Preventing vasospasm improves outcome after aneurysmal subarachnoid hemorrhage: rationale and design of CONSCIOUS-2 and CONSCIOUS-3 trials

Cerebral vasospasm after aneurysmal subarachnoid hemorrhage (aSAH) is a frequent but unpredictable complication associated with poor outcome. Current vasospasm therapies are suboptimal; new therapies are needed. Clazosentan, an endothelin receptor antagonist, has shown promise in phase 2 studies, and two randomized, double-blind, placebo-controlled phase 3 trials (CONSCIOUS-2 and CONSCIOUS-3) are underway to further investigate its impact on vasospasm-related outcome after aSAH. Here, we describe the design of these studies, which was challenging with respect to defining endpoints and standardizing endpoint interpretation and patient care. Main inclusion criteria are: age 18-75 years; SAH due to ruptured saccular aneurysm secured by surgical clipping (CONSCIOUS-2) or endovascular coiling (CONSCIOUS-3); substantial subarachnoid clot; and World Federation of Neurosurgical Societies grades I-IV prior to aneurysm-securing procedure. In CONSCIOUS-2, patients are randomized 2:1 to clazosentan (5 mg/h) or placebo. In CONSCIOUS-3, patients are randomized 1:1:1 to clazosentan 5, 15 mg/h, or placebo. Treatment is initiated within 56 h of aSAH and continued until 14 days after aSAH. Primary endpoint is a composite of mortality and vasospasm-related morbidity within 6 weeks of aSAH (all-cause mortality, vasospasm-related new cerebral infarction, vasospasm-related delayed ischemic neurological deficit, neurological signs or symptoms in the presence of angiographic vasospasm leading to rescue therapy initiation). Main secondary endpoint is extended Glasgow Outcome Scale at week 12. A critical events committee assesses all data centrally to ensure consistency in interpretation, and patient management guidelines are used to standardize care. Results are expected at the end of 2010 and 2011 for CONSCIOUS-2 and CONSCIOUS-3, respectively.

Morphological and functional 3-Tesla magnetic resonance imaging of saphenous vein coronary artery bypass grafts

Evaluation of a novel non-invasive tool for postoperative follow-up of patients postelective saphenous vein coronary artery bypass graft (CABG) was performed. Ten patients were included. Their bypass grafts supplied the right coronary artery (7), marginal branches (1), diagonal branches (2), and the circumflex artery (n=1). Each bypass was examined intraoperatively using Doppler flow measurement. Patients were examined with a 3-Tesla magnetic resonance imaging (MRI) scanner (MAGNETOM Verio, Siemens, Erlangen, Germany) within one week postsurgery using MR-angiography with an intravasal contrast agent and velocity encoded phase-contrast flow measurements. Intraoperative Doppler flow measurements revealed regular flow patterns in all vascular territories supplied. The median intraoperative flow rate was 50 ml/min with an inter-quartile range (IQR) of 42-70 ml/min. The clinical postoperative course was uneventful. MRI showed all grafts to be patent. The median postoperative flow rate was 50 ml/min (IQR: 32-65 ml/min). MRI flow rates agreed well with intraoperative Doppler flow measurements (mean difference: -2.8±20.1 ml/min). This initial study demonstrates that 3-Tesla MRI flow measurements correlated well with Doppler thus reconfirming the graft patency postCABG. Further refinement and broader application of this technique may facilitate follow-up postCABG potentially replacing empiric clinical judgment by reliable non-invasive imaging.

Emergence of four cases of KPC-2 and KPC-3-carrying Klebsiella pneumoniae introduced to Switzerland, 2009-10

We report four epidemiologically unrelated cases of KPC-carrying Klebsiella pneumoniae identified in Switzerland between May 2009 and November 2010. Three cases were transferred from Italy (two KPC-3, one KPC-2) and one from Greece (KPC-2). Resistance to colistin and doxycycline emerged in one KPC-3-carrying K. pneumoniae strain during therapy. These results demonstrate ongoing dissemination of KPC throughout Europe. Rapid and reliable identification of KPC and implementation of control measures is essential to limit spread.

Inbreeding Alters Activities of the Stress-Related Enzymes Chitinases and β-1,3-Glucanases

Pathogenesis-related proteins, chitinases (CHT) and β-1,3-glucanases (GLU), are stress proteins up-regulated as response to extrinsic environmental stress in plants. It is unknown whether these PR proteins are also influenced by inbreeding, which has been suggested to constitute intrinsic genetic stress, and which is also known to affect the ability of plants to cope with environmental stress. We investigated activities of CHT and GLU in response to inbreeding in plants from 13 Ragged Robin (Lychnis flos-cuculi) populations. We also studied whether activities of these enzymes were associated with levels of herbivore damage and pathogen infection in the populations from which the plants originated. We found an increase in pathogenesis-related protein activity in inbred plants from five out of the 13 investigated populations, which suggests that these proteins may play a role in how plants respond to intrinsic genetic stress brought about by inbreeding in some populations depending on the allele frequencies of loci affecting the expression of CHT and the past levels of inbreeding. More importantly, we found that CHT activities were higher in plants from populations with higher levels of herbivore or pathogen damage, but inbreeding reduced CHT activity in these populations disrupting the increased activities of this resistance-related enzyme in populations where high resistance is beneficial. These results provide novel information on the effects of plant inbreeding on plant–enemy interactions on a biochemical level.

14-3-3- and II/3-10-gene expression as molecular markers to address viability and growth activity of Echinococcus multilocularis metacestodes

The present study aimed to search for and characterize parasite molecules, whose expression levels correlate with the viability and growth activity of Echinococcus multilocularis metacestodes. We focused on the expression profiles of 2 parasite-derived genes, 14-3-3 and II/3-10, as putative molecular markers for viability and growth activity of the larval parasite. In experiments in vivo, gene expression levels of 14-3-3 and II/3-10 were relatively quantified by real-time reverse transcription-PCR using a housekeeping gene, beta-actin, as a reference reaction. All three reactions were compared with growth activity of the parasite developing in permissive nu/nu and in non-permissive wild type BALB/c mice. At 2 months p.i., the transcription level of 14-3-3 was significantly higher in parasites actively proliferating in nu/nu mice compared to parasites moderately growing in wild type mice. Immunoblotting experiments confirmed at the protein level that 14-3-3 was over-expressed in parasites derived from nu/nu mice at 2 months p.i. In vitro treatment of E. multilocularis with an anti-echinococcal drug nitazoxanide resulted in a significant decrease of both 14-3-3 and II/3-10 transcription levels found after 8 days of treatment, which correlated with the kinetics of a housekeeping gene, beta-actin. The conclusion is that 14-3-3, combined with II/3-10, exhibits good potential as a molecular marker to assess viability and growth activity of the parasite.

Use of straight and curved 3-dimensional titanium miniplates for fracture fixation at the mandibular angle

PURPOSE: The aim of this follow-up study was to evaluate the clinical usefulness of a new type of 3-dimensional (3D) miniplate for open reduction and monocortical fixation of mandibular angle fractures. PATIENTS AND METHODS: In 20 consecutive patients, noncomminuted mandibular angle fractures were treated with open reduction and fixation using a 2 mm 3D miniplate system in a transoral approach. All patients were systematically monitored until 6 months postoperatively. Among the outcome parameters recorded were infection, hardware failure, wound dehiscence, and sensory disturbance of the inferior alveolar nerve. RESULTS: The mean operation time from incision to wound closure was 65 minutes. Two patients had a mucosal wound dehiscence with no consequences. None developed an infection requiring a plate removal. All but 2 patients had normal sensory function 3 months after surgery. Plate fracture occurred in one patient in whom a preceding surgical removal of the third molar had been the reason for the mandibular fracture. In the absence of clinical symptoms, the patient declined plate removal. On final follow-up, fracture healing was considered clinically complete in all patients. CONCLUSIONS: The 3D plating system described here is suitable for fixation of simple mandibular angle fractures and is an easy-to-use alternative to conventional miniplates. The system may be contraindicated in patients in whom insufficient interfragmentary bone contact causes minor stability of the fracture.

Novel functional profiling approach combining reverse phase protein microarrays and human 3-D ex vivo tissue cultures: expression of apoptosis-related proteins in human colon cancer

Cancer is caused by a complex pattern of molecular perturbations. To understand the biology of cancer, it is thus important to look at the activation state of key proteins and signaling networks. The limited amount of available sample material from patients and the complexity of protein expression patterns make the use of traditional protein analysis methods particularly difficult. In addition, the only approach that is currently available for performing functional studies is the use of serial biopsies, which is limited by ethical constraints and patient acceptance. The goal of this work was to establish a 3-D ex vivo culture technique in combination with reverse-phase protein microarrays (RPPM) as a novel experimental tool for use in cancer research. The RPPM platform allows the parallel profiling of large numbers of protein analytes to determine their relative abundance and activation level. Cancer tissue and the respective corresponding normal tissue controls from patients with colorectal cancer were cultured ex vivo. At various time points, the cultured samples were processed into lysates and analyzed on RPPM to assess the expression of carcinoembryonic antigen (CEA) and 24 proteins involved in the regulation of apoptosis. The methodology displayed good robustness and low system noise. As a proof of concept, CEA expression was significantly higher in tumor compared with normal tissue (p<0.0001). The caspase 9 expression signal was lower in tumor tissue than in normal tissue (p<0.001). Cleaved Caspase 8 (p=0.014), Bad (p=0.007), Bim (p=0.007), p73 (p=0.005), PARP (p<0.001), and cleaved PARP (p=0.007) were differentially expressed in normal liver and normal colon tissue. We demonstrate here the feasibility of using RPPM technology with 3-D ex vivo cultured samples. This approach is useful for investigating complex patterns of protein expression and modification over time. It should allow functional proteomics in patient samples with various applications such as pharmacodynamic analyses in drug development.