Arabidopsis glutathione reductase 2 is indispensable in plastids, while mitochondrial glutathione is safeguarded by additional reduction and transport systems.

A highly negative glutathione redox potential (EGSH ) is maintained in the cytosol, plastids and mitochondria of plant cells to support fundamental processes, including antioxidant defence, redox regulation and iron-sulfur cluster biogenesis. Out of two glutathione reductase (GR) proteins in Arabidopsis, GR2 is predicted to be dual-targeted to plastids and mitochondria, but its differential roles in these organelles remain unclear. We dissected the role of GR2 in organelle glutathione redox homeostasis and plant development using a combination of genetic complementation and stacked mutants, biochemical activity studies, immunogold labelling and in vivo biosensing. Our data demonstrate that GR2 is dual-targeted to plastids and mitochondria, but embryo lethality of gr2 null mutants is caused specifically in plastids. Whereas lack of mitochondrial GR2 leads to a partially oxidised glutathione pool in the matrix, the ATP-binding cassette (ABC) transporter ATM3 and the mitochondrial thioredoxin system provide functional backup and maintain plant viability. We identify GR2 as essential in the plastid stroma, where it counters GSSG accumulation and developmental arrest. By contrast a functional triad of GR2, ATM3 and the thioredoxin system in the mitochondria provides resilience to excessive glutathione oxidation.

Active optics system of the VLT Survey Telescope.

This paper describes the active optics system of the VLT Survey Telescope, the 2.6-m survey telescope designed for visible wavelengths of the European Southern Observatory at Cerro Paranal, in the Atacama desert. The telescope is characterized by a wide field of view (1.42 deg diameter), leading to tighter active optics than in conventional telescopes, in particular for the alignment requirements. We discuss the effects of typical error sources on the image quality and present the specific solutions adopted for wavefront sensing and correction of the aberrations, which are based on the shaping of a monolithic primary mirror and the positioning of the secondary in five degrees of freedom.

Attitudes of family physicians towards adolescent cannabis users: a qualitative study in France.

BACKGROUND: GPs are the health professionals most frequently consulted by adolescents. However, discussion between GPs and adolescents regarding cannabis use does not occur spontaneously. OBJECTIVE: To identify obstacles to the identification and management of cannabis use by adolescents based on GPs' experiences. METHODS: We conducted a qualitative study using focus groups of GPs from the Auvergne area (France). The GPs were selected according to descriptive and strategic variables. Three researchers--an anthropologist, a psychiatrist with expertise in addiction and a GP--performed a thematic analysis. RESULTS: Twenty-four GPs participated in three consecutive focus groups. The GPs were aware of the health risks of cannabis, yet ambivalent about its use by adolescents. The GPs also reported a lack of patient questioning during consultation. The obstacles to the identification and treatment of cannabis use by adolescents identified included lack of GP knowledge about cannabis (e.g. consumption patterns and laws); difficulties in addressing the issue with adolescents, evaluating adolescents' consumption and its impact and proposing support and follow-up and the presence of parents. The GPs were aware that their role lies at the intersection between the medical, personal, familial and social fields. CONCLUSION: Despite these barriers, GPs are willing to ask adolescents about their cannabis use. An adolescent's awareness, environment and receptiveness favour a sustainable therapeutic relationship. Brief intervention is a tool that may be of assistance in this relationship and allow GPs to take the initiative.

CANABIC: CANnabis and Adolescents: effect of a Brief Intervention on their Consumption--study protocol for a randomized controlled trial.

BACKGROUND: Cannabis is the most consumed illegal substance in France. General practitioners (GPs) are the health professionals who are most consulted by adolescents. Brief intervention (BI) is a promising care initiative for the consumption of cannabis, and could be a tool for GPs in caring for adolescents who consume cannabis. The aim of the CANABIC study is to measure the impact of a BI carried out by a GP on the consumption of cannabis by adolescents of 15 to 25 years of age. METHODS: A randomized clustered controlled trial, stratified over three areas (Auvergne, Languedoc-Roussillon, and Rhône - Alpes), comparing an intervention group, which carries out the BI in consultation, and a control group, which ensures routine medical care. The main assessment criterion is the consumption of cannabis by amount of joints per month, at 12 months. The amount necessary to highlight a significant difference between the two groups of 30% of consumption at 12 months is 250 patients (50 GPs, 5 patients per GP; risk α = 5%; power = 90%; intra-cluster correlation coefficient ρ = 0.2; Hawthorne effect = 15%; lost to follow-up rates for GPs = 10% and for patients = 20%). This plan is replicated for the three areas, and therefore a total of 750 patients are expected.The secondary criteria for judgment are the associated consumption of tobacco and alcohol, the perception of the consequences of consumption, and the driving of a vehicle following consumption. DISCUSSION: Research about BI for young cannabis users is underway. The aim of the CANABIC study is to validate a BI suited to adolescents who consume cannabis, which may be performed in the general practice. This would provide a tool for their treatment by a GP, which could be widely distributed during initial or further medical training. TRIAL REGISTRATION: CANABIC is a randomized clustered trial (NCT01433692, registered 2011 Sept 12), PHRC funded: Clinical Research Hospital Program (Governmental Fund, Health Ministry). Date first patient randomized: March 2012.

[The limits of patient education in short-term hospitalization: the case of cardiovascular patients]. / Éducation thérapeutique en court séjour : quelles limites ? Cas de patients cardiovasculaires.

The purpose of this study was to examine the limitations of therapeutic education for patients with cardiovascular risk factors during short hospital stays. The paper presents the results of a qualitative study conducted over the course of a year involving 18 case studies of professional practices and 18 interviews with 5 health professionals and 13 patients. The results show that professionals and patients have conflicting views about the time spent in hospital, as well as conflicting concerns and expectations, thus limiting the effectiveness of educational care. The findings suggest that after acute myocardial infarction or a stroke, patients tend to view themselves as survivors during their experience of short-term hospitalization in a care unit. As a result, short-term hospitalization may not be conducive to the mobilization of patients' cognitive and emotional capacities in a therapeutic education program.

Removing static aberrations from the active optics system of a wide-field telescope.

The wavefront sensor in active and adaptive telescopes is usually not in the optical path toward the scientific detector. It may generate additional wavefront aberrations, which have to be separated from the errors due to the telescope optics. The aberrations that are not rotationally symmetric can be disentangled from the telescope aberrations by a series of measurements taken in the center of the field, with the wavefront sensor at different orientation angles with respect to the focal plane. This method has been applied at the VLT Survey Telescope on the ESO Paranal observatory.

Health care support issues for internationally adopted children: a qualitative approach to the needs and expectations of families.

BACKGROUND: Families of internationally adopted children may face specific problems with which general practitioners (GPs) may not be familiar. The aim of the study was to explore problems faced by families before, during and after the arrival of their internationally adopted child and to assess the usefulness of a specific medical structure for internationally adopted children, which could be a resource for the GP. METHODOLOGY/PRINCIPAL FINDINGS: We conducted a qualitative study using individual semistructured guided conversations and interviewed 21 families that had adopted a total of 26 children internationally in the Puy de Dome department, France, in 2003. Quantitative data were used to describe the pathologies diagnosed and the investigations performed.Our study showed that the history of these families, from the start of the adoption project to its achievement, is complex and warrants careful analysis. Health-care providers should not only consider the medical aspects of adoption, but should also be interested in the histories of these families, which may play a role in the forming of attachments between the adoptee and their adoptive parents and prevent further trouble during the development of the child. We also showed that adoptive parents have similar fears or transient difficulties that may be resolved quickly by listening and reassurance. Most such families would support the existence of a specific medical structure for internationally adopted children, which could be a resource for the general practitioner. However, the health-care providers interviewed were divided on the subject and expressed their fear that a special consultation could be stigmatizing to children and families. CONCLUSIONS/SIGNIFICANCE: A specific consultation with well-trained and experienced practitioners acting in close collaboration with GPs and paediatricians may be of help in better understanding and supporting adopted children and their families.

Glutathione deficiency of the Arabidopsis mutant pad2-1 affects oxidative stress-related events, defense gene expression, and the hypersensitive response.

The Arabidopsis (Arabidopsis thaliana) phytoalexin-deficient mutant pad2-1 displays enhanced susceptibility to a broad range of pathogens and herbivorous insects that correlates with deficiencies in the production of camalexin, indole glucosinolates, and salicylic acid (SA). The pad2-1 mutation is localized in the GLUTAMATE-CYSTEINE LIGASE (GCL) gene encoding the first enzyme of glutathione biosynthesis. While pad2-1 glutathione deficiency is not caused by a decrease in GCL transcripts, analysis of GCL protein level revealed that pad2-1 plants contained only 48% of the wild-type protein amount. In contrast to the wild type, the oxidized form of GCL was dominant in pad2-1, suggesting a distinct redox environment. This finding was corroborated by the expression of GRX1-roGFP2, showing that the cytosolic glutathione redox potential was significantly less negative in pad2-1. Analysis of oxidative stress-related gene expression showed a higher transcript accumulation in pad2-1 of GLUTATHIONE REDUCTASE, GLUTATHIONE-S-TRANSFERASE, and RESPIRATORY BURST OXIDASE HOMOLOG D in response to the oomycete Phytophthora brassicae. Interestingly, oligogalacturonide elicitation in pad2-1 revealed a lower plasma membrane depolarization that was found to act upstream of an impaired hydrogen peroxide production. This impaired hydrogen peroxide production was also observed during pathogen infection and correlated with a reduced hypersensitive response in pad2-1. In addition, a lack of pathogen-triggered expression of the ISOCHORISMATE SYNTHASE1 gene, coding for the SA-biosynthetic enzyme isochorismate synthase, was identified as the cause of the SA deficiency in pad2-1. Together, our results indicate that the pad2-1 mutation is related to a decrease in GCL protein and that the resulting glutathione deficiency negatively affects important processes of disease resistance.

Active optics control of VST telescope secondary mirror.

In telescopes based on active optics, defocus and coma are usually compensated for by secondary mirror movements. They are performed at the Very Large Telescope Survey Telescope (VST) with a hexapod--a parallel robot with six degrees of freedom positioning capability. We describe the application of the two-mirror telescope theory to the VST case and the solutions adopted for the hexapod control. We present the results of performance and reliability tests performed both in the laboratory and at the telescope.

Active optics primary mirror support system for the 2.6 m VST telescope.

The Very Large Telescope Survey Telescope (VST) is equipped with an active optics system in order to correct low-order aberrations. The 2.6 m primary mirror is supported both axially and laterally and is surrounded by several safety devices for earthquake protection. We describe the mirror support system and discuss the results of the qualification test campaign.

The NADPH-dependent thioredoxin system constitutes a functional backup for cytosolic glutathione reductase in Arabidopsis.

Tight control of cellular redox homeostasis is essential for protection against oxidative damage and for maintenance of normal metabolism as well as redox signaling events. Under oxidative stress conditions, the tripeptide glutathione can switch from its reduced form (GSH) to oxidized glutathione disulfide (GSSG), and thus, forms an important cellular redox buffer. GSSG is normally reduced to GSH by 2 glutathione reductase (GR) isoforms encoded in the Arabidopsis genome, cytosolic GR1 and GR2 dual-targeted to chloroplasts and mitochondria. Measurements of total GR activity in leaf extracts of wild-type and 2 gr1 deletion mutants revealed that approximately 65% of the total GR activity is attributed to GR1, whereas approximately 35% is contributed by GR2. Despite the lack of a large share in total GR activity, gr1 mutants do not show any informative phenotype, even under stress conditions, and thus, the physiological impact of GR1 remains obscure. To elucidate its role in plants, glutathione-specific redox-sensitive GFP was used to dynamically measure the glutathione redox potential (E(GSH)) in the cytosol. Using this tool, it is shown that E(GSH) in gr1 mutants is significantly shifted toward more oxidizing conditions. Surprisingly, dynamic reduction of GSSG formed during induced oxidative stress in gr1 mutants is still possible, although significantly delayed compared with wild-type plants. We infer that there is functional redundancy in this critical pathway. Integrated biochemical and genetic assays identify the NADPH-dependent thioredoxin system as a backup system for GR1. Deletion of both, NADPH-dependent thioredoxin reductase A and GR1, prevents survival due to a pollen lethal phenotype.

Real-time imaging of the intracellular glutathione redox potential.

Dynamic analysis of redox-based processes in living cells is now restricted by the lack of appropriate redox biosensors. Conventional redox-sensitive GFPs (roGFPs) are limited by undefined specificity and slow response to changes in redox potential. In this study we demonstrate that the fusion of human glutaredoxin-1 (Grx1) to roGFP2 facilitates specific real-time equilibration between the sensor protein and the glutathione redox couple. The Grx1-roGFP2 fusion protein allowed dynamic live imaging of the glutathione redox potential (E(GSH)) in different cellular compartments with high sensitivity and temporal resolution. The biosensor detected nanomolar changes in oxidized glutathione (GSSG) against a backdrop of millimolar reduced glutathione (GSH) on a scale of seconds to minutes. It facilitated the observation of redox changes associated with growth factor availability, cell density, mitochondrial depolarization, respiratory burst activity and immune receptor stimulation.

Redox-sensitive GFP in Arabidopsis thaliana is a quantitative biosensor for the redox potential of the cellular glutathione redox buffer.

The cellular glutathione redox buffer is assumed to be part of signal transduction pathways transmitting environmental signals during biotic and abiotic stress, and thus is essential for regulation of metabolism and development. Ratiometric redox-sensitive GFP (roGFP) expressed in Arabidopsis thaliana reversibly responds to redox changes induced by incubation with H(2)O(2) or DTT. Kinetic analysis of these redox changes, combined with detailed characterization of roGFP2 in vitro, shows that roGFP2 expressed in the cytosol senses the redox potential of the cellular glutathione buffer via glutaredoxin (GRX) as a mediator of reversible electron flow between glutathione and roGFP2. The sensitivity of roGFP2 toward the glutathione redox potential was tested in vivo through manipulating the glutathione (GSH) content of wild-type plants, through expression of roGFP2 in the cytosol of low-GSH mutants and the endoplasmic reticulum (ER) of wild-type plants, as well as through wounding as an example for stress-induced redox changes. Provided the GSH concentration is known, roGFP2 facilitates the determination of the degree of oxidation of the GSH solution. Assuming sufficient glutathione reductase activity and non-limiting NADPH supply, the observed almost full reduction of roGFP2 in vivo suggests that a 2.5 mm cytosolic glutathione buffer would contain only 25 nm oxidized glutathione disulfide (GSSG). The high sensitivity of roGFP2 toward GSSG via GRX enables the use of roGFP2 for monitoring stress-induced redox changes in vivo in real time. The results with roGFP2 as an artificial GRX target further suggest that redox-triggered changes of biologic processes might be linked directly to the glutathione redox potential via GRX as the mediator.