Advances in polymeric systems for tissue engineering and biomedical applications

The characteristics of tissue engineered scaffolds are major concerns in the quest to fabricate ideal scaffolds for tissue engineering applications. The polymer scaffolds employed for tissue engineering applications should possess multifunctional properties such as biocompatibility, biodegradability and favorable mechanical properties as it comes in direct contact with the body fluids in vivo. Additionally, the polymer system should also possess biomimetic architecture and should support stem cell adhesion, proliferation and differentiation. As the progress in polymer technology continues, polymeric biomaterials have taken characteristics more closely related to that desired for tissue engineering and clinical needs. Stimuli responsive polymers also termed as smart biomaterials respond to stimuli such as pH, temperature, enzyme, antigen, glucose and electrical stimuli that are inherently present in living systems. This review highlights the exciting advancements in these polymeric systems that relate to biological and tissue engineering applications. Additionally, several aspects of technology namely scaffold fabrication methods and surface modifications to confer biological functionality to the polymers have also been discussed. The ultimate objective is to emphasize on these underutilized adaptive behaviors of the polymers so that novel applications and new generations of smart polymeric materials can be realized for biomedical and tissue engineering applications.     

Vacuum infiltration enhances the Agrobacterium-mediated genetic transformation in Indian soybean cultivars

For the first time we have developed a reliable and efficient vacuum infiltration-assisted Agrobacterium-mediated genetic transformation (VIAAT) protocol for Indian soybean cultivars and recovered fertile transgenic soybean plants through somatic embryogenesis. Immature cotyledons were used as an explant and three Agrobacterium tumefaciens strains (EHA 101, EHA 105, and KYRT 1) harbouring the binary vector pCAMBIA1301 were experimented in the co-cultivation. The immature cotyledons were pre-cultured in liquid somatic embryo induction medium prior to vacuum infiltration with the Agrobacterium suspension and co-cultivated for 3 days on co-cultivation medium containing 50 mg l^?1 citric acid, 100 µM acetosyringone, and 100 mg l^?1 l-cysteine. The transformed somatic embryos were selected in liquid somatic embryo induction medium containing 10 mg l^?1 hygromycin and the embryos were germinated in basal medium containing 20 mg l^?1 hygromycin. The presence and integration of the hpt II and gus genes into the soybean genome were confirmed by GUS histochemical assay, polymerase chain reaction, and Southern hybridization. Among the different combinations tested, high transformation efficiency (9.45 %) was achieved when immature cotyledons of cv. Pusa 16 were pre-cultured for 18 h and vacuum infiltrated with Agrobacterium tumefaciens KYRT 1 for 2 min at 750 mm of Hg. Among six Indian soybean cultivars tested, Pusa 16 showed highest transformation efficiency of 9.45 %. The transformation efficiency of this method (VIAAT) was higher than previously reported sonication-assisted Agrobacterium-mediated transformation. These results suggest that an efficient Agrobacterium-mediated transformation protocol for stable integration of foreign genes into soybean has been developed.     

Microbial Communities in Chesapeake & Ohio Canal National Historical Park and Their Function as Indicators of Water Quality

Abstract

The objective of this study was to establish meaningful relationships between prokaryotic community profiles and water quality parameters in different water bodies (spring, stream, cave, and mine) in the middle reach of the Chesapeake & Ohio Canal National Historic Park (C&O), Maryland. The microbial profiles in the water samples were determined using metagenomic analysis. The relationships between microbial phylogenetic profiles and water quality parameters were investigated using principal component analysis (PCA) and redundancy analysis (RDA). The most abundant phyla identified in most samples were Proteobacteria (55.4%), Bacteroidetes (12.3%), Actinobacteria (10.6%), Firmicutes (2.4%), Planktomycetes (1.8%), Verrucomicrobia (1.5%), Chloroflexi (1.5%), and Acidobacteria (1.3%), which are major bacterial and archaeal groups typically observed in natural freshwater environments. PCA showed that water chemistry was determined primarily by the geology of the site and the type of water source (i.e., spring, stream, cave, or mine). Most samples located in carbonate formations correlated with high alkalinity, inorganic carbon, and calcium, representing the typical karstic geochemistry. RDA shows that pH, electrical conductivity, temperature and nutrients including nitrate, phosphate, and sulfate, were significant determinants of the microbial ecology.     

Plasminogen activator: Isolation and purification from Lymphosarcoma of ascites bearing mice

Plasminogen activator secreted by lymphosarcoma (ascites) of mice was purified up to 163-fold by ammonium sulphate fractionation at 35% saturation and chromatography on p-aminobenzamidine-Sepharose 4B. The purified activator contained specific activity of 9980 IU/mg. The plasminogen activator displayed homogeneity by polyacrylamide slab gel electrophoresis and high performance liquid chromatography. The activator consisted of a single polypeptide chain with an apparent molecular weight of 66,000 daltons as determined by sodium dodecyl sulphate-polyacrylamide gel electrophoresis under reducing conditions as well as gel filtration on Sephadex G-100. Distinct differences between this activator and urokinase were discernible in respect of specific activities, fibrin affinity and immunochemical properties. The lymphosarcoma activator appears to be of tissue-type origin since it showed gross similarity to standard tissue plasminogen activator in terms of modes of binding to fibrin and immunological attributes.     

Oxidation of propene and 1-butene byMethylococcus capsulatus andMethylosinus trichosporium

Summary Methane-grown cells ofMethylococcus capsulatus andMethylosinus trichosporium readily oxidized propene and various isomers of butene to their respective epoxides. When examined in a proton NMR spectrum using tris([3-trifluoromethylhydroxymethylene]-d-camphorato), europium III derivative as an optically active chemical shift reagent, the products propylene oxide and 1,2-epoxybutane were found to contain equal amounts of both isomers. Methane-grown cells of both bacteria had considerable levels of reducing equivalents to catalyze the epoxidation of gaseous olefins. Cells depleted of reductants catalyzed the oxidation in the presence of low levels of methanol or formaldehyde with a stoichiometry of about 2:1. The rates of epoxidation of propene and 1-butene in a continuous reactor were 2–3-times that of a batch-wise reaction; the epoxidation activity, however, was lost within 3 h. The inactivation was attributed to the reactivity of the accumulated epoxides in the reactor. Propene and 1-butene oxidation by both bacteria were drastically inhibited by the respective products. Thus, the major problem in the application of microorganisms for production of epoxides from gaseous olefins is the rapid separation of the reactive products.     

Interaction of lysozyme with dyes. II. Binding of bromophenol blue

The binding of lysozyme with bromophenol blue (BPB) at various dye concentrations and pH was carried out at 25 degrees C by equilibrium dialysis, ultraviolet (UV) difference and circular dichroism (CD) spectral techniques. Binding isotherms at pH 5.0 show non-cooperative binding at low dye concentrations, which change over to cooperative binding at higher concentrations indicating biphasic nature. However, binding isotherms at pH 7.0 and 9.0 show cooperative binding only, at all concentrations of the dye. The number of available binding sites decreases with the increase of pH. Gibbs free energy change, calculated on the basis of Wyman's binding potential concept, decreases with the increase of pH. Binding isotherms at pH 5.0 obtained at a lower temperature of 8 degrees C, also indicate the biphasic nature similar to those observed at 25 degrees C, but with a slight decreased strength of binding. The UV difference spectra of the complex do not show any distinct peaks in the 285 to 297 nm region eliminating any possible interaction of BPB with tryptophan and tyrosine residues of the lysozyme molecule. The CD spectra of lysozyme-BPB complex show a decrease in ellipticities with reference to native lysozyme in the near UV and far UV regions. This indicates that the lysozyme-BPB complex has a lower helical content probably due to the conformational changes induced into the native enzyme. The appearance of new positive peaks at 315 nm in the near UV region and at 592 nm in the visible region of the CD spectra may be due to the induced asymmetry into the BPB molecule as a result of its binding to a cationic residue (probably a lysine residue) of lysozyme.(ABSTRACT TRUNCATED AT 250 WORDS)     

FireStem2D – A Two-Dimensional Heat Transfer Model for Simulating Tree Stem Injury in Fires

FireStem2D, a software tool for predicting tree stem heating and injury in forest fires, is a physically-based, two-dimensional model of stem thermodynamics that results from heating at the bark surface. It builds on an earlier one-dimensional model (FireStem) and provides improved capabilities for predicting fire-induced mortality and injury before a fire occurs by resolving stem moisture loss, temperatures through the stem, degree of bark charring, and necrotic depth around the stem. We present the results of numerical parameterization and model evaluation experiments for FireStem2D that simulate laboratory stem-heating experiments of 52 tree sections from 25 trees. We also conducted a set of virtual sensitivity analysis experiments to test the effects of unevenness of heating around the stem and with aboveground height using data from two studies: a low-intensity surface fire and a more intense crown fire. The model allows for improved understanding and prediction of the effects of wildland fire on injury and mortality of trees of different species and sizes.     

Effect of process parameters on 3-hydroxypropionic acid production from glycerol using a recombinant <Emphasis Type="Italic">Escherichia coli</Emphasis>

The stability and specific activity of endo-β-1,4-glucanase III from Trichoderma reesei QM9414 was enhanced, and the expression efficiency of its encoding gene, egl3, was optimized by directed evolution using error-prone PCR and activity screening in Escherichia coli RosettaBlue (DE3) pLacI as a host. Relationship between increase in yield of active enzyme in the clones and improvement in its stability was observed among the mutants obtained in the present study. The clone harboring the best mutant 2R4 (G41E/T110P/K173M/Y195F/P201S/N218I) selected in via second-round mutagenesis after optimal recombinating of first-round mutations produced 130-fold higher amount of mutant enzyme than the transformant with wild-type EG III. Mutant 2R4 produced by the clone showed broad pH stability (4.4–8.8) and thermotolerance (entirely active at 55°C for 30 min) compared with those of the wild-type EG III (pH stability, 4.4–5.2; thermostability, inactive at 55°C for 30 min). k _cat of 2R4 against carboxymethyl-cellulose was about 1.4-fold higher than that of the wild type, though the K _m became twice of that of the wild type.