Stable transmission of targeted gene modification using single-stranded oligonucleotides with flanking LNAs.

Targeted mutagenesis directed by oligonucleotides (ONs) is a promising method for manipulating the genome in higher eukaryotes. In this study, we have compared gene editing by different ONs on two new target sequences, the eBFP and the rd1 mutant photoreceptor betaPDE cDNAs, which were integrated as single copy transgenes at the same genomic site in 293T cells. Interestingly, antisense ONs were superior to sense ONs for one target only, showing that target sequence can by itself impart strand-bias in gene editing. The most efficient ONs were short 25 nt ONs with flanking locked nucleic acids (LNAs), a chemistry that had only been tested for targeted nucleotide mutagenesis in yeast, and 25 nt ONs with phosphorothioate linkages. We showed that LNA-modified ONs mediate dose-dependent target modification and analyzed the importance of LNA position and content. Importantly, when using ONs with flanking LNAs, targeted gene modification was stably transmitted during cell division, which allowed reliable cloning of modified cells, a feature essential for further applications in functional genomics and gene therapy. Finally, we showed that ONs with flanking LNAs aimed at correcting the rd1 stop mutation could promote survival of photoreceptors in retinas of rd1 mutant mice, suggesting that they are also active in vivo.

Changing the awareness of low vitamin D status in a rheumatology population: a pre/post-study.

INTRODUCTION: In 2009 hypovitaminosis D was highly prevalent in a population of Swiss rheumatology patients (86%). We aimed to evaluate the evolution of vitamin D status in the same population two years later, after the results of the first study were disseminated to local physicians and patients, in order to determine the evolution of the problem and the impact of physician information. METHOD: Patients in our rheumatology clinic were screened for 25-OH vitamin D. Results were categorised as: deficient (<10 ng/ml or <25 nmol/l), insufficient (10 to 30 ng/ml or 25 to 75 nmol/l) or normal (>30 ng/ml or >75 nmol/l). We also used another cut-off of 20 ng/ml (50 nmol/l). We evaluated the evolution of 25-OH vitamin D dosages and vitamin D3 prescriptions between 2008 and 2011 in our institution and the number of publications on vitamin D in three important medical journals of the French speaking part of Switzerland. RESULTS: Compared with 2009, significantly more patients had normal results in 2011. Fifty-two percent of patients had levels >20 ng/ml in 2009 and 66% in 2011, difference statistically significant (p = 0.001). During the years separating the two study periods the number of 25-OH vitamin D dosages and the prescription of high doses of vitamin D3 increased in our hospital. In addition the number of publications on vitamin D increased between 2008 and 2011. CONCLUSION: We concluded that lower prevalence in hypovitaminosis D is certainly related to better adherence to daily supplements, and to better information and awareness of the physicians about hypovitaminosis D.

Repositioning precision of coronary arteries measured on X-ray angiography and its implications for coronary MR angiography.

BACKGROUND: To test the hypothesis that intervals with superior beat-to-beat coronary artery repositioning precision exist in the cardiac cycle, to design a coronary MR angiography (MRA) methodology in response, and to ascertain its performance. METHODS: Coronary repositioning precision in consecutive heartbeats was measured on x-ray coronary angiograms of 17 patients and periods with the highest repositioning precision were identified. In response, the temporal order of coronary MRA pulse sequence elements required modification and the T2 -prep now follows (T2 -post) rather than precedes the imaging part of the sequence. The performance of T2 -post was quantitatively compared (signal-to-noise [SNR], contrast-to-noise [CNR], vessel sharpness) to that of T2 -prep in vivo. RESULTS: Coronary repositioning precision is <1 mm at peak systole and in mid diastole. When comparing systolic T2 -post to diastolic T2 -prep, CNR and vessel sharpness remained unchanged (both P = NS) but SNR for muscle and blood increased by 104% and 36% (both P < 0.05), respectively. CONCLUSION: Windows with improved coronary repositioning precision exist in the cardiac cycle: one in peak systole and one in mid diastole. Peak-systolic imaging necessitates a re-design of conventional coronary MRA pulse sequences and leads to image quality very similar to that of conventional mid-diastolic data acquisition but improved SNR. J. Magn. Reson. Imaging 2015;41:1251-1258. © 2014 Wiley Periodicals, Inc.

Cytomegalovirus infection and new-onset post-transplant diabetes mellitus.

We analyzed data from all consecutive kidney transplant patients at our institution between April 2003 and October 2006. We found 15 cases of late-onset cytomegalovirus (CMV) infection, two of which developed concurrent post-transplant diabetes mellitus (PTDM). In these two cases, PTDM was transient and normal glucose tolerance was achieved after an eight-wk therapeutic course of oral valganciclovir. These findings suggest that CMV infection after organ transplantation may be associated with concurrent PTDM. The distinct causative relationship is yet to be determined.

Pre- and post-treatment immunodiagnostic evaluation in human schistosomiasis mansoni

Schistosomiasis control seems to be different in countries were low parasitic burden and asymptomatic clinical patients are the features of majority of cases. Immunological methods must substitute the traditional coprologic techniques used for some decades in the Control Program. Circumoval Precipitin Test (COPT), intradermal test and ELISA with soluble egg antigen (SEA) are evaluated for using as tools for seroepidemiologic studies. COPT and ELISA were performed after treatment to known their utility when impact of chemotherapy must be assessed. One hundred sixty five persons were followed up 3, 6, 9 and 12 months after treatment. The mean sensitivity of CPT studied by age groups was 95.6% which is very important considering that 88.4% of the studied population excreted less than 100 egg/gr of feces, while sensitivity of intradermal test was 58.2%. Children showed the highest ractivity to COPT. When treatment is effective, COPT reactivity progressively disminish until become negative one year later. In the non cure group, the COPT reactivity disminished but never below 20%. ELISA-SEA did not modify one year after treatment. Effort should be made to isolate fractions of eggs Schistosoma mansoni whose antibodies disappear after treatment.

Post-traumatic mutism in children: clinical characteristics, pattern of recovery and clinicopathological correlations.

Among the numerous clinical syndromes observed after severe traumatic head injury, post-traumatic mutism is a disorder rarely reported in adults and not studied in any detail in children. We report seven children between the ages of 3 1/2 and 14 years who sustained severe head injury and developed post-traumatic mutism. We aim to give a precise clinical characterization of this disorder, discuss differential diagnosis and correlations with brain imaging and suggest its probable neurological substrate. After a coma lasting from 5 to 25 days, the seven patients who suffered from post-traumatic mutism went through a period of total absence of verbal production lasting from 5 to 94 days, associated with the recovery of non-verbal communication skills and emotional vocalization. During the first days after the recovery of speech, all patients were able to produce correct small sentences with a hypophonic and monotonous voice, moderate dysarthria, word finding difficulties but no signs of aphasia, and preserved oral comprehension. The neurological signs in the acute phase (III nerve paresis in three of seven patients, signs of autonomic dysfunctions in five of seven patients), the results of the brain imaging and the experimental animal data all suggest the involvement of mesencephalic structures as playing a key role in the aetiology of post-traumatic mutism.

Bimodal magmatism as a consequence of the post-collisional readjustment of the thickened Variscan continental lithosphere (Aiguilles Rouges - Mont Blanc Massifs, Western Alps)

High Precision U-Pb zircon and monazite dating in the Aiguilles Rouges-Mont Blanc area allowed discrimination of three short-lived bimodal magmatic pulses: the early 332 Ma Mg-K Pormenaz monzonite and associated 331 Ma peraluminous Montees Pelissier monzogranite; the 307 Ma cordierite-bearing peraluminous Vallorcine and Fully intrusions; and the 303 Fe-K Mont Blanc syenogranite. All intruded syntectonically along major-scale transcurrent faults at a time when the substratum was experiencing tectonic exhumation, active erosion recorded in detrital basins and isothermal decompression melting dated at 327-320 Ma. Mantle activity and magma mixing are evidenced in all plutons by coeval mafic enclaves, stocks and synplutonic dykes. Both crustal and mantle sources evolve through time, pointing to an increasingly warm continental crust and juvenile asthenospheric mantle sources. This overall tectono-magmatic evolution is interpreted in a scenario of post-collisional restoration to normal size of a thickened continental lithosphere. The latter re-equilibrates through delamination and/or erosion of its mantle root and tectonic exhumation/erosion in an overall extensional regime. Extension is related to either gravitational collapse or back-are extension of a distant subduction zone.

Post operative epidural haematomas following spinal surgery : P58

Introduction: Compressive epidural haematomas occurring following spine surgery are very rare but can potentially lead to irreversible damage. The evacuation of the haematoma as an emergency procedure remains the only effective treatment providing though alerting signs are detected on time. Few studies exist on this subject probably due to its rarity. The etiological factors as well as the place of imaging studies prior to urgent haematoma evacuation remain controversial. Two cases of delayed post-operative compressive epidural haematomas following lumbar-spine surgery were detected in our unit between April 2003 and January 2009. In both cases new onset of pain, aggravation of existing neurological deficit or development of new deficit along with worsening of pre-existing walking difficulties were noted. Emergency computer tomography (CT) could not exclude compression in both cases due to important artefacts. Emergency surgery was performed confirming the presence of haematoma in both cases and leading to a complete neurological recovery following its evacuation. As only risk factors common to both cases we identified drain removal and resuming of thromboprophylaxis. Conclusion: Obstacles in early detection of post-operative compressive epidural haematomas occurring following spine surgery are patients presenting with multiple complaints as well as shift work pattern of staff who might not always be trained in detecting early changes in neurological status. We therefore established a checklist for post-operative neurological observations to be carried out on spine surgery patients during the postoperative period. We describe our adopted attitude considering the etiological factors observed in our unit. Further studies including in a multi centre setting would be necessary in order to ascertain our observations.

New Horizons in Forensic Imaging: Post Mortem CT Angiography - Benefits and Practical Application.

Puropse/Aim: To learn about the developement of post mortem CT angiography, its indications, benefits, pitfalls and practical application. Content Organization: A. Developement of post mortem CT angiography B. Technical prerequisites C. Practical application of post mortem CT angiography (preparation of the body, injection of contrast agent, examination protocol) D. Indications and benefits (including a comparison with conventional autopsy) E. Interpretation of imaging data (with case demonstrations) F. Artifacts, pitfalls and limitations G. Current and potential future use. Summary: This exhibit demonstrates the developement, application and interpretation of post mortem CT angiography. Teaching points: 1. post mortem CT angiography is feasible and useful for identification of the cause of death 2. depending on the indication it can be superior to autopsy 3. limitations and artifacts need to be known for interpreta

Three-dimensional cholangio-spiral CT demonstration of a post-traumatic bile leak in a child.

Bilioma is a rare complication of traumatic liver injury, and the precise site of bile leak is often difficult to demonstrate with a non-invasive technique. We report a case of post-traumatic bile leak in a 15-year-old girl in whom spiral CT after intravenous cholangiography allowed excellent preoperative demonstration of the extent of the liver rupture and an exact location of the bile leak. We think that spiral-CT cholangiography could be an accurate, non-invasive technique to investigate the biliary system in cases of paediatric liver trauma.

Ruptured pseudo-aneurysm of a femoral artery in a drug abuser revealed by post-mortem CT angiography.

A 35-year-old drug addict was found dead in a public toilet with a ruptured groin, which was later diagnosed to be a leaking pseudo-aneurysm. Investigation at the scene revealed impressive external hemorrhage related to a groin wound. Post-mortem computed tomography angiography demonstrated an aneurysm of the right femoral artery with leak of contrast liquid. Signs of blood loss were evident at autopsy, and histological examination revealed necrosis and rupture of the pseudo-aneurysm. Toxicological analyses were positive for methadone, cocaine, citalopram, and benzodiazepines. This is the first case report in the literature of a ruptured femoral pseudo-aneurysm with a post-mortem radiological diagnosis.

Neuroprotection in cerebral ischemia : an effect of c-Jun N-terminal kinase inhibition on the inflammatory response

RESUMESuite à un accident vasculaire cérébral (AVC) ischémique, les cellules gliales ducerveau deviennent activées, de nombreuses cellules inflammatoires pénètrent dans letissu lésé et sécrètent une grande variété de cytokines et chémokines. Aujourd'hui, ilexiste des interrogations sur les effets bénéfiques ou délétères de cette inflammation surla taille de la lésion et le pronostic neurologique.Ce projet vise à évaluer l'effet d'un peptide neuroprotecteur, D-JNKI1, inhibiteur de lavoie pro-apoptotique de signalisation intracellulaire c-Jun N-terminal kinase (JNK), surl'inflammation post-ischémique.Nous montrons d'abord que la microglie est largement activée dans toute la région lésée48 h après l'induction d'une ischémie chez la souris. Cependant, malgré l'inhibition dela mort neuronale par D-JNKI1 évaluée à 48 h, nous n'observons de modification ni del'activation de la microglie, ni de son nombre. Ensuite, nous montrons que le cerveaupeut être protégé même s'il y a une augmentation massive de la sécrétion de médiateursinflammatoires dans la circulation systémique très tôt après induction d'un AVCischémique. De plus, nous notons que la sécrétion de molécules inflammatoires dans lecerveau n'est pas différente entre les animaux traités par D-JNKI1 ou une solutionsaline, bien que nous ayons obtenu une neuroprotection significative chez les animauxtraités.En conclusion, nous montrons que l'inhibition de la voie de JNK par D-JNKI1n'influence pas directement l'inflammation post-ischémique. Ceci suggère quel'inhibition de l'inflammation n'est pas forcément nécessaire pour obtenir en hautdegré de neuroprotection du parenchyme lésé après ischémie cérébrale, et que lesmécanismes inflammatoires déclenchés lors d'une ischémie cérébrale ne sont pasforcément délétères pour la récupération du tissu endommagé.SUMMARYAfter cerebral ischemia, glial cells become activated and numerous inflammatory cellsinfiltrate the site of the lesion, secreting a large variety of cytokines and chemokines. Itis controversial whether this brain inflammation is detrimental or beneficial and how itinfluences lesion size and neurological outcome.This project was aimed at critically evaluating whether the neuroprotective peptide DJNKI,an inhibitor of the pro-apopotic c-Jun N-terminal kinase (JNK) pathway,modulates post-ischemic inflammation in animal models of stroke. Specifically, it wasasked whether JNK inhibition prevents microglial activation and the release ofinflammatory mediators.In the first part of this study, we showed that microglia was activated throughout thelesion 48 h after experimental stroke. However, the activation and accumulation ofmicroglia was not reduced by D-JNKI1, despite a significant reduction of the lesionsize. In the second part of this project, we demonstrated that neuroprotection measuredat 48 h occurs even though inflammatory mediators are released in the plasma veryearly after the onset of cerebral ischemia. Furthermore, we found that secretion ofinflammatory mediators in the brain was not different in groups treated with D-JNKI1or not, despite a significant reduction of the lesion size in the treated group.Altogether, we show that inhibition of the JNK pathway using D-JNKI1 does notinfluence directly post-stroke inflammation. Inhibition of inflammation is therefore notnecessarily required for neuroprotection after cerebral ischemia. Thus, post-strokeinflammation might not be detrimental for the tissue recovery.