Quantitative real-time RT-PCR and chromogenic in situ hybridization: precise methods to detect HER-2 status in breast carcinoma

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Chromogenic in situ hybridization compared with other approaches to evaluate HER2/neu status in breast carcinomas

Human epidermal growth factor receptor 2 (HER2) has been evaluated in breast cancer patients to identify those most likely to benefit from herceptin-targeted therapy. HER2 amplification, detected in 20-30% of invasive breast tumors, is associated with reduced survival and metastasis. The most frequently used technique for evaluating HER2 protein status as a routine procedure is immunohistochemistry (IHC). HER2 copy number alterations have also been evaluated by fluorescence in situ hybridization (FISH) in moderate immunoexpression (IHC 2+) cases. An alternative procedure to evaluate gene amplification is chromogenic in situ hybridization (CISH), which has some advantages over FISH, including the correlation between HER2 status and morphological features. Other methodologies have also been used, such as silver-enhanced in situ hybridization (SISH) and quantitative real-time RT-PCR, to determine the number of HER2 gene copies and expression, respectively. Here we will present a short and comprehensive review of the current advances concerning HER2 evaluation in human breast cancer.

Giardia duodenalis: protein substrates degradation by trophozoite proteases

The present investigation was undertaken to identify and characterize trophozoite proteases of five axenic strains of Giardia duodenalis isolated in Brazil and the reference strain Portland 1 isolated in the United States. Trophozoite cell lysates of each strain were analysed for the pattern of proteins and for proteolytic activity. Samples were tested in SDS-polyacrylamide gel electrophoresis for the protein profiles, and the detection of proteases in cell lysates was performed using substrate gel electrophoresis [gelatin, collagen, bovine serum albumin (BSA) and haemoglobin] and azocasein assays. Indeed, synthetic inhibitors were included in the assays to characterize the protease classes. Differences on the hydrolysis patterns of protein substrates were observed in relation to the substrate composition as much as the Giardia trophozoite strain. The substrate-containing gels revealed hydrolysis bands with molecular masses ranging from > 97 to 20-15 kDa, and most zones were common to the five strains. However, some pronounced differences could be detected in the BTU-11 pattern. Azocasein was also degraded; however, depending on the lysate assayed, the degree of substrate degradation was variable. It was observed that inhibitory effects are substrate-dependent since the activity was predominantly due to cysteine proteases against gelatin, collagen, BSA and azocasein substrates and due to serine against haemoglobin. The presence of aspartic protease and aminopeptidase activity in the lysates was also indicated.

Colonization of rocky and leaf pack substrates by benthic macroinvertebrates in a stream in southeast Brazil.

A comparative analysis was conducted of the colonization by benthic macroinvertebrates of rocky and leaf pack substrates, both natural and artificial. This colonization was evaluated by season, with the objective of ascertaining the influence of rainfall on the rate of colonization. The total density of macroinvertebrates after 21 days of colonization was significantly greater in the dry than in the wet season. When the substrate types were compared, artificial leaf pack substrate presented the smallest density for both seasons. In the wet season, Chironomidae, Leptohyphidae, Hydropsychidae, Elmidae, immature stages of Trichoptera, and Hydroptilidae showed a more representative density. In the dry season, Chironomidae, Baetidae and Oligochaeta were the most abundant taxa. The artificial rocky substrate used in this experiment was the most appropriate, due to its resemblance with natural substrate conditions in terms of the maintenance of the structural integrity of the substrate throughout the experimental period. Successional and seasonal effects were of great relevance, playing an important role in the colonization process.

Biomonitoring of substrates containing sewage sludge: assessment of the feasibility in using the diplopod Rhinocricus padbergi.

Due to the constant release of potentially harmful substances in the environment, it is necessary the use of biomonitoring as a form to assess the impact of these substances on animals that inhabit this environment. Therefore, there is a better understanding of the possible effects that these exogenous substances may cause in the organisms present there. In the present study it was verified the feasibility of the use of the diplopod Rhinocricus padbergi as bioindicator of impacted soils by their exposition to substrates containing different concentrations of sewage sludge from different Sewage Treatment Stations (STSs). It was observed animals' behavior and the survival rate was analyzed by log-rank test (p=0,05). The analysis showed that the animals exposed to pure sewage sludge presented higher mortality index than the specimens exposed to different concentrations of the sludge mixed with soil. In general, the survival rate and the analysis of the behavior of the diplopod R. padbergi showed efficiency of this species in biomonitoring impacted soils.

Influence of Different Substrates on the Production of a Mutant Thermostable Glucoamylase in Submerged Fermentation

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Production of cyclodextrins by CGTase from Bacillus clausii using different starches as substrates

Cyclodextrins ( CDs) are cyclic oligasaccharides composed by D- glucose monomers joined by alpha- 1,4-D glicosidic linkages. The main types of CDs are alpha-,beta-and gamma-CDs consisting of cycles of six, seven, and eight glucose monomers, respectively. Their ability to form inclusion complexes is the most important characteristic, allowing their wide industrial application. The physical property of the CD-complexed compound can be altered to improve stability, volatility, solubility, or bio-availability. The cyclomaltodextrin glucanotransferase ( CGTase, EC 2.4.1.19) is an enzyme capable of converting starch into CD molecules. In this work, the CGTase produced by Bacillus clausii strain E16 was used to produce CD from maltodextrin and different starches ( commercial soluble starch, corn, cassava, sweet potato, and waxy corn starches) as substrates. It was observed that the substrate sources influence the kind of CD obtained and that this CGTase displays a beta- CGTase action, presenting a better conversion of soluble starch at 1.0%, of which 80% was converted in CDs. The ratio of total CD produced was 0: 0.89: 0.11 for alpha/beta/gamma. It was also observed that root and tuber starches were more accessible to CGTase action than seed starch under the studied conditions.

Effect of Carbamide Peroxide Bleaching Gel Concentration on the Bond Strength of Dental Substrates and Resin Composite

Purpose: This study evaluated the effect of bleaching gel containing 10%, 15% and 20% carbamide peroxide (CP) on the bond strength of dental enamel or dentin and resin composite restorations.Methods: The buccal surfaces of 12 bovine tooth crowns were conditioned with 37% phosphoric acid, and the adhesive resin Single Bond 2 and the resin composite Filtek Z350 were used to perform the restorations. The blocks were sectioned to obtain bar specimens. Each specimen group (enamel-E, dentin-D) was divided into four subgroups (n=15): S-artificial saliva; 10-10% CP bleaching; 15-15% CP bleaching; 20-20% CP bleaching. CP was applied for six hours daily for two weeks. The specimens were submitted to the a test in a universal testing machine. The data were analyzed by one-way ANOVA and the Tukey post-hoc test and a correlation analysis (r) was performed.Results: For Group E, the mean value (+/- standard-deviation) was 21.86 (+/- 6.03)a, 18.91 (+/- 8.31)ab, 15.43 (+/- 7.44)b and 10.6 (+/- 4.94)c for ES, E10, E15 and E20, respectively. For Group D, the alpha values were 34.73 (+/- 4.68)a, 35.12 (+/- 13.43)a, 29.67 (+/- 6.84)ab and 24.56 (+/- 6.54)b for DS, D10, D15 and D20, respectively. A negative correlation between the CP concentration and mean values was observed for both the enamel (r=-0.95) and dentin (r=-0.85) groups.Conclusion: In the current study, the bond strength of the restoration to enamel and the restoration to dentin were influenced by the application of CP and was dependent on the CP concentration in the bleaching gel.

Ferroelectric characteristics of SrBi4Ti4O15 thin films grown on Pt/Ti/SiO2/Si substrates by the soft chemical method

Ferroelectric SrBi4Ti4O15 thin films were successfully prepared on a Pt(111)/Ti/SiO2/Si(100) substrate for the first time by spin coating, using the polymeric precursor method. X-ray diffraction patterns of the films indicate that they are polycrystalline in nature. Atomic force microscopy (AFM) analyses showed that the surface of these films is smooth, dense and crack-free with low surface roughness (6.4 nm). At room temperature and at a frequency of 1 MHz, the dielectric constant and the dissipation factor were, respectively, 150 and 0.022. The C-V characteristics of perovskite thin film prepared at low temperature show normal ferrolectric behaviour. The remanent polarization and coercive field for the films deposited were 5.4 mu C/cm(2) and 8 5 kV/cm, respectively. All the capacitors showed good polarization fatigue characteristics at least up to 1 x 10(10) bipolar pulse cycles indicating that SrBi4Ti4O15 thin films can be a promising material for use in nonvolatile memories. (c) 2005 Elsevier B.V. All rights reserved.

CULTIVATION of THE EDIBLE MUSHROOM OUDEMANSIELLA CANARII (JUNGH.) HÖHN. IN LIGNOCELLULOSIC SUBSTRATES

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Reuse of sand, crushed sugarcane and peanut hull-based substrates for cherry tomato cultivation

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Effect of different substrates on composting of poultry litter

The objective was to evaluate the differences between distinct types of litter material and their combinations in the dynamics of degradation on the organic matter fractions and the quality of the final compound. The treatments were established according to material used as substrate for broiler litter: treatment 1 - rice husks; 2 - sugar cane bagasse; 3 - wood shavings; 4 - wood shavings + sugar cane bagasse; 5 - rice husks + sugar cane bagasse; and 6 - Napier grass. The following variables were monitored: temperature, levels of total solids (TS), volatile solids (VS), mass and volume of the pile, fibrous fraction, and levels and reductions of N, P and K during the process. Piles formed with Napier grass and sugar cane bagasse presented the highest average temperatures during composting. The greater average reductions in TS and VS were attained in piles with sugar cane bagasse (68.12 and 73.07%, for TS and VS, respectively). The reductions of greatest volume occurred in piles with sugar cane bagasse (52.08%), followed by Napier grass (50.56%). Poultry litters composed of rice husks and wood shavings presented 13.21 and 10.23% of lignin, respectively, which contributed to the lower degradation of fibrous fraction and degradability. Substrates with lower lignin content were those with greatest organic matter degradation rate and had reduced losses of N levels during the process. Composting performance is affected by the initial substrate used to compose the poultry litter.

Oriented growth of Bi3.25La0.75Ti3O12 thin films on RuO2/SiO2/Si substrates by using the polymeric precursor method: Structural, microstructural and electrical properties

c-axis oriented Bi3.25La0.75Ti3O12 (BLT) thin films were grown on a RuO2 top electrode deposited on a (100) SiO2/Si substrate by the polymeric precursor method. X-ray diffraction and atomic force microscope investigations indicate that the films exhibit a dense, well crystallized microstructure having random orientations with a rather smooth surface morphology. The electrical properties of preferred oriented Bi3.25La0.75Ti3O12 (BLT) thin films deposited on RuO2 bottom electrode leaded to a large remnant polarization (P-r ) of 17.2 mu C/cm(2) and (V-c ) of 1.8 V, fatigue free characteristics up to 10(10) switching cycles and a current density of 2.2 mu A/cm(2) at 5 V. We found that the polarization loss is insignificant with nine write/read voltages at a waiting time of 10,000 s. Independently of the applied electric field the retained switchable polarization approached a nearly steady-state value after a retention time of 10 s.

Photoreflectance studies of optical transitions in cubic GaN grown on GaAs(001) substrates

The optical properties of cubic GaN epitaxial layers were investigated by modulated photoreflectance (PR) and photoluminescence in the temperature interval from 5 to 300 K. The epilayers were grown on GaAs(001) substrates by molecular beam epitaxy using a nitrogen RIF-activated plasma source. The PR spectra show a transition which is well fitted using the third-derivative functional form of the unperturbed dielectric function, which we interpret as band-to-band transition. Our results allow determination of the temperature dependence of the main gap of c-GaN and give insights into the residual strain in the film, as well as allow us to estimate the binding energy of the complex formed by an exciton bound to a neutral acceptor. (C) 2003 Elsevier B.V. B.V. All rights reserved.