Refining abacavir hypersensitivity diagnoses using a structured clinical assessment and genetic testing in the Swiss HIV Cohort Study.

BACKGROUND: We aimed to assess the value of a structured clinical assessment and genetic testing for refining the diagnosis of abacavir hypersensitivity reactions (ABC-HSRs) in a routine clinical setting. METHODS: We performed a diagnostic reassessment using a structured patient chart review in individuals who had stopped ABC because of suspected HSR. Two HIV physicians blinded to the human leukocyte antigen (HLA) typing results independently classified these individuals on a scale between 3 (ABC-HSR highly likely) and -3 (ABC-HSR highly unlikely). Scoring was based on symptoms, onset of symptoms and comedication use. Patients were classified as clinically likely (mean score > or =2), uncertain (mean score > or = -1 and < or = 1) and unlikely (mean score < or = -2). HLA typing was performed using sequence-based methods. RESULTS: From 131 reassessed individuals, 27 (21%) were classified as likely, 43 (33%) as unlikely and 61 (47%) as uncertain ABC-HSR. Of the 131 individuals with suspected ABC-HSR, 31% were HLA-B*5701-positive compared with 1% of 140 ABC-tolerant controls (P < 0.001). HLA-B*5701 carriage rate was higher in individuals with likely ABC-HSR compared with those with uncertain or unlikely ABC-HSR (78%, 30% and 5%, respectively, P < 0.001). Only six (7%) HLA-B*5701-negative individuals were classified as likely HSR after reassessment. CONCLUSIONS: HLA-B*5701 carriage is highly predictive of clinically diagnosed ABC-HSR. The high proportion of HLA-B*5701-negative individuals with minor symptoms among individuals with suspected HSR indicates overdiagnosis of ABC-HSR in the era preceding genetic screening. A structured clinical assessment and genetic testing could reduce the rate of inappropriate ABC discontinuation and identify individuals at high risk for ABC-HSR.

Effect of computerisation on the quality and safety of chemotherapy prescription.

BACKGROUND: Chemotherapy is prescribed according to protocols of several cycles. These protocols include not only therapeutic agents but also adjuvant solvents and inherent supportive care measures. Multiple errors can occur during the prescription, the transmission of documents and the drug delivery processes, and lead to potentially serious consequences. OBJECTIVE: To assess the effect of a computerised physician order entry (CPOE) system on the number of errors in prescription recorded by the centralised chemotherapy unit of a pharmacy service in a university hospital. PATIENTS AND METHODS: Existing chemotherapy protocols were standardised by a multidisciplinary team (composed of a doctor, a pharmacist and a nurse) and a CPOE system was developed from a File Maker Pro database. Chemotherapy protocols were progressively introduced into the CPOE system. The effect of the system on prescribing errors was measured over 15 months before and 21 months after starting computerised protocol prescription. Errors were classified as major (dosage and drug name) and minor (volume or type of infusion solution). RESULTS: Before computerisation, 141 errors were recorded for 940 prescribed chemotherapy regimens (15%). After introduction of the CPOE system, 75 errors were recorded for 1505 prescribed chemotherapy regimens (5%). Of these errors, 69 (92%) were recorded in prescriptions that did not use a computerised protocol. A dramatic decrease in the number of errors was noticeable when 50% of the chemotherapy protocols were prescribed through the CPOE system. CONCLUSION: Errors in chemotherapy prescription nearly disappeared after implementation of CPOE. The safety of chemotherapy prescription was markedly improved.

Critical limits of soil penetration resistance in a rhodic Eutrudox

Soil penetration resistance is an important indicator of soil physical quality and the critical limit of 2 MPa has been widely used to characterize the soil physical quality, in both no-tillage and conventional systems. The aim of this study was to quantify the influence of different tillage and cropping systems on the soil penetration resistance in a Rhodic Eutrudox. The experiment was carried out in a 5 × 2 factorial, completely randomized block design (tillage systems vs cropping systems), with four replications. The tillage systems consisted of: conventional tillage disk harrow; minimum tillage with annual chiseling; minimum tillage with chiseling every three years; no-tillage for 11 consecutive years; and no-tillage for 24 consecutive years. The factor cropping systems was represented by: crop rotation and crop succession. The soil penetration resistance (SPR) was determined in 20 soil samples per treatment and layer (0.0-0.10; 0.10-0.20 and 0.20-0.30 m) for each soil matric potential: -6, -10, -33, -100, -500 kPa. The SPR was determined at a volumetric soil water content equivalent to the fraction of plant-available water of 0.7. There were no differences of soil penetration resistance between the two cropping systems. Differences in soil penetration resistance among tillage systems were related to the matric potential at which the samples were equilibrated. The critical SPR limit of 2 MPa normally used for conventional tillage should be maintained. However, this value of 2 MPa is inappropriate for the physical quality characterization of Rhodic Eutrudox under no-tillage and/or minimum tillage with chiseling. Regardless of the cropping systems, the critical SPR limit should be raised to 3 MPa for minimum tillage with chiseling and to 3.5 MPa for no-tillage.

Binding of double-strand breaks in DNA by human Rad52 protein.

Double-strand breaks (DSBs) in DNA are caused by ionizing radiation. These chromosomal breaks can kill the cell unless repaired efficiently, and inefficient or inappropriate repair can lead to mutation, gene translocation and cancer. Two proteins that participate in the repair of DSBs are Rad52 and Ku: in lower eukaryotes such as yeast, DSBs are repaired by Rad52-dependent homologous recombination, whereas vertebrates repair DSBs primarily by Ku-dependent non-homologous end-joining. The contribution of homologous recombination to vertebrate DSB repair, however, is important. Biochemical studies indicate that Ku binds to DNA ends and facilitates end-joining. Here we show that human Rad52, like Ku, binds directly to DSBs, protects them from exonuclease attack and facilitates end-to-end interactions. A model for repair is proposed in which either Ku or Rad52 binds the DSB. Ku directs DSBs into the non-homologous end-joining repair pathway, whereas Rad52 initiates repair by homologous recombination. Ku and Rad52, therefore, direct entry into alternative pathways for the repair of DNA breaks.

Systematic assessment of items influencing Crohn's disease patient's preferences in selecting an anti-TNF agent (Choose TNF trial)

Background and Aims: The three anti-TNF agents infliximab (IFX), adalimumab (ADA) andcertolizumab pegol (CZP) have demonstrated similar efficacy in induction and maintenanceof response and remission in Crohn's disease (CD) treatment. Given the comparability ofthese drugs, patient's preferences may influence the choice of the product. However, dataon patient's preferences for choosing anti-TNF agents are lacking. We therefore aimed toassess the CD patient's appraisal to select the drug of his choice and to identify factorsguiding this decision.Methods: A prospective survey among anti-TNF-naive CD patientswas performed. Patients were provided a description of the three anti-TNF agents focusingon indication, application mode (s.c. vs. i.v.), application time intervals, setting of application(hospital vs. private practice vs. patient's home), average time to apply the medication permonth, typical side effects, and the scientific evidence of efficacy and safety available for everydrug. Patients answered a questionnaire consisting of 17 questions, covering demographic,disease-specific, and medication data.Results: Hundred patients (47f/53m, mean age 45±16years) completed the questionnaire. Disease duration was <1year in 7%, 1-5 years in 31%,and >5 years in 62% of patients. Disease location was ileal in 33%, colonic in 40%, andileocolonic in 27%. Disease phenotype was inflammatory in 68%, stenosing in 29%, andinternally fistulizing in 3% of patients. Additionally, 20% had perianal fistulizing disease.Patients were already treated with the following drugs: mesalamines 61%, budesonide 44%,prednisone 97%, thiopurines 78%, methotrexate 16%. In total, 30% had already heardabout IFX, 20% about ADA, and 11% about CZP. Thirty-six percent voted for treatmentwith ADA, 28% for CZP, and 25% for IFX, whereas 11% were undecided. The followingfactors influenced the patient's decision for choosing a specific anti-TNF drug (severalanswers possible): side effects 76%, physician's recommendation 66%, application mode54%, efficacy experience 52%, time to spend for therapy 27%, patient's recommendations21%, interactions with other medications 12%. The single most important factor for choosinga specific anti-TNF was (1 answer): side effect profile 35%, physician's recommendation22%, efficacy experience 21%, application mode 13%, patient's recommendations 5%, timespent for therapy 3%, interaction with other medications 1%.Conclusions: The majority ofpatients preferred anti-TNF syringes to infusions. The safety profile of the drugs and thephysician's recommendation are major factors influencing the patient's choice for a specificanti-TNF drug. Patient's issues about safety and lifestyle habits should be taken into accountwhen prescribing specific anti-TNF formulations.

COMPRESSIBLITY AND PENETRABILITY OF LATOSSOLO VERMELHO-AMARELO DISTRÓFICO (OXISOL) UNDER VARIED MANAGEMENT SYSTEMS AND LAND USES

Soil compaction is one of the main degradation causes, provoked by inappropriate agricultural practices that override the limitations of the soil physical properties. Preconsolidation pressure and penetration resistance have proved effective as alternative to assess and identify soil compaction. Based on the interpretation of these physico-mechanical parameters, compaction can be prevented with a better adjusted soil management. This study was performed to generate preconsolidation pressure and penetration resistance models for Latososlo Vermelho-Amarelo distrófico (Oxisol) under various managements and uses; and evaluate which of these would lead to degradation or degradation susceptibility. The study was carried out in Curvelo, MG. Two managements and one land use were evaluated: no-tillage, sheep grazing and natural forest. Undisturbed soil samples collected from the 0-5 cm layer were subjected to uniaxial compression and penetration resistance tests. Preconsolidation pressure models for forest and no-tillage soils were not statistically different, demonstrating a low degradation potential in no-tillage systems. Preconsolidation pressure was higher in soil under sheep grazing at all water retention tensions and penetration resistance values were higher than under native forest indicating animal trampling as a potential degradation factor. Neither management presented penetration resistance values above 2 MPa at field capacity moisture. Only under sheep grazing the soil penetrability was near 2 MPa at field capacity and values greater than 2 MPa at 0.2 kg kg-1.

Stress ulcer prophylaxis in non-critically ill patients: a prospective evaluation of current practice in a general surgery department

Objective: There is little evidence regarding the benefit of stress ulcer prophylaxis (SUP) outside critical care setting. Over-prescription of SUP is not devoid of risks. This prospective study aimed to evaluate the use of proton pump inhibitors (PPIs) for SUP in a general surgery department.Methods: Data collection was performed prospectively during an 8-week period on patients hospitalized in a general surgery department (58 beds) by pharmacists. Patients with a PPI prescription for the treatment of ulcers, gastro-oesophageal reflux disease, oesophagitis or epigastric pain were excluded. Patients admitted twice during the study period were not re-included. The American Society of Health-System Pharmacists guidelines on SUP were used to assess the appropriateness of de novo PPI prescriptions.Results: Among 255 consecutive patients in the study, 138 (54%) received a prophylaxis with PPI, of which 86 (62%) were de novo PPI prescriptions. One-hundred twenty-nine patients (94%) received esomeprazole (according to the hospital drug policy). The most frequent dosage was 40 mg/day. Use of PPI for SUP was evaluated in 67 patients. Fifty-three patients (79%) had no risk factors for SUP. Twelve and 2 patients had one or two risk factors, respectively. At discharge, PPI prophylaxis was continued in 34% of patients with a de novo PPI prescription.Conclusion: This study highlights the overuse of PPIs in non-ICU patients and the inappropriate continuation of PPI prescriptions at discharge.Treatment

Novel anesthetics and other treatment strategies for refractory status epilepticus

Refractory status epilepticus (RSE)-that is, seizures resistant to at least two antiepileptic drugs (AEDs)-is generally managed with barbiturates, propofol, or midazolam, despite a low level of evidence (Rossetti, 2007). When this approach fails, the need for alternative pharmacologic and nonpharmacologic strategies emerges. These have been investigated even less systematically than the aforementioned compounds, and are often used, sometimes in succession, in cases of extreme refractoriness (Robakis & Hirsch, 2006). Several possibilities are reviewed here. In view of the marked heterogeneity of reported information, etiologies, ages, and comedications, it is extremely difficult to evaluate a given method, not to say to compare different strategies among them. Pharmacologic Approaches Isoflurane and desflurane may complete the armamentarium of anesthetics,' and should be employed in a ''close'' environment, in order to prevent intoxication of treating personnel. c-Aminobutyric acid (GABA)A receptor potentiation represents the putative mechanism of action. In an earlier report, isoflurane was used for up to 55 h in nine patients, controlling seizures in all; mortality was, however, 67% (Kofke et al., 1989). More recently, the use of these inhalational anesthetics was described in seven subjects with RSE, for up to 26 days, with an endtidal concentration of 1.2-5%. All patients required vasopressors, and paralytic ileus occurred in three; outcome was fatal in three patients (43%) (Mirsattari et al., 2004). Ketamine, known as an emergency anesthetic because of its favorable hemodynamic profile, is an N-methyl-daspartate (NMDA) antagonist; the interest for its use in RSE derives from animal works showing loss of GABAA efficacy and maintained NMDA sensitivity in prolonged status epilepticus (Mazarati & Wasterlain, 1999). However, to avoid possible neurotoxicity, it appears safer to combine ketamine with GABAergic compounds (Jevtovic-Todorovic et al., 2001; Ubogu et al., 2003), also because of a likely synergistic effect (Martin & Kapur, 2008). There are few reported cases in humans, describing progressive dosages up to 7.5 mg/kg/h for several days (Sheth & Gidal, 1998; Quigg et al., 2002; Pruss & Holtkamp, 2008), with moderate outcomes. Paraldehyde acts through a yet-unidentified mechanism, and appears to be relatively safe in terms of cardiovascular tolerability (Ramsay, 1989; Thulasimani & Ramaswamy, 2002), but because of the risk of crystal formation and its reactivity with plastic, it should be used only as fresh prepared solution in glass devices (Beyenburg et al., 2000). There are virtually no recent reports regarding its use in adults RSE, whereas rectal paraldehyde in children with status epilepticus resistant to benzodiazepines seems less efficacious than intravenous phenytoin (Chin et al., 2008). Etomidate is another anesthetic agent for which the exact mechanism of action is also unknown, which is also relatively favorable regarding cardiovascular side effects, and may be used for rapid sedation. Its use in RSE was reported in eight subjects (Yeoman et al., 1989). After a bolus of 0.3 mg/kg, a drip of up to 7.2 mg/kg/h for up to 12 days was administered, with hypotension occurring in five patients; two patients died. A reversible inhibition of cortisol synthesis represents an important concern, limiting its widespread use and implying a careful hormonal substitution during treatment (Beyenburg et al., 2000). Several nonsedating approaches have been reported. The use of lidocaine in RSE, a class Ib antiarrhythmic agent modulating sodium channels, was reviewed in 1997 (Walker & Slovis, 1997). Initial boluses up to 5 mg/kg and perfusions of up to 6 mg/kg/h have been mentioned; somewhat surprisingly, at times lidocaine seemed to be successful in controlling seizures in patients who were refractory to phenytoin. The aforementioned dosages should not be overshot, in order to keep lidocaine levels under 5 mg/L and avoid seizure induction (Hamano et al., 2006). A recent pediatric retrospective survey on 57 RSE episodes (37 patients) described a response in 36%, and no major adverse events; mortality was not given (Hamano et al., 2006 Verapamil, a calcium-channel blocker, also inhibits P-glycoprotein, a multidrug transporter that may diminish AED availability in the brain (Potschka et al., 2002). Few case reports on its use in humans are available; this medication nevertheless appears relatively safe (under cardiac monitoring) up to dosages of 360 mg/day (Iannetti et al., 2005). Magnesium, a widely used agent for seizures elicited by eclampsia, has also been anecdotally reported in RSE (Fisher et al., 1988; Robakis & Hirsch, 2006), but with scarce results even at serum levels of 14 mm. The rationale may be found in the physiologic blockage of NMDA channels by magnesium ions (Hope & Blumenfeld, 2005). Ketogenic diet has been prescribed for decades, mostly in children, to control refractory seizures. Its use in RSE as ''ultima ratio'' has been occasionally described: three of six children (Francois et al., 2003) and one adult (Bodenant et al., 2008) were responders. This approach displays its effect subacutely over several days to a few weeks. Because ''malignant RSE'' seems at times to be the consequence of immunologic processes (Holtkamp et al., 2005), a course of immunomodulatory treatment is often advocated in this setting, even in the absence of definite autoimmune etiologies (Robakis & Hirsch, 2006); steroids, adrenocorticotropic hormone (ACTH), plasma exchanges, or intravenous immunoglobulins may be used alone or in sequential combination. Nonpharmacologic Approaches These strategies are described somewhat less frequently than pharmacologic approaches. Acute implantation of vagus nerve stimulation (VNS) has been reported in RSE (Winston et al., 2001; Patwardhan et al., 2005; De Herdt et al., 2009). Stimulation was usually initiated in the operation room, and intensity progressively adapted over a few days up to 1.25 mA (with various regimens regarding the other parameters), allowing a subacute seizure control; one transitory episode of bradycardia/asystole has been described (De Herdt et al., 2009). Of course, pending identification of a definite seizure focus, resective surgery may also be considered in selected cases (Lhatoo & Alexopoulos, 2007). Low-frequency (0.5 Hz) transcranial magnetic stimulation (TMS) at 90% of the resting motor threshold has been reported to be successful for about 2 months in a patient with epilepsia partialis continua, but with a weaning effect afterward, implying the need for a repetitive use (Misawa et al., 2005). More recently, TMS was applied in a combination of a short ''priming'' high frequency (up to 100 Hz) and longer runs of low-frequency stimulations (1 Hz) at 90-100% of the motor threshold in seven other patients with simple-partial status, with mixed results (Rotenberg et al., 2009). Paradoxically at first glance, electroconvulsive treatment may be found in cases of extremely resistant RSE. A recent case report illustrates its use in an adult patient with convulsive status, with three sessions (three convulsions each) carried out over 3 days, resulting in a moderate recovery; the mechanism is believed to be related to modification of the synaptic release of neurotransmitters (Cline & Roos, 2007). Therapeutic hypothermia, which is increasingly used in postanoxic patients (Oddo et al., 2008), has been the object of a recent case series in RSE (Corry et al., 2008). Reduction of energy demand, excitatory neurotransmission, and neuroprotective effects may account for the putative mechanism of action. Four adult patients in RSE were cooled to 31_-34_C with an endovascular system for up to 90 h, and then passively rewarmed over 2-50 h. Seizures were controlled in two patients, one of whom died; also one of the other two patients in whom seizures continued subsequently deceased. Possible side effects are related to acid-base and electrolyte disturbances, and coagulation dysfunction including thrombosis, infectious risks, cardiac arrhythmia, and paralytic ileus (Corry et al., 2008; Cereda et al., 2009). Finally, anecdotic evidence suggests that cerebrospinal fluid (CSF)-air exchange may induce some transitory benefit in RSE (Kohrmann et al., 2006); although this approach was already in use in the middle of the twentieth century, the mechanism is unknown. Acknowledgment A wide spectrum of pharmacologic (sedating and nonsedating) and nonpharmacologic (surgical, or involving electrical stimulation) regimens might be applied to attempt RSE control. Their use should be considered only after refractoriness to AED or anesthetics displaying a higher level of evidence. Although it seems unlikely that these uncommon and scarcely studied strategies will influence the RSE outcome in a decisive way, some may be interesting in particular settings. However, because the main prognostic determinant in status epilepticus appears to be related to the underlying etiology rather than to the treatment approach (Rossetti et al., 2005, 2008), the safety issue should always represent a paramount concern for the prescribing physician. Conclusion The author confirms that he has read the Journal's position on issues involved in ethical publication and affirms that this paper is consistent with those guidelines.

Amélioration de l'hyperglycémie hospitalière: création d'un nouveau modèle de gouvernance [Improvement of hospital hyperglycemia: creation of a new governance model?].

The report of significant decrease of the inpatient hospital mortality and morbidity with an efficient insulin therapy has demonstrated the need of a good glycaemic control for patients hospitalised in acute care. However, one is faced with numerous difficulties in the hospital management of patients with hyperglycaemia, errors often occur when prescribing insulin, and the management skills are insufficient. Our goal is to change the medical and nursing practices to evolve towards an efficient and safe management of the hospitalised patient. The model we lay out in this article is based upon observation of the therapeutic support of patients with a chronic condition, whilst using a systemic management approach.

Prospective monitoring of cefepime in intensive care unit adult patients.

INTRODUCTION: Cefepime has been associated with a greater risk of mortality than other beta-lactams in patients treated for severe sepsis. Hypotheses for this failure include possible hidden side-effects (for example, neurological) or inappropriate pharmacokinetic/pharmacodynamic (PK/PD) parameters for bacteria with cefepime minimal inhibitory concentrations (MIC) at the highest limits of susceptibility (8 mg/l) or intermediate-resistance (16 mg/l) for pathogens such as Enterobacteriaceae, Pseudomonas aeruginosa and Staphylococcus aureus. We examined these issues in a prospective non-interventional study of 21 consecutive intensive care unit (ICU) adult patients treated with cefepime for nosocomial pneumonia. METHODS: Patients (median age 55.1 years, range 21.8 to 81.2) received intravenous cefepime at 2 g every 12 hours for creatinine clearance (CLCr) >or= 50 ml/min, and 2 g every 24 hours or 36 hours for CLCr < 50 ml/minute. Cefepime plasma concentrations were determined at several time-points before and after drug administration by high-pressure liquid chromatography. PK/PD parameters were computed by standard non-compartmental analysis. RESULTS: Seventeen first-doses and 11 steady states (that is, four to six days after the first dose) were measured. Plasma levels varied greatly between individuals, from two- to three-fold at peak-concentrations to up to 40-fold at trough-concentrations. Nineteen out of 21 (90%) patients had PK/PD parameters comparable to literature values. Twenty-one of 21 (100%) patients had appropriate duration of cefepime concentrations above the MIC (T>MIC >or= 50%) for the pathogens recovered in this study (MIC <or= 4 mg/l), but only 45 to 65% of them had appropriate coverage for potential pathogens with cefepime MIC >or= 8 mg/l. Moreover, 2/21 (10%) patients with renal impairment (CLCr < 30 ml/minute) demonstrated accumulation of cefepime in the plasma (trough concentrations of 20 to 30 mg/l) in spite of dosage adjustment. Both had symptoms compatible with non-convulsive epilepsy (confusion and muscle jerks) that were not attributed to cefepime-toxicity until plasma levels were disclosed to the caretakers and symptoms resolved promptly after drug arrest. CONCLUSIONS: These empirical results confirm the suspected risks of hidden side-effects and inappropriate PK/PD parameters (for pathogens with upper-limit MICs) in a population of ICU adult patients. Moreover, it identifies a safety and efficacy window for cefepime doses of 2 g every 12 hours in patients with a CLCr >or= 50 ml/minute infected by pathogens with cefepime MICs <or= 4 mg/l. On the other hand, prompt monitoring of cefepime plasma levels should be considered in case of lower CLCr or greater MICs.

Outpatient Quick Diagnosis Units for the evaluation of suspected severe diseases: an observational, descriptive study

Background: Hospitals in countries with public health systems have recently adopted organizational changes to improve efficiency and resource allocation, and reducing inappropriate hospitalizations has been established as an important goal. AIMS: Our goal was to describe the functioning of a Quick Diagnosis Unit in a Spanish public university hospital after evaluating 1,000 consecutive patients. We also aimed to ascertain the degree of satisfaction among Quick Diagnosis Unit patients and the costs of the model compared to conventional hospitalization practices. DESIGN: Observational, descriptive study. METHODS: Our sample comprised 1,000 patients evaluated between November 2008 and January 2010 in the Quick Diagnosis Unit of a tertiary university public hospital in Barcelona. Included patients were those who had potentially severe diseases and would normally require hospital admission for diagnosis but whose general condition allowed outpatient treatment. We analyzed several variables, including time to diagnosis, final diagnoses and hospitalizations avoided, and we also investigated the mean cost (as compared to conventional hospitalization) and the patients' satisfaction. RESULTS: In 88% of cases, the reasons for consultation were anemia, anorexia-cachexia syndrome, febrile syndrome, adenopathies, abdominal pain, chronic diarrhea and lung abnormalities. The most frequent diagnoses were cancer (18.8%; mainly colon cancer and lymphoma) and Iron-deficiency anemia (18%). The mean time to diagnosis was 9.2 days (range 1 to 19 days). An estimated 12.5 admissions/day in a one-year period (in the internal medicine department) were avoided. In a subgroup analysis, the mean cost per process (admission-discharge) for a conventional hospitalization was 3,416.13 Euros, while it was 735.65 Euros in the Quick Diagnosis Unit. Patients expressed a high degree of satisfaction with Quick Diagnosis Unit care. CONCLUSIONS: Quick Diagnosis Units represent a useful and cost-saving model for the diagnostic study of patients with potentially severe diseases. Future randomized study designs involving comparisons between controls and intervention groups would help elucidate the usefulness of Quick Diagnosis Units as an alternative to conventional hospitalization.

Involvement of the RasGAP derived fragment N in the resistance of pancreatic beta cells towards apoptosis

RESUME DESTINE AUX NON SCIENTIFIQUESLe diabète est une maladie associée à un excès de glucose (sucre) dans le sang. Le taux de glucose sanguin augmente lorsque l'action d'une hormone, l'insuline, responsable du transport du glucose du sang vers les tissus de l'organisme diminue, ou lorsque les quantités d'insuline à disposition sont inadéquates.L'une des causes communes entre les deux grands types de diabète connus, le type 1 et le type 2, est la disparition des cellules beta du pancréas, spécialisées dans la sécrétion d'insuline, par mort cellulaire programmée aussi appelée apoptose. Alors que dans le diabète de type 1, la destruction des cellules beta est causée par notre propre système immunitaire, dans le diabète de type 2, la mort de ces cellules, est principalement causée par des concentrations élevées de graisses saturés ou de molécules impliquées dans l'inflammation que l'on rencontre en quantités augmentées chez les personnes obèses. Etant donné l'augmentation épidémique du nombre de personnes obèses de par le monde, on estime que le nombre de personnes diabétiques (dont une majorité sont des diabétiques de type 2), va passer de 171 million en l'an 2000, à 366 million en l'an 2030, expliquant la nécessité absolue de mettre au point de nouvelles stratégies thérapeutique pour combattre cette maladie.L'apoptose est un processus complexe dont la dérégulation induit de nombreuses affections allant du cancer jusqu'au diabète. L'activation de caspase 3, une protéine clé contrôlant la mort cellulaire, était connue pour systématiquement mener à la mort cellulaire programmée. Ces dernières années, notre laboratoire a décrit des mécanismes de survie qui sont activés par caspase 3 et qui expliquent sans doute pourquoi son activation ne mène pas systématiquement à la mort cellulaire. Lorsqu'elle est faiblement activée, caspase 3 clive une autre protéine appelée RasGAP en deux protéines plus courtes dont l'une, appelée le fragment Ν a la particularité de protéger les cellules contre l'apoptose.Durant ma thèse, j'ai été impliqué dans divers projets destinés à mieux comprendre comment le fragment Ν protégeait les cellules contre l'apoptose et à savoir s'il pouvait être utilisé comme outil thérapeutique dans les conditions de survenue d'un diabète expérimental. C'est dans ce but que nous avons créé une souris transgénique, appelée RIP-N, exprimant le fragment Ν spécifiquement dans les cellules beta. Comme attendu, les cellules beta de ces souris étaient plus résistantes à la mort induite par des composés connus pour induire le diabète, comme certaines molécules induisant l'inflammation ou les graisses saturées. Nous avons ensuite pu montrer que les souris RIP-N étaient plus résistantes à la survenue d'un diabète expérimental que ce soit par l'injection d'une drogue induisant l'apoptose des cellules beta, que ce soit dans un fond génétique caractérisé par une attaque spontanée des cellules beta par le système immunitaire ou dans le contexte d'un diabète de type 2 induit par l'obésité. Dans plusieurs des modèles animaux étudiés, nous avons pu montrer que le fragment Ν protégeait les cellules en activant une voie protectrice bien connue impliquant successivement les protéines Ras, PI3K et Akt ainsi qu'en bloquant la capacité d'Akt d'activer le facteur NFKB, connu pour être délétère pour la survie de la cellule beta. La capacité qu'a le fragment Ν d'activer Akt tout en prévenant l'activation de NFKB par Akt est par conséquent particulièrement intéressante dans l'intégration des signaux régulant la mort cellulaire dans le contexte de la survenue d'un diabète.La perspective d'utiliser le fragment Ν comme outil thérapeutique dépendra de notre capacité à activer les signaux protecteurs induits par le fragment Ν depuis l'extérieur de la cellule ou de dériver des peptides perméables aux cellules possédant les propriétés du fragment N.2 SUMMARYDiabetes mellitus is an illness associated with excess blood glucose. Blood glucose levels raise when the action of insulin decreases or when insulin is provided in inappropriate amounts. In type 1 diabetes (T1D) as well as in type 2 diabetes (T2D), the insulin secreting beta cells in the pancreas undergo controlled cell death also called apoptosis. Whereas in T1D, beta cells are killed by the immune system, in T2D, they are killed by several factors, among which are increased blood glucose levels, increased levels of harmful lipids or pro-inflammatory cytokines that are released by the dysfunctional fat tissue of obese people. Given the epidemic increase in the number of obese people throughout the world, the number of diabetic people (a majority of which are type 2 diabetes) is estimated to rise from 171 million affected people in the year 2000 to 366 million in 2030 explaining the absolute requirement for new therapies to fight the disease.Apoptosis is a very complex process whose deregulation leads to a wide range of diseases going from cancer to diabetes. Caspase 3 although known as a key molecule controlling apoptosis, has been shown to have various other functions. In the past few years, our laboratory has described a survival mechanism, that takes place at low caspase activity and that might explain how cells that activate their caspases for reasons other than apoptosis survive. In such conditions, caspase 3 cleaves another protein called RasGAP into two shorter proteins, one of which, called fragment N, protects cells from apoptosis.We decided to check whether fragment Ν could be used as a therapeutical tool in the context of diabetes inducing conditions. We thus derived a transgenic mouse line, called RIP-N, in which the expression of fragment Ν is restricted to beta cells. As expected, the beta cells of these mice were more resistant ex-vivo to cell death induced by diabetes inducing factors. We then showed that the RIP-N transgenic mice were resistant to streptozotocin induced diabetes, a mouse model mimicking type 1 diabetes, which correlated to fewer number of apoptotic beta cells in the pancreas of the transgenic mice compared to their controls. The RIP-N transgene also delayed overt diabetes development in the NOD background, a mouse model of autoimmune type 1 diabetes, and delayed the occurrence of obesity induced hyperglycemia in a mouse model of type 2-like diabetes. Interestingly, fragment Ν was mediating its protection by activating the protective Akt kinase, and by blocking the detrimental NFKB factor. Our future ability to activate the protective signals elicited by fragment Ν from the outside of cells or to derive cell permeable peptides bearing the protective properties of fragment Ν might condition our ability to use this protein as a therapeutic tool.3 RESUMELe diabète est une maladie associée à un excès de glucose plasmatique. La glycémie augmente lorsque l'action de l'insuline diminue ou lorsque les quantités d'insuline à disposition sont inadéquates. Dans le diabète de type 1 (D1) comme dans le diabète de type 2 (D2), les cellules beta du pancréas subissent la mort cellulaire programmée aussi appelée apoptose. Alors que dans le D1 les cellules beta sont tuées par le système immunitaire, dans le D2 elles sont tuées par divers facteurs parmi lesquels on trouve des concentrations élevées de glucose, d'acides gras saturés ou de cytokines pro-inflammatoires qui sont sécrétées en concentrations augmentées par le tissu adipeux dysfonctionnel des personnes obèses. Etant donné l'augmentation épidémique du nombre de personnes obèses de par le monde, on estime que le nombre de personnes diabétiques (dont une majorité sont des diabétiques de type 2), va passer de 171 million en l'an 2000, à 366 million en l'an 2030, justifiant la nécessité absolue de mettre au point de nouvelles stratégies thérapeutique pour combattre cette maladie.L'apoptose est un processus complexe dont la dérégulation induit de nombreuses affections allant du cancer jusqu'au diabète. Caspase 3, bien que connue comme étant une protéine clé contrôlant l'apoptose a bien d'autres fonctions démontrées. Ces dernières années, notre laboratoire a décrit un mécanisme de survie qui est activé lorsque caspase 3 est faiblement activée et qui explique probablement comment des cellules qui ont activé leurs caspases pour une autre raison que l'apoptose peuvent survivre. Dans ces conditions, caspase 3 clive une autre protéine appelée RasGAP en deux protéines plus courtes dont l'une, appelée le fragment Ν a la particularité de protéger les cellules contre l'apoptose.Nous avons donc décidé de vérifier si le fragment Ν pouvait être utilisé comme outil thérapeutique dans les conditions de survenue d'un diabète expérimental. Pour se faire, nous avons créé une souris transgénique, appelée RIP-N, exprimant le fragment Ν spécifiquement dans les cellules beta. Comme attendu, les cellules beta de ces souris étaient plus résistantes ex-vivo à la mort induite par des facteurs pro-diabétogènes. Nous avons ensuite pu montrer que les souris RIP-N étaient plus résistantes à la survenue d'un diabète induit par la streptozotocine, un drogue mimant la survenue d'un D1 et que ceci était corrélée à une diminution du nombre de cellules en apoptose dans le pancréas des souris transgéniques comparé à leurs contrôles. L'expression du transgène a aussi eu pour effet de retarder la survenue d'un diabète franc dans le fond génétique NOD, un modèle génétique de diabète de type 1 auto-immun, ainsi que de retarder la survenue d'une hyperglycémie dans un modèle murin de diabète de type 2 induit par l'obésité. Dans plusieurs des modèles animaux étudiés, nous avons pu montrer que le fragment Ν protégeait les cellules en activant la kinase protectrice Akt ainsi qu'en bloquant le facteur délétère NFKB. La perspective d'utiliser le fragment Ν comme outil thérapeutique dépendra de notre capacité à activer les signaux protecteurs induits par le fragment Ν depuis l'extérieur de la cellule ou de dériver des peptides perméables aux cellules possédant les propriétés du fragment

Reduced choroidal neovascularization by AAV-anti-VEGF shRNA delivery

VEGF plays an essential role in ocular angiogenic diseases including the late-stage form of AMD, the primary cause of vision loss in the western world. Over-expression of VEGF leads to development of vasculature emanating from the choroid, invading the subretinal space through breaks in Bruch's membrane. Strategies leading to long-term suppression of inappropriate ocular angiogenesis are required. A panel of 10 shRNAs targeting the coding region of human VEGF165 was tested in HEK293 cells and in the human retinal pigment epithelial cell line, ARPE-19. VEGF knock-down up to 92% was achieved by co-transfecting shRNAexpressing constructs with plasmid encoding the Renilla luciferase gene fused to the VEGF165 sequence. For in vivo delivery of the most potent shRNA cassette, both single-stranded and self-complementary rAAV vectors were packaged in serotype 8 capsids. Intramuscular administration in mice led to localized expression and 96% knock-down of endogenous VEGF. Using eGFP as a marker, efficient gene transfer of retinal pigment epithelial cells, the cells thought to be responsible for the abnormal VEGF production, was obtained by subretinal delivery of rAAV2.8 vectors. The capacity of rAAV-encoded shRNAs to silence endogenous VEGF gene expression was evaluated in the laser-induced murine model of choroidal neovascularization (CNV). In this mouse model of AMD, sizes of the CNV were found to be significantly reduced following rAAV-shRNA subretinal delivery. Thus, our results indicate that gene transfer combining AAV-mediated delivery with triggering of the endogenous RNAi pathway can be used for anti-VEGF therapy and holds great promise for the treatment of AMD.

First explicit criteria to decide on the appropriateness of therapy of ulcerative colitis: the European EPATUC panel

Background: Ulcerative colitis (UC) is a chronic disease with a wide variety of treatment options many of which are not evidence based. Supplementing available guidelines, which are often broadly defined, consensus-based and generally not tailored to specifically reflect the individual patient situation, we developed explicit appropriateness criteria to assist, and improve treatment decisions. Methods: We used the RAND appropriateness method which does not force consensus. An extensive literature review was compiled based on and supplementing, where necessary, the ECCO UC 2011 guidelines. EPATUC (endorsed by ECCO) was formed by 7 gastroenterologists, 2 surgeons and 2 general practitioners from throughout Europe. Clinical scenarios reflecting practice were rated on a 9-point scale from 1 (extremely inappropriate) to 9 (extremely appropriate), based on the expert's experience and the available literature. After extensive discussion, all scenarios were re-rated at a two-day panel meeting. Median and disagreement (D) were used to categorize ratings into 3 categories: appropriate (A), uncertain (U) and inappropriate (I). Results: 718 clinical scenarios were rated, structured in 13 main clinical presentations: not refractory (n = 64) or refractory (n = 33) proctitis, mild to moderate left-sided (n = 72) or extensive (n = 48) colitis, severe colitis (n = 36), steroid- dependant colitis (n = 36), steroid-refractory colitis (n = 55), acute pouchitis (n = 96), maintenance of remission (n = 248), colorectal cancer prevention (n = 9) and fulminant colitis (n = 9). Overall, 100 indications were judged appropriate (14%), 129 uncertain (18%) and 489 inappropriate (68%). Disagreement between experts was very low (6%). Conclusions: For the very first time, explicit appropriateness criteria for therapy of UC were developed that allow both specific and rapid therapeutic decision making and prospective assessment of treatment appropriateness. Comparison of these detailed scenarios with patient profiles encountered in the Swiss IBD cohort study indicates good concordance. EPATUC criteria will be freely accessible on the internet (epatuc.ch)

Prevalence, incidence and indications of hypno-sedative treatments in hospitalized patients aged 75 and older

Introduction Sleep impairment is a common problem in older persons and is associated with adverse health and economic consequences. The most prescribed hypnosedatives in Switzerland are benzodiazepines, benzodiazepine-like hypnosedatives and clomethiazole. In the elderly persons, these medications have been associated with decreased cognitive performance, an increased risk of addiction and an increased risk of falls. The main aim of this study was to determine the prevalence, incidence and indications of hypnosedative prescriptions in older in-patients admitted to a Swiss university hospital. Furthermore, we planned to investigate the possible relationship between hypnosedatives' prescription and patients' falls during the hospital stay and we compared the prevalence of hypnosedative drugs' prescribing between geriatric and non-geriatric clinical wards.Materials & Methods This observational study included all patients aged 75 years or older admitted to a Swiss university hospital for 24 h or more. Prescription-related data were recorded every day for 26 consecutive days in the two geriatric wards, and twice, a week apart, in three non-geriatric wards.Results A total of 204 patients (mean age 84.7 ± 0.7 years, 66% women) were included. The prevalence of hypnosedative drugs prescription was 73% and the incidence was 28% on the two geriatric wards. The most common indications for a prescription on geriatric wards was insomnia (51%), followed by continuation of usual treatment (12%). The most frequently prescribed hypnosedative drug was clomethiazole (64% of the patients), followed by lorazepam (11%). We were not able to demonstrate any significant correlation between the number of hypnosedative drugs prescribed and the number of falls during hospital stay. The prevalence of hypnosedative drugs prescriptions was similar between geriatric and non-geriatric wards. The proportion of patients treated by at least one hypnosedative increased between hospital admission (32%) and discharge (45%).Discussions, Conclusion This study reveals a high prevalence of hypnosedative prescription in medical in-patients aged 75 years or older. Main indication was insomnia and the most frequently prescribed hypnosedative molecule was clomethiazole. Prescription incidence was also very high and the proportion of patients being prescribed a hypnosedative increased during hospital stay. Systematic interventions need to be integrated in order to reduce elderly inpatient exposure to treatments that are associated with adverse health outcomes.Conference information: ESCP 40th International Symposium on Clinical Pharmacy. Clinical Pharmacy: Connecting Care and Outcomes Dublin Ireland, 19-21 October 2011