Polymer Encapsulated Liposomes for Oral Co-Delivery of Curcumin and Hydroxytyrosol

Curcumin (Cur) is a hydrophobic polyphenol from the rhizome of Curcuma spp., while hydroxytyrosol (HT) is a water-soluble polyphenol from Olea europaea. Both show outstanding antioxidant properties but suffer from scarce bioavailability and low stability in biological fluids. In this work, the co-encapsulation of Cur and HT into liposomes was realized, and the liposomal formulation was improved using polymers to increase their survival in the gastrointestinal tract. Liposomes with different compositions were formulated: Type 1, composed of phospholipids and cholesterol; Type 2, also with a PEG coating; and Type 3 providing an additional shell of Eudragit^® S100, a gastro-resistant polymer. Samples were characterized in terms of size, morphology, ζ-potential, encapsulation efficiency, and loading capacity. All samples were subjected to a simulated in vitro digestion and their stability was investigated. The Eudragit^®S100 coating demonstrated prevention of early releases of HT in the mouth and gastric phases, while the PEG shell reduced bile salts and pancreatin effects during the intestinal digestion. In vitro antioxidant activity showed a cumulative effect for Cur and HT loaded in vesicles. Finally, liposomes with HT concentrations up to 40 μM and Cur up to 4.7 μM, alone or in combination, did not show cytotoxicity against Caco-2 cells.     

Strain-specific polyketide synthase genes of Aspergillus niger

In silico comparison of 34 putative pks genes in Aspergillus niger strain CBS 513.88 versus A. niger strain ATCC 1015 genome revealed significant nucleotide identity (>95% covering a minimum of 99% of the gene sequence) for 31 of these genes (approximately 91%). A. niger CBS 513.88 harbors three putative pks genes (An01g01130, An11g05940, and An15g07920), for which nucleotide identity was not found in A. niger ATCC 1015. To compare the results of the in silico analysis with the in vivo situation, experimental data were obtained for a large number of A. niger strains obtained from different substrates and geographical regions. Three putative pks genes that were found to be variable between the two A. niger strains using bioinformatics tools were in fact strain-specific genes based on experimental data. The PCR amplification signals for the An01g01130, An11g05940, and An15g07920 pks genes were detected in only 97%, 71%, and 26% of the strains, respectively. Southern blot analyses confirmed the PCR data. Because one of the strain-specific pks genes (An15g07920) is located in a putative ochratoxin cluster, we focused our investigation on that region. We assessed the ochratoxin production capability of the 119 A. niger strains and found a positive association between the presence of this pks gene and the capability of the respective strain to produce ochratoxin.     

Campylobacter jejuni loss of culturability in aqueous microcosms and ability to resuscitate in a mouse model

Water is known as one of the main transmission routes of Campylobacter and contributes to increase the number of sporadic infections and outbreaks. Campylobacter jejuni persists in the environment, especially in water, in a viable but non-culturable (VBNC) form that is thought to be a possible cause of water-borne outbreaks. In this study, we evaluated the loss of culturability and viability of 9 C. jejuni strains of clinical origin and one ATCC reference strain when kept at 4 degrees C in artificial sea water (ASW). Culturability was measured as colony-forming units while viability was evaluated by CTC-DAPI double staining and the combined CTC-specific fluorescent antibody technique (CTC-FA). When cultured on Columbia Agar plates, strains exhibited different growth profiles which allowed to classify them into three different groups. Both techniques used to monitor the viability of the bacterial cells showed that C. jejuni strains survived in the VBNC form in the microcosms through a period lasting from 138 to 152 days. The recovery of C. jejuni VBNC forms to culturability, as evidenced by cell division, was obtained by passage in the mouse intestine. Our results indicate that C. jejuni VBNC cells were able to remain in this state for a few months and regain their culturability after in vivo passage depending on their lasting in the VBNC state, which affects the number of respiring bacteria. In fact, the resuscitation was achieved when the number of respiring bacteria became higher than 10(4) cell/ml. Therefore, a relatively high microbial titer of respiring bacteria in the VBNC state seems to be important for the resuscitation and subsequent intestinal colonisation.     

Enzymatic de-glycosylation of rutin improves its antioxidant and antiproliferative activities

Bioavailability and biological properties of flavonoid glycosides can be improved after the enzymatic hydrolysis of specific glycosyl groups. In this study, we evaluate the antioxidant and antiproliferative potential of rutin after enzymatic hydrolysis performed by α-l-rhamnosidases (hesperidinase from Penicillium sp. and naringinase from Penicillium decumbens) previously heated at 70 °C for 30 min to inactivate the undesirable β-d-glucosidase activity. The highest in vitro antioxidant activity determined by DPPH radical scavenging was achieved with rutin hydrolyzed by hesperidinase. Rutin was predominantly bioconverted into quercetin-3-glucoside. There was no statistical difference between xanthine oxidase inhibition by rutin before and after hydrolysis. However, in vitro inhibitory activity against ten human tumor cell lines showed that hydrolyzed rutin exerted a more potent antiproliferative effect than quercetin and rutin on various cancer cell lines, specially glioma, and ovarian and breast adenocarcinomas. These results indicate that quercetin-3-glucoside could be a promising functional derivative obtained by rutin hydrolysis.     

Disrupting the methionine biosynthetic pathway in Candida guilliermondii: characterization of the MET2 gene as counter‐selectable marker

Candida guilliermondii (teleomorph Meyerozyma guilliermondii) is an ascomycetous species belonging to the fungal CTG clade. This yeast remains actively studied as a result of its moderate clinical importance and most of all for its potential uses in biotechnology. The aim of the present study was to establish a convenient transformation system for C. guilliermondii by developing both a methionine auxotroph recipient strain and a functional MET gene as selection marker. We first disrupted the MET2 and MET15 genes encoding homoserine‐O‐acetyltransferase and O‐acetylserine O‐acetylhomoserine sulphydrylase, respectively. The met2 mutant was shown to be a methionine auxotroph in contrast to met15 which was not. Interestingly, met2 and met15 mutants formed brown colonies when cultured on lead‐containing medium, contrary to the wild‐type strain, which develop as white colonies on this medium. The MET2 wild‐type allele was successfully used to transfer a yellow fluorescent protein (YFP) gene‐expressing vector into the met2 recipient strain. In addition, we showed that the loss of the MET2‐containing YFP‐expressing plasmid can be easily observed on lead‐containing medium. The MET2 wild‐type allele, flanked by two short repeated sequences, was then used to disrupt the LYS2 gene (encoding the α‐aminoadipate reductase) in the C. guilliermondii met2 recipient strain. The resulting lys2 mutants displayed, as expected, auxotrophy for lysine. Unfortunately, all our attempts to pop‐out the MET2 marker (following the recombination of the bordering repeat sequences) from a target lys2 locus were unsuccessful using white/brown colony colour screening. Nevertheless, this MET2 transformation/disruption system represents a new versatile genetic tool for C. guilliermondii. Copyright © 2014 John Wiley & Sons, Ltd.     

Exposure to polycyclic aromatic hydrocarbons through consumption of edible marine species in Catalonia, Spain

The concentrations of 16 polycyclic aromatic hydrocarbons (PAHs; naphthalene, acenaphtylene, acenaphtene, fluorene, phenanthrene, anthracene, fluoranthene, pyrene, benzo[a]anthracene, chrysene, benzo[b]fluoranthene, benzo[k]fluoranthene, benzo[a]pyrene, indeno[1,2,3-c,d]pyrene, dibenzo[a,h]anthracene, and benzo[g,h,i]perylene) were determined by gas chromatography-mass spectrometry in samples of 14 edible marine species (sardine, tuna, anchovy, mackerel, swordfish, salmon, hake, red mullet, sole, cuttlefish, squid, clam, mussel, and shrimp) collected in March and April 2005. These species are widely consumed by the population of Catalonia, Spain. PAH intake was also estimated for eight age and sex groups of this population. Mussel, clam, and shrimp had the highest PAH concentrations (22.4, 21.5, and 15.9 ng/g of fresh weight, respectively). In contrast, sole (2.5 ng/g of fresh weight) and cuttlefish and squid (both 3.0 ng/g of fresh weight) had the lowest mean concentrations. The highest PAH intake was found in women and girls (5.3 and 5.2 ng/kg/day, respectively), but female adolescents and female seniors had the lowest PAH intakes (3.3 ng/kg/day in both groups). The intake of benzo[a]pyrene and six other PAHs that are probably human carcinogens through consumption of these marine species would be associated with 0.27/10(6) increase in the risk of development of cancer over a 70-year life span.     

A test house study of pesticides and pesticide degradation products following an indoor application

Preexisting pesticide degradates are a concern for pesticide biomonitoring studies as exposure to them may result in overestimation of pesticide exposure. The purpose of this research was to determine whether there was significant formation and movement, of pesticide degradates over a 5‐week period in a controlled indoor setting after insecticide application. Movement of the pesticides during the study was also evaluated. In a simulated crack and crevice application, commercially available formulations of fipronil, propoxur, cis/trans‐permethrin, and cypermethrin were applied to a series of wooden slats affixed to the wall in one room of an unoccupied test house. Floor surface samples were collected through 35 days post‐application. Concentrations of the pesticides and the following degradates were determined: 2‐iso‐propoxyphenol, cis/trans 3‐(2,2‐dichlorovinyl)‐3‐3‐dimethyl‐(1‐cyclopropane) carboxylic acid, 3‐phenoxybenzoic acid, fipronil sulfone, fipronil sulfide, and fipronil desulfinyl. Deltamethrin, which had never been applied, and chlorpyrifos, which had been applied several years earlier, and their degradation products, cis‐3‐(2,2‐dibromovinyl)‐2,2‐dimethylcyclopropane‐1‐carboxylic acid, and, 3,5,6‐trichloro‐2‐pyridinol, respectively, were also measured. Propoxur was the only insecticide with mass movement away from the application site. There was no measurable formation or movement of the degradates. However, all degradates were present at low levels in the formulated product. These results indicate longitudinal repetitive sampling of indoor degradate levels during short‐term studies, is unnecessary.     

Hereditary Prostate Cancer: Genes Related,Target Therapy and Prevention

Prostate cancer (PCa) is globally the second most diagnosed cancer type and the most common cause of cancer-related deaths in men. Family history of PCa, hereditary breast and ovarian cancer (HBOC) and Lynch syndromes (LS), are among the most important risk factors compared to age, race, ethnicity and environmental factors for PCa development. Hereditary prostate cancer (HPCa) has the highest heritability of any major cancer in men. The proportion of PCa attributable to hereditary factors has been estimated in the range of 5–15%. To date, the genes more consistently associated to HPCa susceptibility include mismatch repair (MMR) genes (MLH1, MSH2, MSH6, and PMS2) and homologous recombination genes (BRCA1/2, ATM, PALB2, CHEK2). Additional genes are also recommended to be integrated into specific research, including HOXB13, BRP1 and NSB1. Importantly, BRCA1/BRCA2 and ATM mutated patients potentially benefit from Poly (ADP-ribose) polymerase PARP inhibitors, through a mechanism of synthetic lethality, causing selective tumor cell cytotoxicity in cell lines. Moreover, the detection of germline alterations in MMR genes has therapeutic implications, as it may help to predict immunotherapy benefits. Here, we discuss the current knowledge of the genetic basis for inherited predisposition to PCa, the potential target therapy, and the role of active surveillance as a management strategy for patients with low-risk PCa. Finally, the current PCa guideline recommendations are reviewed.     

巨人的花园——对不同尺度棕地的差异化策略

近几十年来，旧工业区转型的项目不断增加，范围日益扩大，内涵不断扩充，被认为是城市改造的主要途径。工业区的转型不仅是自身的需要，而且经常能提供一个现实的机会来设想与城市在功能和关系上的深远变化。因为实施的连续性和话题的扩展性，这种转型要放在社会、技术和国际化变化的语境下，考虑典型的城市改造过程中的周期性活力，作为持续演变的效果来进行分析。这种改善往往受到多方面影响，包括：该地区最初的设计、与城市中断了的互动、新的机遇以及潜在的需求。所以，转型不仅意味着现存物质空间的改变，往往还作为观念工具，提供区域层面的解决之道。