Enhanced green fluorescence by the expression of an Aequorea victoria green fluorescent protein mutant in mono- and dicotyledonous plant cells.

The expression of the jellyfish green fluorescent protein (GFP) in plants was analyzed by transient expression in protoplasts from Nicotiana tabacum, Arabidopsis thaliana, Hordeum vulgare, and Zea mays. Expression of GFP was only observed with a mutated cDNA, from which a recently described cryptic splice site had been removed. However, detectable levels of green fluorescence were only emitted from a small number of protoplasts. Therefore, other mutations in the GFP cDNA leading to single-amino acid exchanges in the chromophore region, which had been previously studied in Escherichia coli, were tested in order to improve the sensitivity of this marker protein. Of the mutations tested so far, the exchange of GFP amino acid tyrosine 66 to histidine (Y66H) led to detection of blue fluorescence in plant protoplasts, while the exchange of amino acid serine 65 to cysteine (S65C) and threonine (S65T) increased the intensity of green fluorescence drastically, thereby significantly raising the detection level for GFP. For GFP S65C, the detectable number of green fluorescing tobacco (BY-2) protoplasts was raised up to 19-fold, while the fluorimetricly determined fluorescence was raised by at least 2 orders of magnitude.     

A nationwide study of decisions to forego life-prolonging treatment in Dutch medical practice

BACKGROUND: Decisions to withhold or withdraw life-prolonging treatment in terminally ill patients are common in some areas of medical practice. Information about the frequency and background of these decisions is generally limited to specific clinical settings. This article describes the practice of withholding or withdrawing life-prolonging treatment in the Netherlands. METHODS: Questionnaires were sent to the attending physicians of a stratified sample of 6060 of all 43002 cases of death in the Netherlands from August 1 through November 30, 1995. The questions concerned the treatments foregone, the patient characteristics, and the decision-making process. The response rate was 77%. RESULTS: A nontreatment decision was made in 30% (95% confidence interval, 28%-31%) of all deaths in the Netherlands in 1995; this is an increase compared with 28% (95% confidence interval, 26%-29%) in 1990; in 20% of all deaths, this decision was the most important end-of-life decision. Artificial nutrition or hydration and antibiotics were the treatments most frequently foregone, each accounting for 25% of cases in which a nontreatment decision was made. Nursing-home physicians withheld or withdrew treatment more often than clinical specialists or general practitioners in 52%, 35%, and 17% of all deaths they were involved with, respectively. Of the patients in whom a nontreatment decision was the most important end-of-life decision, 26% were competent; of those, 93% were involved in the decision making. In 17% of patients, the nontreatment decision was made without being discussed with the patient or the patient's relatives and without knowledge of the patient's wishes. Life was shortened by an estimated 24 hours or less in 42% and 1 month or more in 8% of patients. CONCLUSIONS: Decisions to forego life-prolonging treatment are frequently made end-of-life decisions in the Netherlands and may be increasing. Most of these decisions do not involve high-technology treatments, and the consequences, in terms of shortening of life, are relatively small.     

A MUTANT OF Escherichia coli PERMITTING REPLICATION OF TWO F FACTORS

Control mechanisms of replication of bacterial genetic elements are poorly understood at present. We have studied one such mechanism involving replication of the F factor in Escherichia coli. The F factor can replicate either autonomously in F(+) or F' strains, or as an integral part of the chromosome in Hfr strains. We have shown that presence of either an integrated F factor or a free F factor prevents replication of a second free F factor. Two integrated F factors can replicate in the same cell.The present experiments show that when a F'lac element was transferred by mating into an Hfr strain, it had to become integrated into the chromosome in order to persist. With a recombination-deficient (recA) Hfr strain as recipient, the frequency of F'lac integration was greatly reduced. This permitted us to isolate a mutant Hfr strain in which the F'lac element was able to replicate autonomously. The mutation has most likely occurred in the integrated F factor itself. Availability of this new recA mutant Hfr strain facilitates genetic analysis of the F factor, since in recA(+) Hfr strains frequent integration of a free F factor into the chromosome obscures recognition of complementation and recombination between two free F factors.     

Cost-effectiveness of primary prevention of paediatric asthma: a decision-analytic model

Background

Many children stand to benefit from being asthma-free for life with primary (i.e., prenatally started) prevention addressing one environmental exposure in a unifaceted (UF) approach or at least two in a multifaceted (MF) approach. We assessed the cost-effectiveness of primary prevention programmes for Dutch children in a decision-analytic framework.

Methods

A decision-analytic tree model analysing healthcare costs and asthma cases prevented was developed to compare usual care (UC) with two UF and three MF programmes on the primary prevention of asthma amongst children. Programmes were evaluated through incremental cost-effectiveness ratios and net monetary benefits. Decision and parameter uncertainty were subjected to value-of-information analyses.

Results

The current UC and one of three MF programmes dominated the other alternatives. The MF programme was more costly but also more effective than UC at an incremental cost-effectiveness ratio of €8,209.20/additional asthma case prevented. The value of perfect information to reduce uncertainty was €291.6M at its lowest. Most of the uncertainty in the cost-effectiveness threshold was attributable to the probability and cost estimates for low-risk children.

Conclusion

This study supports the feasibility of a structured programme that simultaneously addresses exposure to house dust mites, pet dander, environmental tobacco, and breast-feeding as a cost-effective alternative to UC in the primary prevention of asthma amongst children.     

Evaluation of an imaging biomarker,Dixon quantitative chemical shift imaging,in Gaucher disease: lessons learned

Gaucher disease (GD) is the first lysosomal storage disorder for which specific therapy became available. The infiltration of bone marrow by storage cells plays an important part in the pathophysiology of skeletal complications and can be quantified by measurements of bone marrow fat fraction (Ff). Ff measurements by Dixon quantitative chemical shift imaging (QCSI) are standard for the follow-up care of GD patients at the Academic Medical Center. Several criteria should be met in order for these measurements to qualify as an imaging biomarker. These include: 1) The presence of the imaging biomarker is closely coupled or linked to the presence of the target disease or condition; 2) The detection and/or quantitative measurement of the biomarker is accurate, reproducible, and feasible over time, and; 3) The changes measured over time in the imaging biomarker are closely coupled, or linked, to the success or failure of the therapeutic effect and the true end point for the medical therapy being evaluated. This review assesses the use of Ff measurements by QCSI as a biomarker for GD in light of these criteria. In addition potential pitfalls are discussed including: degenerative disc disease; vertebral collapse and infection; haematological malignancies; focal fatty deposits; age; menopause; phase and repositioning errors, and; fat surrounding the basivertebral vein. QCSI measurements of Ff can be used as an imaging biomarker for GD taking these pitfalls into account. It is one of the first biomarkers, in particular imaging biomarkers, for GD that has been systematically evaluated and could be a valuable tool in clinical trials comparing different treatments or dosing regimens.     

Variations in radioiodine ablation: decision-making after total thyroidectomy

European Journal of Nuclear Medicine and Molecular Imaging - The role of radioiodine treatment following total thyroidectomy for differentiated thyroid cancer is changing. The last major revision...     

A technique for detecting GABA in the human brain with PRESS localization and optimized refocusing spectral editing radiofrequency pulses

A proton magnetic resonance spectral editing technique is presented that uses PRESS excitation to achieve spatially localized measurements of brain gamma-aminobutyric acid (GABA). The homonuclear difference spectroscopy technique employs a frequency selective inversion pulse to suppress the creatine resonance at 3.0 ppm. The timing of this pulse is optimized to maximize the suppression of creatine by minimizing the effect of the editing pulse on the 3.0 ppm resonances. The PRESS excitation achieves three dimensional spatial localization in a single acquisition making it less sensitive to patient motion than multiple acquisition techniques. The performance and utility of this technique were evaluated by phantom experiments and by in vivo measurements of brain GABA concentration in 10 normal subjects.     

The Role of the Cerebellum in Degenerative Ataxias and Essential Tremor: Insights From Noninvasive Modulation of Cerebellar Activity

Over the last three decades, measuring and modulating cerebellar activity and its connectivity with other brain regions has become an emerging research topic in clinical neuroscience. The most important connection is the cerebellothalamocortical pathway, which can be functionally interrogated using a paired-pulse transcranial magnetic stimulation paradigm. Cerebellar brain inhibition reflects the magnitude of suppression of motor cortex excitability after stimulating the contralateral cerebellar hemisphere and therefore represents a neurophysiological marker of the integrity of the efferent cerebellar tract. Observations that cerebellar noninvasive stimulation techniques enhanced performance of certain motor and cognitive tasks in healthy individuals have inspired attempts to modulate cerebellar activity and connectivity in patients with cerebellar diseases in order to achieve clinical benefit. We here comprehensively explore the therapeutic potential of these techniques in two movement disorders characterized by prominent cerebellar involvement, namely the degenerative ataxias and essential tremor. The article aims to illustrate the (patho)physiological insights obtained from these studies and how these translate into clinical practice, where possible by addressing the association with cerebellar brain inhibition. Finally, possible explanations for some discordant interstudy findings, shortcomings in our current understanding, and recommendations for future research will be provided. © 2019 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.     

Advantages of diagnosing bacterial spondylitis with magnetic resonance imaging

Summary We report on 16 cases of suspected spondylitis in which we used magnetic resonance (MR) imaging to confirm or exclude the diagnosis. MR has several advantages, one of which is to permit diagnosis of this disease in the early stages without major risks. In addition, MR permits recognition of complications such as paravertebral or intraspinal abscess formation with a high security and accuracy. Moreover, it is possible to show spondylitic alterations in three different planes. To differentiate this disease from metastatic or tumorous lesions the technique with T₁- and T₂-weighted images is helpful. As a result, MR imaging can shorten the time between onset and diagnosis of spondylitis.

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Zusammenfassung Es wird über 16 Patienten mit hämatogener bakterieller Spondylitis oder Verdacht auf einen entzündlichen Wirbelsäulenprozeß berichtet, bei denen die Magnetresonanztomographie (MR) diagnostisch angewandt wurde. Diese Methode erlaubt aufgrund ihrer technischen Bedingungen unabhängig von Röntgenstrahlen eine Spondylitis frühzeitig zu erfassen und kann entsprechende Veränderungen in drei verschiedenen Ebenen abbilden. Außerdem kann das Ausmaß eventueller begleitender paravertebraler und/oder intraspinaler Abszesse mit hoher Genauigkeit und Sicherheit dargestellt werden. Dies ist besonders für die operative Therapie vorteilhaft. Weiterhin ist aufgrund der Zuhilfenahme sog. T₁- und T₂-gewichteter Bilder die Differential-diagnostik zu anderen Erkrankungen der Wirbelsäule möglich. Auch können ausgeprägtere Veränderungen der kalkhaltigen Gewebsanteile dargestellt werden. Im Vergleich zu anderen diagnostischen Methoden besitzt die MR mehrere Vorteile in einer Methoden Dies dürfte einen wesentlichen Beitrag zur Frühdiagnose dieser Krankheit leisten.