely (i) silsesquioxane (SSQO) nanoparticles Surrounded by the methacrylate pendant groups and the methanol/MMA mixture. and (ii) aggregation zones or islands containing correlated SSQO nanoparticles, embedded in the liquid medium. The (29)Si NMR results Show that the inner Structures of SSQO nanoparticles produced at pH 1 and 3 were built Up of polyhedral structures. mainly cagelike octamers and small linear oligomers, respectively. Irrespective of MMA and methanol contents, for a [H(2)O]/[MPTS] ratio higher than or equal to 1, the SSQO nailoparticles produced at pH I exhibit an average condensation degree (CD approximate to 69-87%) and average radius of gyration (R(g) approximate to 2.5 angstrom) larger than those produced at pH 3 (CD approximate to 48-67% and R(g) approximate to 1.5 angstrom). Methanol appears to act as a redispersion agent, by decreasing the number of particles inside the aggregation zones, while the addition of MMA induces a swelling of the aggregation zones.

Structural Characterization of Photopolymerizable Binary Liposomes Containing Diacetylenic and Saturated Phospholipids

The use of liposomes to encapsulate materials has received widespread attention for drug delivery, transfection, diagnostic reagent, and as immunoadjuvants. Phospholipid polymers form a new class of biomaterials with many potential applications in medicine and research. Of interest are polymeric phospholipids containing a diacetylene moiety along their acyl chain since these kinds of lipids can be polymerized by Ultra-Violet (UV) irradiation to form chains of covalently linked lipids in the bilayer. In particular the diacetylenic phosphatidylcholine 1,2-bis(10,12-tricosadiynoyl)- sn-glycero-3-phosphocholine (DC8,9PC) can form intermolecular cross-linking through the diacetylenic group to produce a conjugated polymer within the hydrocarbon region of the bilayer. As knowledge of liposome structures is certainly fundamental for system design improvement for new and better applications, this work focuses on the structural properties of polymerized DC8,9PC:1,2-dimyristoyl-sn-glycero-3-phusphocholine (DMPC) liposomes. Liposomes containing mixtures of DC8,9PC and DMPC, at different molar ratios, and exposed to different polymerization cycles, were studied through the analysis of the electron spin resonance (ESR) spectra of a spin label incorporated into the bilayer, and the calorimetric data obtained from differential scanning calorimetry (DSC) studies. Upon irradiation, if all lipids had been polymerized, no gel-fluid transition would be expected. However, even samples that went through 20 cycles of UV irradiation presented a DSC band, showing that around 80% of the DC8,9PC molecules were not polymerized. Both DSC and ESR indicated that the two different lipids scarcely mix at low temperatures, however few molecules of DMPC are present in DC8,9PC rich domains and vice versa. UV irradiation was found to affect the gel fluid transition of both DMPC and DC8,9PC rich regions, indicating the presence of polymeric units of DC8,9PC in both areas, A model explaining lipids rearrangement is proposed for this partially polymerized system.

Thermal and structural properties of commercial dental resins light-cured with blue emitting diodes (LEDs)

We have investigated the thermal and structural properties of different commercial dental resins: Filtek(TM) Z-350, Grandio(A (R)), Tetric Ceram(A (R)), and TPH Spectrum(A (R)). The purpose of the present study was to evaluate quantitatively the photo-polymerization behavior and the effect of filler contents on the kinetic cures of the dental resins by using Differential Scanning Calorimetry (DSC) and Fourier Transform Infrared Spectroscopy (FT-IR) techniques. We have successfully obtained the low and high glass transition T (g) values of the dental composite resins from DSC curves. It was also observed a good agreement between the both T (g) values, activation energies from thermal degradation, and the degree of conversion obtained for all samples. The results have shown that Tetric Ceram(A (R)) dental resin presented the higher T (g) values, activation energy of 215 +/- A 6 KJ mol(-1), and the higher degree of conversion (63%) when compared to the other resins studied herein.

Measurement of shrinkage of composite resin by laser speckle contrast analysis

The aim of this study was to evaluate the shrinkage of a microhybrid dental composite resin photo-activated by one LED with different power densities by means of speckle technique. The dental composite resin Filtek (TM) Z-250 (3M/ESPE) at color A(2) was used for the samples preparation. Uncured composite was packed in a metallic mold and irradiated during 20 s from 100 to 1000 mW cm(-2). For the photo-activation of the samples, it was used a LED prototype (Light Emission Diode) with wavelength centered at 470 nm and adjustable power density until 1 W cm(-2). The speckle patterns obtained from the bottom composite surfaces were monitored using a CCD camera without lens. The speckle field is recorded in a digital picture and stored by CCD camera as the carrier of information on the displacement of the tested surface. The calculated values were obtained for each pair of adjacent patterns and the changes in speckle contrast as a function of time were obtained from six repeated measurements. The speckle contrasts obtained from the bottom surface with 100 mW cm(-1) were smaller than those than the other power densities. The higher power densities provided the higher shrinkage.

Enzyme activity of horseradish peroxidase immobilized in chitosan matrices in alternated layers

The successful immobilization of enzymes such as horseradish peroxidase (HRP) in solid films is essential for applications in sensors and for fundamental studies aimed at identifying possible biotechnological devices. In this study we show that HRP can be immobilized in alternated layers with chitosan as the template material. The activity of HRP in HRP/chitosan films was preserved for several weeks, and could be detected optically upon monitoring the reaction with pyrogallol. The morphology of the film displayed stripes that disappeared after reaction with pyrogallol. Though the activity in the HRP/chitosan film was lower than in a homogeneous solution or in an LB film investigated earlier, the response was linear for a considerable period of time, which may be advantageous for sensing hydrogen peroxide. (C) 2009 Elsevier B.V. All rights reserved.

Influence of pre-heat treatment and different light-curing units on Vickers hardness of a microhybrid composite resin

The aim of this study was to evaluate the hardness of a dental composite resin submitted to temperature changes before photo-activation with two light-curing unite (LCUs). Five samples (4 mm in diameter and 2 mm in thickness) for each group were made with pre-cure temperatures of 37, 54, and 60A degrees C. The samples were photo-activated with a conventional quartz-tungsten-halogen (QTH) and blue LED LCUs during 40 s. The hardness Vickers test (VHN) was performed on the top and bottom surfaces of the samples. According to the interaction between light-curing unit and different pre-heating temperatures of composite resin, only the light-curing unit provided influences on the mean values of initial Vickers hardness. The light-curing unit based on blue LED showed hardness mean values more homogeneous between the top and bottom surfaces. The hardness mean values were not statistically significant difference for the pre-cure temperature used. According to these results, the pre-heating of the composite resin provide no influence on Vickers hardness mean values, however the blue LED showed a cure more homogeneous than QTH LCU.

Changes on degree of conversion of dual-cure luting light-cured with blue LED

The indirect adhesive procedures constitute recently a substantial portion of contemporary esthetic restorative treatments. The resin cements have been used to bond tooth substrate and restorative materials. Due to recently introduction of the self-bonding resin luting cement based on a new monomer, filler and initiation technology has become important to study the degree of conversion of these new materials. In the present work the polymerization reaction and the filler content of dual-cured dental resin cements were studied by means of infra-red spectroscopy (FT-IR) and thermogravimetry (TG). Twenty specimens were made in a metallic mold (8 mm diameter x 1 mm thick) from each of 2 cements, PanaviaA (R) F2.0 (Kuraray) and RelyX (TM) Unicem Applicap (3M/ESPE). Each specimen was cured with blue LED with power density of 500 mW/cm(2) for 30 s. Immediately after curing, 24 and 48 h, and 7 days DC was determined. For each time interval 5 specimens were pulverized, pressed with KBr and analyzed with FT-IR. The TG measurements were performed in Netzsch TG 209 under oxygen atmosphere and heating rate of 10A degrees C/min from 25 to 700A degrees C. A two-way ANOVA showed DC (%) mean values statistically significance differences between two cements (p < 0.05). The Tukey`s test showed no significant difference only for the 24 and 48 h after light irradiation for both resin cements (p > 0.05). The Relx-Y (TM) Unicem mean values were significantly higher than PanaviaA (R) F 2.0. The degree of conversion means values increasing with the storage time and the filler content showed similar for both resin cements.

Synthesis of azopolymers with controlled structure and photoinduced birefringence in their LB films

The molecular architecture of azopolymers may be controlled via chemical synthesis and with selection of a suitable film-forming method, which is important for improving their properties for practical uses. Here we address the main challenge of combining the photoinduced birefringence features of azopolymers with the higher thermal and mechanical stabilities of poly(methyl methacrylate) (PMMA) using Atom Transfer Radical Polymerization (ATRP) to synthesize diblock- and triblock-copolymers of an azomonomer and the monomer methyl methacrylate. Langmuir-Blodgett (LB) films made with the copolymers mixed with cadmium stearate displayed essentially the same optically induced birefringence characteristics, in terms of maximum and residual birefringence and time for writing, as the mixed LB films with the homopolymer poly[4-(N-ethyl-N-(2-methacryloxyethyl))amino-2`-chloro-4`-nitroazobenzene] (HPDR13), also synthesized via ATRP. In fact, the controlled architecture of HPDR13 chains led to Langmuir films that could be more closely packed and reach higher collapse pressures than the corresponding films obtained with HPDR13-conv synthesized via conventional radicalar polymerization. This allowed LB films to be fabricated from neat HPDR13, which was not possible with HPDR13-conv. The enhanced organization in the LB films produced with controlled azopolymer chains, however, led to a smaller free volume available for isomerization of the azochromophores, thus yielding a lower photoinduced birefringence than in the HPDR13-conv films. The combination of ATRP synthesis and LB technology is then promising to obtain optical storage in films with improved thermal and mechanical processabilities, though a further degree of control must be sought to exploit film organization while maintaining the necessary free volume in the films. (C) 2008 Elsevier Ltd. All rights reserved.

3D Cell-Migration Studies using Two-Photon Engineered Polymer Scaffolds

We use two-photon polymerization to fabricate 3D scaffolds with precise control over pore size and shape for studying cell migration in 3D. These scaffolds allow movement of cells in all directions. The fabrication, imaging, and quantitative analysis method developed here can be used to do systematic cell studies in 3D.