Peptide degradation is a critical determinant for cell-penetrating peptide uptake

Cell-penetrating peptide mediated uptake of labels appears to follow an equilibrium-like process. However, this assumption is only valid if the peptides are stabile. Hence, in this study we investigate intracellular and extracellular peptide degradation kinetics of two fluorescein labeled cell-penetrating peptides, namely MAP and penetratin, in Chinese hamster ovarian cells. The degradation and uptake kinetics were assessed by RP-HPLC equipped with a fluorescence detector. We show that MAP and penetratin are rapidly degraded both extracellularly and intracellularly giving rise to several degradation products. Kinetics indicates that intracellularly, the peptides exist in (at least) two distinct pools: one that is immediately degraded and one that is stabile. Moreover, the degradation could be decreased by treating the peptides with BSA and phenanthroline and the uptake was significantly reduced by cytochalasin B, chloroquine and energy depletion. The results indicate that the extracellular degradation determines the intracellular peptide concentration in this system and therefore the stability of cell-penetrating peptides needs to be evaluated.     

Molecular and biotechnological aspects of xylanases

Hemicellulolytic microorganisms play a significant role in nature by recycling hemicellulose, one of the main components of plant polysaccharides. Xylanases (EC 3.2.1.8) catalyze the hydrolysis of xylan, the major constituent of hemicellulose. The use of these enzymes could greatly improve the overall economics of processing lignocellulosic materials for the generation of liquid fuels and chemicals. Recently cellulase-free xylanases have received great attention in the development of environmentally friendly technologies in the paper and pulp industry. In microorganisms that produce xylanases low molecular mass fragments of xylan and their positional isomers play a key role in regulating its biosynthesis. Xylanase and cellulase production appear to be regulated separately, although the pleiotropy of mutations, which causes the elimination of both genes, suggests some linkage in the synthesis of the two enzymes. Xylanases are found in a cornucopia of organisms and the genes encoding them have been cloned in homologous and heterologous hosts with the objectives of overproducing the enzyme and altering its properties to suit commercial applications. Sequence analyses of xylanases have revealed distinct catalytic and cellulose binding domains, with a separate non-catalytic domain that has been reported to confer enhanced thermostability in some xylanases. Analyses of three-dimensional structures and the properties of mutants have revealed the involvement of specific tyrosine and tryptophan residues in the substrate binding site and of glutamate and aspartate residues in the catalytic mechanism. Many lines of evidence suggest that xylanases operate via a double displacement mechanism in which the anomeric configuration is retained, although some of the enzymes catalyze single displacement reactions with inversion of configuration. Based on a dendrogram obtained from amino acid sequence similarities the evolutionary relationship between xylanases is assessed. In addition the properties of xylanases from extremophilic organisms have been evaluated in terms of biotechnological applications.     

Emission enhancement in the luminescence performance of warm red light-emitting LiSrVO4:Pr3+ phosphors

Warm red-emitting praseodymium-doped LiSrVO₄ phosphors were synthesized via solid-state reaction. The phase formation was verified using an X-ray diffraction study and the morphology was investigated using a scanning electron microscope study. The LiSrVO₄:Pr³⁺ phosphors emitted red light when exposed to ultraviolet light, indicating their possibility for use in warm white light-emitting diodes (WLEDs). Furthermore, the effect of charge compensators on the luminescence characteristics was addressed. The decay time was investigated using time-resolved photoluminescence. Furthermore, thermal quenching was analyzed through temperature-dependent photoluminescence spectra. Their sensitivity was calculated using temperature-dependent decay time analysis. The colour purity of the emitted light could be measured by photometric analysis. This comprehensive investigation provides a thorough understanding of the luminescence properties of phosphors for WLED applications.     

Allergen Challenge Induces Ifng Dependent GTPases in the Lungs as Part of a Th1 Transcriptome Response in a Murine Model of Allergic Asthma

According to the current paradigm, allergic airway inflammation is mediated by Th2 cytokines and pro-inflammatory chemokines. Since allergic inflammation is self-limited, we hypothesized that allergen challenge simultaneously induces anti-inflammatory genes to counter-balance the effects of Th2 cytokines and chemokines. To identify these putative anti-inflammatory genes, we compared the gene expression profile in the lungs of ragweed-sensitized mice four hours after challenge with either PBS or ragweed extract (RWE) using a micro-array platform. Consistent with our hypothesis, RWE challenge concurrently upregulated Th1-associated early target genes of the Il12/Stat4 pathway, such as p47 and p65 GTPases (Iigp, Tgtp and Gbp1), Socs1, Cxcl9, Cxcl10 and Gadd45g with the Th2 genes Il4, Il5, Ccl2 and Ccl7. These Th1-associated genes remain upregulated longer than the Th2 genes. Augmentation of the local Th1 milieu by administration of Il12 or CpG prior to RWE challenge further upregulated these Th1 genes. Abolition of the Th1 response by disrupting the Ifng gene increased allergic airway inflammation and abrogated RWE challenge-induced upregulation of GTPases, Cxcl9, Cxcl10 and Socs1, but not Gadd45g. Our data demonstrate that allergen challenge induces two sets of Th1-associated genes in the lungs: 1) Ifng-dependent genes such as p47 and p65 GTPases, Socs1, Cxcl9 and Cxcl10 and 2) Ifng-independent Th1-inducing genes like Gadd45g. We propose that allergen-induced airway inflammation is regulated by simultaneous upregulation of Th1 and Th2 genes, and that persistent unopposed upregulation of Th1 genes resolves allergic inflammation.     

Do Interventions Designed to Support Shared Decision-Making Reduce Health Inequalities? A Systematic Review and Meta-Analysis

Background

Increasing patient engagement in healthcare has become a health policy priority. However, there has been concern that promoting supported shared decision-making could increase health inequalities.

Objective

To evaluate the impact of SDM interventions on disadvantaged groups and health inequalities.

Design

Systematic review and meta-analysis of randomised controlled trials and observational studies.

Data Sources

CINAHL, the Cochrane Register of Controlled Trials, the Cochrane Database of Systematic Reviews, EMBASE, HMIC, MEDLINE, the NHS Economic Evaluation Database, Open SIGLE, PsycINFO and Web of Knowledge were searched from inception until June 2012.

Study Eligibility Criteria

We included all studies, without language restriction, that met the following two criteria: (1) assess the effect of shared decision-making interventions on disadvantaged groups and/or health inequalities, (2) include at least 50% of people from disadvantaged groups, except if a separate analysis was conducted for this group.

Results

We included 19 studies and pooled 10 in a meta-analysis. The meta-analyses showed a moderate positive effect of shared decision-making interventions on disadvantaged patients. The narrative synthesis suggested that, overall, SDM interventions increased knowledge, informed choice, participation in decision-making, decision self-efficacy, preference for collaborative decision making and reduced decisional conflict among disadvantaged patients. Further, 7 out of 19 studies compared the intervention''s effect between high and low literacy groups. Overall, SDM interventions seemed to benefit disadvantaged groups (e.g. lower literacy) more than those with higher literacy, education and socioeconomic status. Interventions that were tailored to disadvantaged groups'' needs appeared most effective.

Conclusion

Results indicate that shared decision-making interventions significantly improve outcomes for disadvantaged patients. According to the narrative synthesis, SDM interventions may be more beneficial to disadvantaged groups than higher literacy/socioeconomic status patients. However, given the small sample sizes and variety in the intervention types, study design and quality, those findings should be interpreted with caution.     

Ascominuta lignicola, a new loculoascomycete from submerged wood in Hong Kong

Ascominuta lignicola gen. et sp. nov. is described and illustrated based on a specimen collected on submerged wood in Hong Kong.Ascominuta lignicola is characterised by relatively small ascomata, globose, 4-spored asci, and ascospores with an elaborate sheath.Ascominuta lignicola is compared withMassarina, Mycosphaerella andWettsteinina. The placement ofAscominuta in the Ascomycetesincertae sedis is discussed.     

Zinc finger nucleases: custom-designed molecular scissors for genome engineering of plant and mammalian cells

Custom-designed zinc finger nucleases (ZFNs), proteins designed to cut at specific DNA sequences, are becoming powerful tools in gene targeting—the process of replacing a gene within a genome by homologous recombination (HR). ZFNs that combine the non-specific cleavage domain (N) of FokI endonuclease with zinc finger proteins (ZFPs) offer a general way to deliver a site-specific double-strand break (DSB) to the genome. The development of ZFN-mediated gene targeting provides molecular biologists with the ability to site-specifically and permanently modify plant and mammalian genomes including the human genome via homology-directed repair of a targeted genomic DSB. The creation of designer ZFNs that cleave DNA at a pre-determined site depends on the reliable creation of ZFPs that can specifically recognize the chosen target site within a genome. The (Cys₂His₂) ZFPs offer the best framework for developing custom ZFN molecules with new sequence-specificities. Here, we explore the different approaches for generating the desired custom ZFNs with high sequence-specificity and affinity. We also discuss the potential of ZFN-mediated gene targeting for ‘directed mutagenesis’ and targeted ‘gene editing’ of the plant and mammalian genome as well as the potential of ZFN-based strategies as a form of gene therapy for human therapeutics in the future.     

Hydrolysis of cellulose derived from steam exploded bagasse by Penicillium cellulases: Comparison with commercial cellulase

A complete cellulase from Penicillium pinophilum was evaluated for the hydrolysis of α-cellulose derived from steam exploded sugarcane bagasse and other cellulosic substrates. α-Cellulose at 1% substrate concentration was completely hydrolyzed by Penicillium cellulase within 3 h wherein at 10% the hydrolysis was 100% within 24 h with an enzyme loading of 10 FPU/g. The hydrolysate yielded glucose as major end product as analyzed by HPLC. Under similar conditions, hydrolysis of Sigmacell (microcrystalline cellulose), CP-123 (pulverized cellulose powder) and ball milled Solka Floc were 42%, 56% and 52%, respectively. Further the hydrolysis performance of Penicillium sp. cellulase is compared with Trichoderma reesei cellulase (Accellerase^TM 1000) from Genencore. The kinetics of hydrolysis with respect to enzyme and substrate concentration will be presented.