Thermoreversible protein hydrogel as cell scaffold

A thermoreversible fibrillar hydrogel has been formed from an aqueous lysozyme solution in the presence of dithiothreitol (DTT). Its physical properties and potential as a tissue engineering scaffold have been explored. Hydrogels were prepared by dissolving 3 mM protein in a 20 mM DTT/water mixture, heating to 85 degrees C and cooling at room temperature. No gel was observed for the equivalent sample without DTT. The elastic nature of the gel formed was confirmed by rheology, and the storage modulus of our gel was found to be of the same order of magnitude as for other cross-linked biopolymers. Micro differential scanning calorimetry (microDSC) experiments confirmed that the hydrogel was thermally reversible and that gelation and melting occurs through a solid-liquid-like first-order transition. Infrared spectroscopy of the hydrogel and transmission electron microscopy studies of very dilute samples revealed the presence of beta-sheet-rich fibrils that were approximately 4-6 nm in diameter and 1 mum in length. These fibrils are thought to self-assemble along their long axes to form larger fibers that become physically entangled to form the three-dimensional network observed in both cryo-scanning electron microscopy (cryo-SEM) and small-angle neutron scattering (SANS) studies. The hydrogel was subsequently cultured with 3T3 fibroblasts and cells spread extensively after 7 days and stretched actin filaments formed that were roughly parallel to each other, indicating the development of organized actin filaments in the form of stress fibers in cells.     

Sequential IL-23 and IL-17 and increased Mmp8 and Mmp14 expression characterize the progression of an experimental model of periodontal disease in type 1 diabetes

Molecular mechanisms responsible for periodontal disease (PD) and its worsening in type 1 Diabetes Mellitus (DM1) remain unknown. Cytokine profile and expression levels of collagenases, Mmp14, and tissue inhibitors were determined, as were the numbers of neutrophils and macrophages in combined streptozotocin-induced DM1 and ligature-induced PD models. Increased IL-23 (80-fold) and Mmp8 expression (25-fold) was found in DM1. Ligature resulted in an IL-1β/IL-6 profile, increased expression of Mmp8, Mmp13, and Mmp14 (but not Mmp1), and transient expression of Timp1 and Reck in non-diabetics. PD in DM1 involved IL-1β (but not IL-6) and IL-23/IL-17, reduced IL-6 and IL-10, sustained Mmp8 and Mmp14, increased Mmp13 and reduced Reck expression in association with 20-fold higher counts of neutrophils and macrophages. IL-23 and Mmp8 expression are hallmarks of DM1. In association with the IL-1/IL-6 (Th1) response in PD, one found a secondary IL-17 (Th17) pathway in non-diabetic rats. Low IL-6/TNF-α suggest that the Th1 response was compromised in DM1, while IL-17 indicates a prevalence of the Th17 pathway, resulting in high neutrophil recruitment. Mmp8, Mmp13, and Mmp14 expression seems important in the tissue destruction during PD in DM1. PD-associated IL-1/IL-6 (Th1), IL-10, and Reck expression are associated with the acute-to-chronic inflammation transition, which is lost in DM1. In conclusion, IL-23/IL-17 are associated with the PD progression in DM1.     

Yeast as a model system for studying lipid homeostasis and function

Lipids are essential eukaryotic cellular constituents. Lipid metabolism has a strong impact on cell physiology, and despite good progress in this area, many important basic questions remain unanswered concerning the functional diversity of lipid species and on the mechanisms that cells employ to sense and adjust their lipid composition. Combining convenient experimental tractability, a large degree of conservation of metabolic pathways with other eukaryotes and the relative simplicity of its genome, proteome and lipidome, yeast represents the most advantageous model organism for studying lipid homeostasis and function. In this review we will focus on the importance of yeast as a model organism and some of the innovative advantages for the lipid research field.     

Chronic methylphenidate administration alters antioxidant defenses and butyrylcholinesterase activity in blood of juvenile rats

Methylphenidate (MPH), a psychostimulant that affects both dopaminergic and noradrenergic systems, is one of the most frequently prescribed treatments for attention-deficit hyperactivity disorder. The present study investigated the effects of chronic administration of MPH on some parameters of oxidative stress, as well as on butyrylcholinesterase (BuChE) activity in blood of young rats. Rats received intraperitoneal injections of MPH (2.0 mg/kg) once a day, from the 15th to the 45th day of age or an equivalent volume of 0.9% saline solution (controls). Two hours after the last injection, animals were euthanized, and blood was collected. Results demonstrated that MPH did not alter the dichlorofluorescein formed, decreased both thiobarbituric acid reactive substances and total non-enzymatic radical-trapping antioxidant, and increased superoxide dismutase and catalase activities, suggesting that this psychostimulant may alter antioxidant defenses. BuChE activity was increased in blood of juvenile rats subjected to chronic MPH administration. These findings suggest that MPH may promote peripheral oxidative adaptations and cholinergic changes.     

Comparative study with two different enrichments in the culture media used in the disinfectant efficacy assay

Recent changes in Brazilian legislation for commercial disinfectants have been published due to the recent epidemic of nosocomial infections caused by rapidly growing mycobacteria (RGM) in many states of Brazil over the last 8 years. One of these documents requires that all the manufacturers provide evidence of efficacy of sterilizing and disinfectant products, used for semi critical medical devices, against the Mycobacterium bovis BCG Moreau and Mycobacterium abscessus subsp. bolletii INCQS 00594 strains by using the Confirmative in vitro Test for Determining Tuberculocidal Activity of Disinfectants recommended by the Association of Official Analytical Chemists. These changes have caused additional costs and increased problems for importation of enrichment products at national laboratories where disinfectant efficacy assay service is performed. Middlebrook ADC Enrichment (ADC) is provided by a unique manufacturer and used in the official protocol. The aim of the present study was to evaluate an alternative in house low-cost enrichment composed of fetal bovine serum and glucose (FBSG) with ADC for performance of disinfectant efficacy assay against mycobacteria. After obtaining the growth curves for M. abscessus ATCC 19977, M. abscessus subsp. bolletii INCQS 00594, Mycobacterium chelonae ATCC 35752, and Mycobacterium fortuitum ATCC 6841 by using ADC enrichment and FBSG in Kirchners and 7H9 culture media. Through statistical analysis via the Kruskal-Wallis test on the evaluation of microorganism growth rate, it was observed that there was no inhibition of RGM growth by any of the enrichments used. These results suggest that low-cost enrichment FBSG may be used as a potential substitute of ADC for composition of media for mycobacterial growth, including in disinfectant tests.     

Beta-lactams resistance and presence of class 1 integron in Pseudomonas spp. isolated from untreated hospital effluents in Brazil

The aim of the present study was to investigate the resistance profile, to detect the presence of beta-lactam resistance genes, phenotypic expression of efflux pump systems and class 1 integrons in Pseudomonas spp. strains obtained from untreated hospital effluents. Effluent samples were collected from four hospitals in Porto Alegre, RS, Brazil. Pseudomonas were isolated on MacConkey agar plates and the identification was confirmed by 16S rRNA PCR and biochemical tests. Susceptibility testing was determined by disk-diffusion method using 11 different beta-lactams and MIC assays were performed on isolates resistant to imipenem and ceftazidime. The beta-lactamase genes bla _IMP, bla _VIM, bla _SPM-1, bla _OXA-23-like, bla _OXA-24-like, bla _OXA-51-like and the intl1 gene from class 1 integron were analysed by PCR. One hundred and twenty-four isolates were recovered and the most common species was Pseudomonas pseudoalcaligenes. The resistance found among the isolates was considered high, 62 (50%) isolates were multiresistant. No isolate carrying the beta-lactamase genes tested was found among the strains. Seven isolates showed reduction of MIC for imipenem and ceftazidime in the presence of cyanide m-chlorophenylhydrazone, indicating the hyper expression of efflux pumps. From the 124 isolates, 52 (41.9%) were identified as carrying the class 1 integron gene, intI1. Untreated hospital effluents could be a source of environmental contamination due to discharge of antimicrobial resistant bacteria which can carry integron class 1 and act as a reservoir of resistance genes and have efflux pump systems.     

Influence of temperature, light and nutrients on the growth rates of the macroalga Gracilaria domingensis in synthetic seawater using experimental design

In the present study, the daily relative growth rates (DRGR, in percent per day) of the red macroalga Gracilaria domingensis in synthetic seawater was investigated for the combined influence of five factors, i.e., light (L), temperature (T), nitrate (N), phosphate (P), and molybdate (M), using a statistical design method. The ranges of the experimental cultivation conditions were T, 18–26°C; L, 74–162?μmol photons m^?2?s^?1; N, 40–80?μmol?L^?1; P, 8–16?μmol?L^?1; and M, 1–5?nmol?L^?1. The optimal conditions, which resulted in a maximum growth rate of ≥6.4% d^?1 from 7 to 10?days of cultivation, were determined by analysis of variance (ANOVA) multivariate factorial analysis (with a 2⁵ full factorial design) to be L, 74?μmol photons m^?2?s^?1; T, 26°C; N, 80?μmol?L^?1; P, 8?μmol?L^?1; and M, 1?nmol?L^?1. In additional, these growth rate values are close to the growth rate values in natural medium (von Stosch medium), i.e., 6.5–7.0% d^?1. The results analyzed by the ANOVA indicate that the factors N and T are highly significant linear terms, X _L, (α?=?0.05). On the other hand, the only significant quadratic term (X _Q) was that for L. Statistically significant interactions between two different factors were found between T vs. L and N vs. T. Finally, a two-way (linear/quadratic interaction) model provided a quite reasonable correlation between the experimental and predicted DRGR values (R _adjusted ² ?=?0.9540).     

Spatial–Temporal Variation in Orchid Bee Communities (Hymenoptera: Apidae) in Remnants of Arboreal Caatinga in the Chapada Diamantina Region, State of Bahia, Brazil

The spatial and temporal distribution of organisms is a fundamental aspect of biological communities. The present study focused on three remnants of arboreal Caatinga in northeastern Brazil between May, 2009 and April, 2010. A total of 627 euglossine males were captured in traps baited with artificial aromatic compounds. The specimens belonged to 14 species and four genera: Euglossa Latreille, Eulaema Lepeletier, Eufriesea Cockerell, and Exaerete Hoffmannsegg. Eulaema nigrita Lepeletier (41.6), Euglossa carolina Nemésio (15.3%), Eulaema marcii Nemésio (13.6%), and Euglossa melanotricha Moure (12.8%) were the most common species sampled. The distribution of collected specimens per fragment was as follows: Braúna (280?ha)??259 individuals belonging to 14 species; Cambuí (179?ha)??161 individuals from eight species; and Pindoba (100?ha)??207 individuals represented by seven species. Braúna had the highest diversity (H???=?1.91) and estimated species richness. The largest fragment was the main source of the observed variation in species richness and abundance, indicating a non-random pattern of spatial distribution. The analysis of environmental factors indicated that seasonal variation in these factors was the principal determinant of species occurrence and abundance.     

Stressed but stable: canopy loss decreased species synchrony and metabolic variability in an intertidal hard-bottom community

The temporal stability of aggregate community properties depends on the dynamics of the component species. Since species growth can compensate for the decline of other species, synchronous species dynamics can maintain stability (i.e. invariability) in aggregate properties such as community abundance and metabolism. In field experiments we tested the separate and interactive effects of two stressors associated with storminess--loss of a canopy-forming species and mechanical disturbances--on species synchrony and community respiration of intertidal hard-bottom communities on Helgoland Island, NE Atlantic. Treatments consisted of regular removal of the canopy-forming seaweed Fucus serratus and a mechanical disturbance applied once at the onset of the experiment in March 2006. The level of synchrony in species abundances was assessed from estimates of species percentage cover every three months until September 2007. Experiments at two sites consistently showed that canopy loss significantly reduced species synchrony. Mechanical disturbance had neither separate nor interactive effects on species synchrony. Accordingly, in situ measurements of CO(2)-fluxes showed that canopy loss, but not mechanical disturbances, significantly reduced net primary productivity and temporal variation in community respiration during emersion periods. Our results support the idea that compensatory dynamics may stabilise aggregate properties. They further suggest that the ecological consequences of the loss of a single structurally important species may be stronger than those derived from smaller-scale mechanical disturbances in natural ecosystems.