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Promoting the implementation of practices that are supported by research: the National Implementing Evidence-Based Practice Project.

The National Implementing Evidence-Based Practice Project is an ongoing effort to promote the implementation of effective practices for adults who have severe mental illnesses. The project members designed and developed integrated packages of materials and services to help practice sites implement evidence-based practices and is field-testing the approach in eight states. These implementations are being evaluated carefully to learn how to make the technology transfer process more efficient in the future. This article describes the project and provides some early reflections on the implementation experience.

Adolescent↗

Detection of fungal DNA by real-time polymerase chain reaction: evaluation of 2 methodologies in experimental pulmonary aspergillosis.

The capabilities of 2 quantitative polymerase chain reaction (PCR) assays for detecting pulmonary aspergillosis were analyzed. Both methodologies were real-time (RT) based and were compared with quantitative cultures and galactomannan (GM) antigen detection in a rabbit model of invasive aspergillosis. A total of 106 samples including blood, serum, lung, and brain from 3 controls and 9 infected New Zealand rabbits were analyzed. The RT-PCR methodologies were an Aspergillus fumigatus-specific assay using fluorescent resonance energy transfer technology targeting a highly conserved region of the fungal 18S rRNA gene and a panfungal assay to amplify the internal transcribed spacer regions 1 and 2 from fungal rRNA gene complex, employing SYBRGreen fluorescent dye as a detector. The specificity for both PCR base assays, culture, and GM determination was 100%. The sensitivity of the specific PCR assay was 88.9% in lung samples, 66.6% in serum, 55.5% in blood, and 33.3 in brain specimens. The panfungal assay had a sensitivity of 33.3% in lung and serum samples, being brain and blood specimens invariably negative. Otherwise, 100% of the lungs resulted positive for culture, and all serum samples showed a GM index above 1.0 after 2 days of infection. The specific RT-PCR assay is a reliable technique to detect A. fumigatus DNA in vivo comparable to cultures and GM determination. The panfungal RT-PCR assay exhibited low sensitivity to diagnose invasive aspergillosis in rabbits advising against its clinical introduction.

Animals↗

An evaluation of workshop training in motivational interviewing for addiction and mental health clinicians.

We evaluated a 2-day training workshop on motivational interviewing (MI) for addiction and mental health clinicians (n = 22). Clinicians completed the helpful responses questionnaire (HRQ) and taped interactions with a standardized patient (SP). Independent, blinded coders rated the tapes using the motivational interviewing skills code (MISC). Post-training assessment showed significant increase on the HRQ and two of four MISC summary scores for SP interviews. At 2-month follow-up, means of the HRQ and two MISC summary scores remained higher than baseline, but declined from post-training. Some MI skills improved to a greater degree than others, and a subset of clinicians (> 40%) showed continued improvement at follow-up. Results were consistent across interviews with different SPs, supporting the use of this assessment method. Findings highlight the need for additional research on the use of standardized patients, alternative assessment techniques, and effective methods of technology transfer for MI skill acquisition and retention.

Adult↗

Photocatalytic degradation of methyl red by TiO2: comparison of the efficiency of immobilized nanoparticles versus conventional suspended catalyst.

The photocatalytic efficiency of supported TiO2 nanoparticles (mean size 6 nm), immobilized onto the inner walls of a cylindrical glass photoreactor was compared versus the performance of conventional TiO2 Degussa P25 catalyst. For this purpose the degradation of methyl red dye was used as evaluation test. The obtained results showed that the TiO2 Degussa P25 catalyst is more efficient than the supported nanoparticles. The poorer performance of the nanosized catalyst can be ascribed to the fact that the immobilization procedure turns out, in spite of the extremely high surface to volume ratio, in an overall reduction of active surface area available for target molecule adsorption, due to the low porosity of the supported catalyst layer. The kinetics of the investigated processes were monitored and a study on the reaction products and intermediates was carried out in order to evaluate possible difference in the reaction pathway in presence of immobilized nanoparticles versus suspended catalyst. The results demonstrate that the mechanisms of parent dye degradation in presence of supported TiO2 nanoparticles are the same as those occurring in presence of TiO2 Degussa P25 catalyst. The present work describe the results obtained on the feasibility of scaling up the colloidal nanocrystal-based photocatalysis experiment: the comparison with a well standardized degradation method performed with a known material can allow a realistic evaluation of the advantages and the limits of the investigated nanoparticle towards the ultimate technology transfer.

Azo Compounds↗

Modeling behavioral and neuronal symptoms of Alzheimer's disease in mice: a role for intraneuronal amyloid.

The amyloid Abeta-peptide (Abeta) is suspected to play a critical role in the cascade leading to AD as the pathogen that causes neuronal and synaptic dysfunction and, eventually, cell death. Therefore, it has been the subject of a huge number of clinical and basic research studies on this disease. Abeta is typically found aggregated in extracellular amyloid plaques that occur in specific brain regions enriched in nAChRs in Alzheimer's disease (AD) and Down syndrome (DS) brains. Advances in the genetics of its familiar and sporadic forms, together with those in gene transfer technology, have provided valuable animal models that complement the traditional cholinergic approaches, although modeling the neuronal and behavioral deficits of AD in these models has been challenging. More recently, emerging evidence indicates that intraneuronal accumulation of Abeta may also contribute to the cascade of neurodegenerative events and strongly suggest that it is an early, pathological biomarker for the onset of AD and associated cognitive and other behavioral deficits. The present review covers these studies in humans, in in vitro and in transgenic models, also providing more evidence that adult 3xTg-AD mice harboring PS1M146V, APPSwe, tauP301L transgenes, and mimicking many critical hallmarks of AD, show cognitive deficits and other behavioral alterations at ages when overt neuropathology is not yet observed, but when intraneuronal Abeta, synaptic and cholinergic deficits can already be described.

Alzheimer Disease↗

Ectopic expression of doublecortin protects adult rat progenitor cells and human glioma cells from severe oxygen and glucose deprivation.

Doublecortin (DCX) is a microtubule-associated protein expressed in migrating neuroblasts. DCX expression is increased in subventricular zone (SVZ) cells migrating to the boundary of an ischemic lesion after induction of middle cerebral artery occlusion (MCAO) in adult rats and mice. We tested the hypothesis that DCX, in addition to being a marker of migrating neuroblasts, serves to protect neuroblasts from conditions of stress, such as oxygen and glucose deprivation (OGD). Using gene transfer technology, we overexpressed DCX in rat SVZ and U-87 human glioma cells. The cells remained viable against severe OGD, up to 32 h exhibiting 1% apoptosis compared with 100% apoptosis in control. In addition, these genetically modified cells upregulated expression of E-, VE- and N-cadherin, molecules that promote endothelial survival signals via the VE-cadherin/vascular endothelial growth factor receptor-2/phosphoinositide 3-kinase (PI3-K)/AKT/beta-catenin pathway and inactivate the proapoptotic factor Bad. DCX overexpression also significantly increased cell migration in SVZ tissue explants and U-87 cells and significantly upregulated microtubule-associated protein-2 (MAP2) and nestin protein levels in SVZ and U-87 cells compared with wild-type control cells. Knocking down DCX expression in DCX overexpressing SVZ and U-87 cells with DCX small interfering RNA (siRNA), confirmed the specificity of DCX on cell survival against OGD, and the DCX induced upregulation of E-, VE- and N-cadherin, MAP2 and nestin. In NIH3T3 cells, DCX overexpression had no effect on cell survival against OGD, and indicating that the protective effects of DCX was restricted to brain cells e.g. SVZ and U-87 cells. Our data suggest a novel and an important role for DCX as a protective agent for migrating neuroblasts and tumor cells.

Analysis of Variance↗

Purification and scale-up of a recombinant heavy chain fragment C of botulinum neurotoxin serotype E in Pichia pastoris GS115.

A recombinant C-terminus heavy chain fragment from botulinum neurotoxin serotype E (BoNT/E) is proposed as a vaccine against the serotype E neurotoxin. This fragment, rBoNTE(Hc), was produced intracellular in Pichia pastoris GS115 by a three-step fermentation process, i.e., glycerol batch phase and a glycerol fed-batch phase to achieve high cell densities, followed by a methanol fed-batch induction phase. The rBoNTE(Hc) protein was purified from the soluble fraction of cell lysates using three ion-exchange chromatography steps (SP Sepharose Fast Flow, Q Sepharose Fast Flow, Sp Sepharose High Performance) and polished with a hydrophobic charge induction chromatography step (MEP HyperCel). Method development at the bench scale was achieved using 7-380 mL columns and the process was performed at the pilot scale using 0.5-3.1 L columns in preparation for technology transfer to cGMP manufacturing. The purification process resulted in greater than 98% pure rBoNTE(Hc) based on HPLC and yielded up to 1.01g of rBoNTE(Hc)/kg cells at the bench scale and 580mg vaccine/kg cells at the pilot scale. N-terminal sequencing showed that the purified rBoNTE(Hc) N-terminus is intact and was found to protect mice against a challenge of 1000 mouse intraperitoneal LD50's of BoNT/E.

Amino Acid Sequence↗

Patenting for the research scientist: an update.

Academic institutional research constantly produces results worthy of patent protection, but coping with the demands of patent law presents considerable challenges to bioscientists working in these institutions. Inventors need, however, to be aware of recent patent office guidelines and court decisions if they are to seek useful intellectual property as a basis for technology transfer to industry.

Animals↗

Translation of an experimental oral vaccine formulation into a commercial product.

An effective experimental vaccine may fail to become a therapeutic reality for a number of scientific, regulatory or commercial reasons. In this review, we share some of our personal experiences as University-based researchers and provide an account of some of the problems that we have encountered during preliminary scale-up and assessment of an oral influenza vaccine formulation. Many of the problems we have faced have been non-scientific and related to identifying project-funding sources, finding suitable contract manufacturing companies that are GMP compliant, and protecting intellectual property generated from the scientific studies. The review is intended as a practical guide that will allow other researchers to adopt effective strategies to permit the translation of an effective experimental formulation to a viable commercial product.

Administration, Oral↗

Gene therapy for inborn errors of liver metabolism.

In humans, treatment for inborn errors of liver metabolism has focused on dietary, drug, and cell therapies. However, significant morbidity and mortality still remain and alternative and/or adjunctive strategies are needed. It has been 15 years since human gene therapy trials were initiated for genetic diseases. While significant progress has been made in the preclinical arena in a variety of disease models, sustained effect in humans has still eluded the field of inborn errors of liver metabolism. At this time, achievement of clinical efficacy in different animal models has been reported with multiple gene transfer technologies. Current efforts are aimed at fully understanding host-vector interactions, and how manipulation of these interactions can help us to increase the therapeutic index of any specific therapy. As with any therapy, it will be the balance of this index and the disease natural history which will determine the future of clinical studies and their outcomes.

DNA↗

Genetic modification of T lymphocytes for adoptive immunotherapy.

Adoptive transfer of T lymphocytes is a promising therapy for malignancies-particularly of the hemopoietic system-and for otherwise intractable viral diseases. Efforts to broaden the approach have been limited by the physiology of the T cells themselves and by a range of immune evasion mechanisms developed by tumor cells. In this review we show how genetic modification of T cells is being used preclinically and in patients to overcome these limitations, by incorporation of novel receptors, resistance mechanisms, and control genes. We also discuss how the increasing safety and effectiveness of gene transfer technologies will lead to an increase in the use of gene-modified T cells for the treatment of a wider range of disorders.

Genetic Vectors↗

Nematode parasite control of livestock in the tropics/subtropics: the need for novel approaches.

Because parasites are more abundant, small ruminants in the tropical/subtropical regions of the world experience much greater ravages from internal parasitic disease than those in the temperate regions. In the tropics/subtropics, the limiting ecological factor influencing the severity of parasitism is rainfall, as temperatures almost always favour hatching and development of the free-living stages. Attempts to expand sheep and goat production by replacing traditional village production systems, which rarely involve anthelmintic treatment, with large-scale intensive commercial enterprises invariably induce complete reliance on anthelmintics to control nematode parasites. This has led to the widespread development of high level, multiple anthelmintic resistance throughout the tropics/subtropics, and in certain regions this has reached the ultimate disastrous scenario of total chemotherapeutic failure. Immediate concerted efforts are needed to resolve this crisis. Significant benefits are likely to emerge from research into non-chemotherapeutic approaches to nematode parasite control, such as grazing management, worm vaccines, breed selection and biological control. However, it is likely that none, in isolation or collectively, will completely replace the need for effective anthelmintics. What is needed is the integration of all methods of parasite control as they come to hand, with the underlying aim of reducing the use and thus preserving the effectiveness of anthelmintics. Although cheap and simple procedures, based on sound epidemiological principles, can achieve dramatic benefits in worm control, they have been poorly adopted by livestock owners. Clearly then, the greatest need is for technology transfer and education programmes, but these activities are generally found to be chronically under-resourced.

Animal Husbandry↗

Pancreatic cancer growth is inhibited by blockade of VEGF-RII.

BACKGROUND: Angiogenesis is important in the development and progression of pancreatic cancer. Therefore antiangiogenic therapy targeting endothelial cells may represent a promising therapeutic option. The aim of the study was to evaluate antiangiogenic therapy as a potential therapeutic option in pancreatic cancer. METHODS: Replication-deficient retroviruses encoding truncated VEGF-RII were used to block vascular endothelial growth factor (VEGF) signaling. Tumor growth of 3 pancreatic cancer cell lines was assayed in a nude mouse model in which each pancreatic cancer cell line was subcutaneously inoculated together with retrovirus-producing cells. Expression of VEGF was assayed by RT-PCR and by enzyme-linked immunosorbent assay. Oxygen tension in tumors was determined polarographically. RESULTS: All 3 pancreatic cancer cell lines expressed VEGF mRNA, with the highest VEGF secretion seen in MIA PaCa-2 cells. In vivo therapeutic intervention through dominant negative inhibition of VEGF-RII significantly reduced the growth rate of subcutaneous tumors and inhibited tumor neoangiogenesis. Tumor oxygenation, however, was not altered in xenograft tumors treated with dominant negative retroviruses. CONCLUSION: The ligand/receptor system consisting of VEGF and VEGF-RII seems to be of biologic significance in the pathogenesis of pancreatic cancer growth. Therefore therapeutic intervention in this angiogenic system by a retroviral-based gene transfer technology represents a rational and feasible new technique to inhibit tumor growth.

Animals↗

Gene therapy and the aging nervous system.

In recent years, the first attempts have been made to apply gene transfer technology to protect neurons from death following neurological insults. There has been sufficient progress in this area that it becomes plausible to consider similar gene therapy approaches meant to delay aspects of aging of the nervous system. In this review, we briefly consider such progress and how it might be applied to the realm of the aging brain. Specifically, we consider: (a) the means of delivery of such therapeutic genes; (b) the choice of such genes; and (c) technical elaborations in gene delivery systems which can more tightly regulate the magnitude and duration of transgene protection.

Aging↗

Origins of mammalian hematopoiesis: in vivo paradigms and in vitro models.

Though a topic of medical interest for centuries, our understanding of vertebrate hematopoietic or "blood-forming" tissue development has improved greatly only in recent years and given a series of scientific and technical milestones. Key among these observations was the description of procedures that allowed the transplantation of blood-forming activity. Beyond this, other advances include the creation of a variety of knock-out animals (mice and more recently zebrafish), microdissection of embryonic and fetal blood-forming tissues, hematopoietic stem (HSC) and progenitor cell (HPC) colony-forming assays, the discovery of cytokines with defined hematopoietic activities, gene transfer technologies, and the description of lineage-specific surface antigens for the identification and purification of pluripotent and differentiated blood cells. The availability of both murine and human embryonic stem cells (ESC) and the delineation of in vitro systems to direct their differentiation have now been added to this analytical arsenal. Such tools have allowed researchers to interrogate the complex developmental processes behind both primitive (yolk sac or extraembryonic) and definitive (intraembryonic) hematopoietic tissue formation. Using ES cells, we hope to not only gain additional basic insights into hematopoietic development but also to develop platforms for therapeutic use in patients suffering from hematological disease. In this review, we will focus on points of convergence and divergence between murine and human hematopoiesis in vivo and in vitro, and use these observations to evaluate the literature regarding attempts to create hematopoietic tissue from embryonic stem cells, the pitfalls encountered therein, and what challenges remain.

Animals↗

Comparison of two manipulation methods to produce in vitro fertilized, biopsied and vitrified bovine embryos.

The objective of this study was to compare the overall efficiency, measured by in vitro embryonic survival, and practical value of bovine in vitro embryo production, biopsy, vitrification, and direct transfer technology using 2 different manipulation methods for biopsy. Slaughterhouse-derived oocytes were matured in vitro, fertilized (Day 0) with frozen-thawed, Percoll-separated spermatozoa and cultured on a granulosa cell monolayer. In Experiment 1, one or two blastomeres were expelled from Day 4 embryos by mechanical force through a hole made by partial zona dissection. Using a darning needle hole system for individual culture of biopsied embryos from Day 4 to Day 7.5, the blastocyst per oocyte rate was 50%, and 76% of the blastocysts survived subsequent vitrification and direct in-straw rehydration. Attempts to increase the cell number of the biopsies by further in vitro culture were unsuccessful. In Experiment 2, Day 7 and Day 8 embryos were manually biopsied before or after vitrification. When biopsy was performed before vitrification, 98% of the embryos survived manipulation, and 86% of these re-expanded after vitrification and in-straw dilution. Biopsy after vitrification was less efficient, since only 69% of the embryos survived both processes. The cumulative efficiency of embryo production, Day 7.5 biopsy and vitrification--in-straw direct rehydration was lower (P < 0.001) than that of Day 4 biopsy and Day 7.5 vitrification (29 vs 38%, respectively). However, a Day 7.5 biopsy may have the more practical application since the size of the biopsy is larger and the process is not as time-consuming as the long-term individual culture of the biopsied embryos.

Journal Article↗

Space product development: bringing the benefits of space down to Earth.

In fulfilling the National Aeronautics and Space Administration's (NASA) responsibility to encourage the fullest commercial use of space the Space Product Development (SPD) Program, within the Microgravity Research Program Office (MRPO) located at the Marshall Space Flight Center (MSFC) in Huntsville, Alabama, is managing an organization of Commercial Space Centers (CSC's) that have successfully employed methods for encouraging private industries to exploit the benefits of space-based research. Unique research opportunities of the space environment are being made available to private industry in an effort to develop new, competitive products; create jobs; and enhance the country's quality of life. Over 200 commercial research activities have been conducted in space by the CSC's and their industrial partners during the last several years. The success of this research is evidenced by the increasing amount of industrial participation in commercial microgravity research and the potential products nearing marketability.

Biotechnology↗