Plasmid pFCE4: a new system of Escherichia coli expression-modification vectors

Two versatile expression-modification vectors were obtained by inserting the origin of replication (ori) of phage f1 into the expression vector pOTS. The resulting plasmids produce large amounts of coding or noncoding ssDNA (depending on ori orientation in pFCE4+ and pFCE4-) and excrete it into the medium as virus-like particles following infection with phage f1. These features make them suitable for dideoxy chain termination sequencing, oligonucleotide directed mutagenesis and gene expression without further manipulations. The human IFN alpha-2 gene, lacking the codon for the first amino acid, cysteine, was efficiently expressed by these vectors.     

Reducing Frequent Visits to the Emergency Department: A Systematic Review of Interventions

Objective

The objective of this study was to establish the effectiveness of interventions to reduce frequent emergency department (ED) use among a general adult high ED-use population.

Methods

Systematic review of the literature from 1950-January 2015. Studies were included if they: had a control group (controlled trials or comparative cohort studies), were set in an ED or acute care facility, and examined the impact of an intervention to reduce frequent ED use in a general adult population. Studies reporting non-original data or focused on a specific patient population were excluded. Study design, patient population, intervention, the frequency of ED visits, and costs of frequent ED use and/or interventions were extracted and narratively synthesized.

Results

Among 17 included articles, three intervention categories were identified: case management (n = 12), individualized care plans (n = 3), and information sharing (n = 2). Ten studies examining case management reported reductions in mean (-0.66 to -37) or median (-0.1 to -20) number of ED visits after 12-months; one study reported an increase in mean ED visits (+2.79); and one reported no change. Of these, 6 studies also reported reduced hospital costs. Only 1 study evaluating individualized care plans examined ED utilization and found no change in median ED visits post-intervention. Costs following individualized care plans were also only evaluated in 1 study, which reported savings in hospital costs of $742/patient. Evidence was mixed regarding information sharing: 1 study reported no change in mean ED visits and did not examine costs; whereas the other reported a decrease in mean ED visits (-16.9) and ED cost savings of $15,513/patient.

Conclusions

The impact of all three frequent-user interventions was modest. Case management had the most rigorous evidence base, yielded moderate cost savings, but with variable reductions in ED use. Future studies evaluating non-traditional interventions, tailoring to patient subgroups or socio-cultural contexts, are warranted.     

A novel nitro-oxy substituted analogue of rofecoxib reduces human colon cancer cell growth

Rofecoxib is a specific COX-2 inhibitor able to exert antiproliferative activity against colorectal cancer cells. It was withdrawn from the market after the demonstration of an increased risk of cardiovascular complications after prolonged use. Nevertheless, it remains an interesting compound for laboratory research as an experimental COX-2 inhibitor. In this study, the antiproliferative activity of a novel dinitro-oxy-substituted analogue of rofecoxib (NO-rofe), potentially less cardiotoxic, has been investigated in vitro on human colon cancer cells and compared with the action of the parent drug. Due to the fact that COX-2 inhibition is the main characteristic of coxibs, we performed all experiments in COX-2-overexpressing (HT-29) and COX-2-negative (SW-480) human colon cancer cells, to elucidate whether the observed effects were dependent on COX-2 inhibition. Moreover, experiments were performed in order to evaluate whether COX-2 pharmacological inhibition may affect beta-catenin/E-cadherin signaling pathway. NO-rofe exerted a significant antiproliferative activity on COX-2 positive HT-29 human colon cancer cells, being less effective on the COX-2 negative SW-480 human colon cancer cell line. In particular, the rofecoxib analogue retained similar potencies with respect to COX-2 inhibition but was much more active than rofecoxib in inhibiting the growth of human colon cancer cells in vitro. In addition, this novel compound resulted in the induction of membrane β-catenin/E-cadherin expression, a feature that may significantly contribute to its antiproliferative activity.     

BJ-PI2, A non-hemorrhagic metalloproteinase from Bothrops jararaca snake venom

Background

Envenoming by Bothrops jararaca can result in local pain, edema, hemorrhage and necrosis, partially mediated by snake venom metalloproteinases (SVMPs). Here, we describe the characterization of BJ-PI2, a P-I class SVMP from B. jararaca venom, and its local tissue actions.

Methods

BJ-PI2 was purified by a combination of gel filtration, anion-exchange chromatography and reverse phase HPLC, and identified by mass spectrometry. Clotting and fibrin(ogen)olytic activities were assayed using conventional methods. Hemorrhagic activity and changes in vascular permeability were examined in rat dorsal skin. Myonecrosis and inflammatory activity were examined in mouse gastrocnemius muscle.

Results

BJ-PI2 was a 23.08 kDa single-chain polypeptide. Tryptic fragments showed highest homology with SVMP insularinase A from Bothrops insularis, but also with B. jararaca SVMP bothrojaractivase; less similarity was observed with B. jararaca SVMPs BJ-PI and jararafibrases II and IV. BJ-PI2 did not clot fibrinogen or rat citrated plasma but had α- and β-fibrinogenolytic activity (inhibited by EDTA and 1,10-phenanthroline but not by PMSF) and attenuated coagulation after plasma recalcification. BJ-PI2 had fibrinolytic activity. BJ-PI2 increased the vascular permeability of rat dorsal skin (inhibited by 1,10-phenanthroline). BJ-PI2 was not hemorrhagic or myonecrotic but caused migration of inflammatory cells. In contrast, venom was strongly hemorrhagic and myonecrotic but caused less infiltration of inflammatory cells.

Conclusions

BJ-PI2 is a non-hemorrhagic, non-myonecrotic, non-coagulant P-I class SVMP that may enhance vascular permeability and inflammatory cell migration in vivo.

General significance

BJ-PI2 contributes to enhanced vascular permeability and inflammatory cell migration after envenoming, but not to venom-induced hemorrhage and necrosis.     

Complete Genome Sequence of Mycobacterium massiliense

Mycobacterium massiliense is a rapidly growing bacterium associated with opportunistic infections. The genome of a representative isolate (strain GO 06) recovered from wound samples from patients who underwent arthroscopic or laparoscopic surgery was sequenced. To the best of our knowledge, this is the first announcement of the complete genome sequence of an M. massiliense strain.     

Comparison of the angles and corresponding moments in the knee and hip during restricted and unrestricted squats

ABSTRACT: Lorenzetti, S, Gülay, T, Stoop, M, List, R, Gerber, H, Schellenberg, F, and Stüssi, E. Comparison of the angles and corresponding moments in the knee and hip during restricted and unrestricted squats. J Strength Cond Res 26(10): 2829-2836, 2012-The aim of this study was to compare the angles and corresponding moments in the knee and hip during squats. Twenty subjects performed restricted and unrestricted squats with barbell loads that were 0, ?, and ? their body weight. The experimental setup consisted of a motion capture system and 2 force plates. The moments were calculated using inverse dynamics. During the unrestricted squats, the maximum moments in the knee were significantly higher, and those in the hip were significantly lower than during restricted squats. At the lowest position, the maximum knee flexion angles were approximately 86° for the restricted and approximately 106° for the unrestricted techniques, whereas the maximum hip flexion angle was between 95° and 100°. The higher moments in the hip during restricted squats suggest a higher load of the lower back. Athletes who aim to strengthen their quadriceps should consider unrestricted squats because of the larger knee load and smaller back load.     

Pharmacokinetics of intrathecal transferrin-ricin a chain immunotoxin.

We have studied the pharmacokinetics of an anti-transferrin receptor immunotoxin following intrathecal (i.t.) and intravenous (i.v.) bolus inoculation in healthy rats. After i.t. inoculation of 4.9 microg transferrin-ricin A-chain (Tfn-RTA) we have measured the immunotoxin concentration in the cerebrospinal fluid (CSF), in the brain tissue and in the peripheral blood. After i.v. administration of 4.9 microg Tfn-RTA the concentration of Tfn-RTA immunotoxin was evaluated in the peripheral blood. We found that the clearance of Tfn-RTA from the CSF is rapid (9.1 microLmin(-1)), the immunotoxin then diffuses into the brain tissue and in the peripheral blood where it reaches concentrations below the MTC50 (Minimum Toxin Concentration 50%). The rate of immunotoxin elimination from the peripheral blood following either i.v. or i.t. administration are similar (kel = 0.0021 min(-1) vs. 0.0025 min(-1)). Thus, in the healthy rat the immunotoxin does not accumulate following i.t. inoculation, reaching non toxic concentrations in the brain tissue and in the peripheral blood, whereas in the CSF as well as at the interface CSF/brain tissue the immunotoxin may reach potentially therapeutic concentrations. In conclusion we believe that the i.t. inoculation of an immunotoxin could be considered a potentially useful route of administration in the treatment of leptomeningeal carcinomatosis.