A three-dimensional model of the human blood-brain barrier to analyse the transport of nanoparticles and astrocyte/endothelial interactions [version 1; referees: 2 approved with reservations]

The aim of this study was to develop a three-dimensional (3D) model of the human blood-brain barrier in vitro, which mimics the cellular architecture of the CNS and could be used to analyse the delivery of nanoparticles to cells of the CNS. The model includes human astrocytes set in a collagen gel, which is overlaid by a monolayer of human brain endothelium (hCMEC/D3 cell line). The model was characterised by transmission electron microscopy (TEM), immunofluorescence microscopy and flow cytometry. A collagenase digestion method could recover the two cell types separately at 92-96% purity. Astrocytes grown in the gel matrix do not divide and they have reduced expression of aquaporin-4 and the endothelin receptor, type B compared to two-dimensional cultures, but maintain their expression of glial fibrillary acidic protein. The effects of conditioned media from these astrocytes on the barrier phenotype of the endothelium was compared with media from astrocytes grown conventionally on a two-dimensional (2D) substratum. Both induce the expression of tight junction proteins zonula occludens-1 and claudin-5 in hCMEC/D3 cells, but there was no difference between the induced expression levels by the two media. The model has been used to assess the transport of glucose-coated 4nm gold nanoparticles and for leukocyte migration. TEM was used to trace and quantitate the movement of the nanoparticles across the endothelium and into the astrocytes. This blood-brain barrier model is very suitable for assessing delivery of nanoparticles and larger biomolecules to cells of the CNS, following transport across the endothelium

Glucose-coated gold nanoparticles transfer across human brain endothelium and enter astrocytes in vitro

The blood-brain barrier prevents the entry of many therapeutic agents into the brain. Various nanocarriers have been developed to help agents to cross this barrier, but they all have limitations, with regard to tissue-selectivity and their ability to cross the endothelium. This study investigated the potential for 4 nm coated gold nanoparticles to act as selective carriers across human brain endothelium and subsequently to enter astrocytes. The transfer rate of glucose-coated gold nanoparticles across primary human brain endothelium was at least three times faster than across non-brain endothelia. Movement of these nanoparticles occurred across the apical and basal plasma membranes via the cytosol with relatively little vesicular or paracellular migration; antibiotics that interfere with vesicular transport did not block migration. The transfer rate was also dependent on the surface coating of the nanoparticle and incubation temperature. Using a novel 3-dimensional co-culture system, which includes primary human astrocytes and a brain endothelial cell line hCMEC/D3, we demonstrated that the glucose-coated nanoparticles traverse the endothelium, move through the extracellular matrix and localize in astrocytes. The movement of the nanoparticles through the matrix was >10 µm/hour and they appeared in the nuclei of the astrocytes in considerable numbers. These nanoparticles have the correct properties for efficient and selective carriers of therapeutic agents across the blood-brain barrier

Evidence of Long Term Benefit of Praziquantel Treatment Against Schistosoma mansoni in Kigungu Fishing Village of Entebbe, Uganda

Praziquantel (PZQ) is efficacious against all species of schistosome: Schistosoma mansoni; Schistosoma haematobium; Schistosoma japonicum and other parasites like the Taenia species. This cross-sectional cohorts study was carried out in Kigungu fishing village along Lake Victoria shore in Entebbe Uganda. Our analysis was based on examining microscopically three slides from a single stool specimen from each of base line cohorts 945.These included children and adults, participants from both sexes in Kigungu fishing village in Entebbe Uganda. Nine hundred and one (901) of the cohorts were re-examined after six months and 625 of the same cohorts who were examined at the baseline and after six months were re-examined 18 months later. The slides were prepared using modified Kato/Katz (Odongo-Aginya) method. The infection proportion with Schistosoma mansoni at baseline was 448 (47.5%) but this was reduced to 244 (25.8%) 18 months after treatment with a single oral dose of praziquantel at 40mg/kg. However 495 (52.5%) were negative at the baseline study. The cure proportion after six was significant {(P=0.00), (OR4.63) CI at 95% (3.53-6.06)}. Similarly the cure proportion after 18 months was significant {(P=0.00), (OR2.2) CI at 95% (1.87-3.34)}. The force of re-infection after six months was significant {(P=0.0001), (OR 0.47) CI at 95% (0.31-0.71)}. Nevertheless the force of re-infection was not significant after 18 months {(P=0.766), (OR 0.95) CI at 95% (0.68-1.34)} eggs excretion did not reach the level of the pre-treatment intensity. The egg reduction was 69.3%. This was associated with age and pre-treatment intensity < 400 eggs per gram (epg) of faeces and age groups ≥ 30 years. The egg reduction also resulted in marked decrease in clinical symptoms in the participants. Our study suggests evidence of long-term benefit of praziquantel in Kigungu and that the re-infection occurred more commonly in younger age group than in the older patients.Key words: Praziquantel; Schistosoma mansoni; Kigungu; Entebbe; Uganda

Design management: changing roles of the professions

This paper sets out to explore how recent changes in procurement in construction have affected the roles that professions play in the design process. It discusses how professions that traditionally took the role of design manager now find themselves participating within previously unforeseen contexts, working in multidisciplinary teams led by contractors and with changed responsibilities at the design stage. Supply chain members who were not previously involved during the early project phases are being engaged at the earliest phases of the project life cycle and even taking leadership roles while designers sometimes work as supply chain partners. A study of design in construction and other sectors shows that in dealing with design management issues it is critical to deepen appreciation for the unique characteristics of design and the design process. The paper argues that contractors and designers taking on design management roles in a dynamic industry seeking to explore best practice and innovative approaches to procurement and in the delivery of projects need to acquire new skills, management education and develop the necessary qualities

Gene therapy for Wiskott-Aldrich syndrome in a severely affected adult

Until recently, hematopoietic stem cell transplantation was the only curative option for Wiskott-Aldrich syndrome (WAS). The first attempts at gene therapy for WAS using a \u3d2-retroviral vector improved immunological parameters substantially but were complicated by acute leukemia as a result of insertional mutagenesis in a high proportion of patients. More recently, treatment of children with a state-of-the-art self-inactivating lentiviral vector (LV-w1.6 WASp) has resulted in significant clinical benefit without inducing selection of clones harboring integrations near oncogenes. Here, we describe a case of a presplenectomized 30-year-old patient with severe WAS manifesting as cutaneous vasculitis, inflammatory arthropathy, intermittent polyclonal lymphoproliferation, and significant chronic kidney disease and requiring long-term immunosuppressive treatment. Following reduced-intensity conditioning, there was rapid engraftment and expansion of a polyclonal pool of transgene-positive functional T cells and sustained gene marking in myeloid and B-cell lineages up to 20 months of observation. The patient was able to discontinue immunosuppression and exogenous immunoglobulin support, with improvement in vasculitic disease and proin-flammatory markers. Autologous gene therapy using a lentiviral vector is a viable strategy for adult WAS patients with severe chronic disease complications and for whom an allogeneic procedure could present an unacceptable risk. This trial was registered at www.clinicaltrials.gov as #NCT01347242

The rs10993994 risk allele for prostate cancer results in clinically relevant changes in microseminoprotein-beta expression in tissue and urine

Microseminoprotein-beta (MSMB) regulates apoptosis and using genome-wide association studies the rs10993994 single nucleotide polymorphism in the MSMB promoter has been linked to an increased risk of developing prostate cancer. The promoter location of the risk allele, and its ability to reduce promoter activity, suggested that the rs10993994 risk allele could result in lowered MSMB in benign tissue leading to increased prostate cancer risk

Vasectomy by ligation and excision, with or without fascial interposition: a randomized controlled trial [ISRCTN77781689]

BACKGROUND: Randomized controlled trials comparing different vasectomy occlusion techniques are lacking. Thus, this multicenter randomized trial was conducted to compare the probability of the success of ligation and excision vasectomy with, versus without, fascial interposition (i.e. placing a layer of the vas sheath between two cut ends of the vas). METHODS: The trial was conducted between December 1999 and June 2002 with a single planned interim analysis. Men requesting vasectomies at eight outpatient clinics in seven countries in North America, Latin America, and Asia were included in the study. The men were randomized to receive vasectomy with versus without fascial interposition. All surgeons performed the vasectomies using the no-scalpel approach to the vas. Participants had a semen analysis two weeks after vasectomy and then every four weeks up to 34 weeks. The primary outcome measure was time to azoospermia. Additional outcome measures were time to severe oligozoospermia (<100 000 sperm/mL) and vasectomy failure based on semen analyses. RESULTS: We halted recruitment after the planned interim analysis, when 841 men had been enrolled. Fascial interposition decreased time to azoospermia (hazard ratio [HR], 1.35; P < 0.0001) and time to severe oligozoospermia (HR, 1.32; P < 0.0001) and reduced failures based on semen analysis by about half, from 12.7% (95% confidence interval [CI], 9.7 to 16.3) to 5.9% (95% CI, 3.8 to 8.6) (P < 0.0001). Older men benefited less from fascial interposition than younger men in terms of the speed of achieving azoospermia. However, the number of vasectomy failures was reduced to a similar degree in all age groups. Slightly more adverse events occurred in the fascial interposition group, but the difference was not significant. These failure rates may appear high to practitioners in countries such as the USA, but they are similar to results from other careful studies of ligation and excision techniques. CONCLUSION: Fascial interposition significantly improves vasectomy success when ligation and excision is the method of vas occlusion. A limitation of this study is that the correlation between postvasectomy sperm concentrations and risk of pregnancy is not well quantified

AGRONOMIC QUALITIES OF GENETIC PYRAMIDS OF COMMON BEAN DEVELOPED FOR MULTIPLE-DISEASE-RESISTANCE

Multiple co-infections by different pathogens on common bean ( Phaseolus vulgaris L.) affect its productivity and cause complete crop loss in susceptible varieties. Therefore, gene pyramiding using marker assisted selection (MAS) and backcrossing, provide alternative cost-effective control measures to bean diseases. However, in the process of developing pyramids, linkage drags were likely to affect the qualities of progeny lines, hence, special attention was paid to this situation. The objective of this study was thus to assess the agronomic qualities of advanced genetic pyramids developed from a four-way cross for multiple disease resistance. The disease resistance genes (R) pyramided from four parents were: Co42 and Co-5 from G2333; Phg-2 from MEX54; Pythium ultimum Dennis from MLB49-89A and I &amp; bc3 from MCM5001. The progeny lines were planted in an incomplete block design, and replicated thrice for two seasons (2015A and 2015B) in fields at CIAT, Kawanda in Uganda. Agronomic traits were highly heritable (0.6), except number of pods per plant (&lt; 0.3). Backcrossing generated high-yielding bean lines, with 270 - 290 seed per plant and early maturity (95-100 days). Nine superior lines with desirable qualities, such as earliness (95 days), high seed rate (290 seeds per plant), and climbing ability, were obtained. Pyramiding R genes did not affect yield traits, except time to flowering and number of flower buds per plant due to transgressive segregation.Les co-infections multiples par de diff\ue9rents pathog\ue8nes sur le haricot commun ( Phaseolus vulgaris L.) affectent sa productivit\ue9 et causent la perte totale des vari\ue9t\ue9s susceptibles de la culture. Par cons\ue9quent, la pyramide des g\ue8nes en utilisant la s\ue9lection assist\ue9e par des marqueurs (MAS) et le r\ue9trocroisement, fournissent des mesures alternatives de contr\uf4le moins ch\ue8res des maladies du haricot. N\ue9anmoins, dans le processus du d\ue9veloppement des pyramides, les poids des liaisons affectent probablement les qualit\ue9s des lign\ue9es de prog\ue9nitures, de ce fait, une attention particuli\ue8re \ue9tait port\ue9e \ue0 cette situation. L\u2019objectif de cette \ue9tude \ue9tait d\u2019\ue9valuer les qualit\ue9s agronomiques des pyramides g\ue9n\ue9tiques avanc\ue9es d\ue9velopp\ue9es d\u2019un croisement de quatre parents pour la r\ue9sistance aux maladies multiples. Les g\ue8nes pyramid\ue9s de r\ue9sistance (R) \ue0 la maladie de quatre parents \ue9taient\ua0: Co42 et Co-5 de G2333; Phg-2 de MEX54; Pythium ultimum Dennis de MLB49-89A et I &amp; bc3 de MCM5001. Les lign\ue9es de prog\ue9nitures \ue9taient plant\ue9es en arrangement de block incomplet et r\ue9pliqu\ue9 trois fois pendant deux saisons (2015A et 2015B) dans les champs \ue0 CIAT, Kawanda en Ouganda. Les traits agronomiques \ue9taient hautement h\ue9ritables (0,6), \ue0 l\u2019exception du nombre de gousses par plant (&lt;0.3). Le r\ue9trocroisement a g\ue9n\ue9r\ue9 des lign\ue9es d\u2019haricot \ue0 haut rendement, avec 270-290 graines par plant et \ue0 maturit\ue9 pr\ue9coce (95-100 jours). Neuf lign\ue9es sup\ue9rieures avec des qualit\ue9s d\ue9sirables, telles que la pr\ue9cocit\ue9 (95 jours), taux de graines \ue9lev\ue9s (290 graines par plant) et l\u2019habilit\ue9 grimpante, \ue9taient obtenues. La pyramide des g\ue8nes R n\u2019avait pas affect\ue9 les traits li\ue9s au rendement, \ue0 l\u2019exception de la p\ue9riode de floraison et le nombre de bougeons de fleurs par plant d\ufb \ue0 la s\ue9gr\ue9gation transgressive

Protecting Endangered Species: Do the Main Legislative Tools Work?

It is critical to assess the effectiveness of the tools used to protect endangered species. The main tools enabled under the U.S. Endangered Species Act (ESA) to promote species recovery are funding, recovery plan development and critical habitat designation. Earlier studies sometimes found that statistically significant effects of these tools could be detected, but they have not answered the question of whether the effects were large enough to be biologically meaningful. Here, we ask: how much does the recovery status of ESA-listed species improve with the application of these tools? We used species' staus reports to Congress from 1988 to 2006 to quantify two measures of recovery for 1179 species. We related these to the amount of federal funding, years with a recovery plan, years with critical habitat designation, the amount of peer-reviewed scientific information, and time listed. We found that change in recovery status of listed species was, at best, only very weakly related to any of these tools. Recovery was positively related to the number of years listed, years with a recovery plan, and funding, however, these tools combined explain <13% of the variation in recovery status among species. Earlier studies that reported significant effects of these tools did not focus on effect sizes; however, they are in fact similarly small. One must conclude either that these tools are not very effective in promoting species' recovery, or (as we suspect) that species recovery data are so poor that it is impossible to tell whether the tools are effective or not. It is critically important to assess the effectiveness of tools used to promote species recovery; it is therefore also critically important to obtain population status data that are adequate to that task

Effectiveness of vasectomy using cautery

BACKGROUND: Little evidence supports the use of any one vas occlusion method. Data from a number of studies now suggest that there are differences in effectiveness among different occlusion methods. The main objectives of this study were to estimate the effectiveness of vasectomy by cautery and to describe the trends in sperm counts after cautery vasectomy. Other objectives were to estimate time and number of ejaculations to success and to determine the predictive value of success at 12 weeks for final status at 24 weeks. METHODS: A prospective, non-comparative observational study was conducted between November 2001 and June 2002 at 4 centers in Brazil, Canada, the UK, and the US. Four hundred men who chose vasectomy were enrolled and followed for 6 months. Sites used their usual cautery vasectomy technique. Earlier and more frequent than normal semen analyses (2, 5, 8, 12, 16, 20, and 24 weeks after vasectomy) were performed. Planned outcomes included effectiveness (early failure based on semen analysis), trends in sperm counts, time and number of ejaculations to success, predictive value of success at 12 weeks for the outcome at 24 weeks, and safety evaluation. RESULTS: A total of 364 (91%) participants completed follow-up. The overall failure rate based on semen analysis was 0.8% (95% confidence interval 0.2, 2.3). By 12 weeks 96.4% of participants showed azoospermia or severe oligozoospermia (< 100,000 sperm/mL). The predictive value of a single severely oligozoospermia sample at 12 weeks for vasectomy success at the end of the study was 99.7%. One serious unrelated adverse event and no pregnancies were reported. CONCLUSION: Cautery is a very effective method for occluding the vas. Failure based on semen analysis is rare. In settings where semen analysis is not practical, using 12 weeks as a guideline for when men can rely on their vasectomy should lessen the risk of failure compared to using a guideline of 20 ejaculations after vasectomy