An in vivo method for measuring turbulence in mechanical prosthesis leakage jets

This work introduces a method for the in vivo measurement and analysis of turbulence within the leakage of a mechanical heart valve. Several analysis techniques were applied to ultrasound measurements acquired within the atrium of a pig, and error associated with these techniques was analyzed. The technique chosen applies cyclic averaging to mean and maximum velocity measurements within small, normalized phase windows to calculate Reynolds normal stresses in the direction of the ultrasound beam. Maximum shear stresses are estimated from these normal stresses using an analytical technique. The stresses observed were smaller than those reported from previous in vitro simulations.     

'One bead two compound libraries' for detecting chemical and biochemical conversions

When combinatorial chemistry was introduced 13 years ago, the expectations were high for the delivery of results, particularly in the pharmaceutical industry. However, combinatorial chemistry was implemented independently of the application for which the products were going to be used. Resins developed only for efficient solid-phase synthesis were used and products were employed in existing assays developed for traditional solution studies. There was almost no assay or technology development and the use of real combinatorial methods soon had to give way to high-throughput synthesis and traditional screening. However, during recent years more sophisticated resins and assay techniques have been developed that may result in a second and more successful implementation of real integrated combinatorial chemistry. The first in this line of new developments is the 'one bead two compound' assay, in which the resin bead in addition to a combinatorial library member contains a reporter compound that can act as a beacon to monitor the activity of the library member. This powerful concept can be generally applied in all fields of combinatorial chemistry including drug, catalysts and material development.     

Acidosis has opposite effects on neuronal survival during hypoxia and reoxygenation

To study the effect of extracellular acidosis on apoptosis and necrosis during ischemia and reoxygenation, we exposed human post-mitotic NT2-N neurones to oxygen and glucose deprivation (OGD) followed by reoxygenation. In some experiments, pH of the cell medium was lowered to 5.9 during either OGD or reoxygenation or both. Staurosporine, used as a positive control for apoptosis, caused Poly(ADP-ribose)-polymerase (PARP) cleavage and nuclear fragmentation, but no PARP cleavage and little fragmentation were seen after OGD. Low molecular weight DNA fragments were found after staurosporine treatment, but not after OGD. No protective effect of caspase inhibitors was seen after 3 h of OGD and 21 h of reoxygenation, but after 45 h of reoxygenation caspase inhibition induced a modest improvement in 3-(4,5-dimethylthiazol-2-yl)2,5-diphenyltetrazolium bromide (MTT) cleavage. While acidosis during OGD accompanied by neutral medium during reoxygenation protected the neurones (MTT: 228 +/- 117% of neutral medium, p < 0.001), acidosis during reoxygenation only was detrimental (MTT: 38 +/- 25%, p < 0.01). We conclude that apoptotic mechanisms play a minor role after OGD in NT2-N neurones. The effect of acidosis on neuronal survival depends on the timing of acidosis, as acidosis was protective during OGD and detrimental during reoxygenation.     

The role of phosphatidylinositol 3-kinase in neural cell adhesion molecule-mediated neuronal differentiation and survival

The neural cell adhesion molecule, NCAM, is known to stimulate neurite outgrowth from primary neurones and PC12 cells presumably through signalling pathways involving the fibroblast growth factor receptor (FGFR), protein kinase A (PKA), protein kinase C (PKC), the Ras-mitogen activated protein kinase (MAPK) pathway and an increase in intracellular Ca2+ levels. Stimulation of neurones with the synthetic NCAM-ligand, C3, induces neurite outgrowth through signalling pathways similar to the pathways activated through physiological, homophilic NCAM-stimulation. We present here data indicating that phosphatidylinositol 3-kinase (PI3K) is required for NCAM-mediated neurite outgrowth from PC12-E2 cells and from cerebellar and dopaminergic neurones in primary culture, and that the thr/ser kinase Akt/protein kinase B (PKB) is phosphorylated downstream of PI3K after stimulation with C3. Moreover, we present data indicating a survival-promoting effect of NCAM-stimulation by C3 on cerebellar and dopaminergic neurones induced to undergo apoptosis. This protective effect of C3 included an inhibition of both DNA-fragmentation and caspase-3 activation. The survival-promoting effect of NCAM-stimulation was also shown to be dependent on PI3K.     

Predicted molecular structure of the mammalian cell entry protein Mce1A of Mycobacterium tuberculosis

The proposed role of the mammalian cell entry protein 1A (Mce1A) of Mycobacterium tuberculosis is to facilitate invasion of host cells. The structure of Mce1A was modelled on the basis of the crystal structure of Colicin N of Escherichia coli by fold prediction and threading. Mce1A, as the model predicts, is an alpha/beta protein consisting of two major (alpha and beta) domains, connected by a long alpha helix. The model further revealed that the protein contains 12 helices, 9 strands, and 1 turn. The final model of Mce1A was verified through the program VERIFY 3D and more than 90% of the residues were in the favourable region. A mouse monoclonal antibody, TB1-5 76C, is directed to an epitope within a 60-mer peptide that has been shown to promote uptake of bacteria in mammalian cells. We show here that the epitope could be narrowed down to a core of 4 amino acids, TPKD. Upstream flanking residues, KRR also contributed to binding. Mce2A does not promote uptake in mammalian cells and sequence comparison of Mce1A and Mce2A indicates that the epitope mediates uptake. The epitope was located at the surface of the Mce1A model at the distal beta strand-loop region in the beta domain. The localization of this epitope in the model confirms its potential role in promoting uptake of M. tuberculosis in host cells.     

Stem cells from the adult human brain develop into functional neurons in culture

Recent research communications indicate that the adult human brain contains undifferentiated, multipotent precursors or neural stem cells. It is not known, however, whether these cells can develop into fully functional neurons. We cultured cells from the adult human ventricular wall as neurospheres and passed them at the individual cell level to secondary neurospheres. Following dissociation and plating, the cells developed the antigen profile of the three main cell types in the brain (GFAP, astrocytes; O2, oligodendrocytes; and beta-III-tubulin/NeuN, neurons). More importantly, the cells developed the electrophysiological profiles of neurons and glia. Over a period of 3 weeks, neuron-like cells went through the same phases as neurons do during development in vivo, including up-regulation of inward Na+ -currents, drop in input resistance, shortening of the action potential, and hyperpolarization of the cell membrane. The cells developed overshooting action potentials with a mature configuration. Recordings in voltage-clamp mode displayed both the fast inactivating TTX-sensitive sodium current (INa) underlying the rising phase of the action potential and the two potassium currents terminating the action potential in mature neurons (IA and IK, sensitive to 4-AP and TEA, respectively). We have thus demonstrated that the human ventricular wall contains multipotent cells that can differentiate into functionally mature neurons.     

The external promoter in the guinea pig 5S rRNA gene is different from the rodent promoter

The guinea pig has about 100 copies of the 5S rRNA gene per haploid genome and they are present in 2.1 kb tandem repeats. Three bona fide 5S rRNA genes and four pseudo genes were sequenced. The conserved external promoter (D box) found in rodents and primates is only partially present in the guinea pig. The "D box like" sequence in guinea pig only has eight of the 12 nucleotides in the conserved D box. The results are in accordance with investigations showing that the guinea pig is not a rodent. Conserved sequences in the non-transcribed spacer can therefore be useful in phylogenetic studies.     

Bone bank service in Odense, Denmark. Audit of the first ten years with bone banking at the Department of Orthopaedics, Odense University Hospital

There has been an increase in the demand for allograft bone in recentyears. The Odense University Hospital bone bank has been in function since1990,and this paper outlines our results during the 10 year period 1990–1999.Potential donors were screened by contemporary banking techniques which includea social history, donor serum tests for HIV, hepatitis B and C, and graftmicrobiology. The bones were stored at –80 °C. No typeofsecondary sterilisation was made. 423 femoral heads were approved and donatedto300 patients,1–6 heads/operation. The allografts have been used mainly toreconstruct defects at revision hip arthroplasty (34%), and for fracturesurgery(24%). 7 % of all transplanted patients were reoperated because of infection.Inthe hip revision group the infection rate was 4 %. There were no cases ofdisease transmission. During the 10 year period there was a change in theclinical use of the allografts. In the first years the allografts were mainlyused for spinal fusion surgery, but today the majority are used in hip revisionand fracture surgery. The clinical results correspond to those reported inlarger international series.     

Cardiac function and metabolism in Type 2 diabetic mice after treatment with BM 17.0744, a novel PPAR-alpha activator

Hearts from diabetic db/db mice, a model of Type 2 diabetes, exhibit left ventricular failure and altered metabolism of exogenous substrates. Peroxisome proliferator-activated receptor-alpha (PPAR-alpha) ligands reduce plasma lipid and glucose concentrations and improve insulin sensitivity in db/db mice. Consequently, the effect of 4- to 5-wk treatment of db/db mice with a novel PPAR-alpha ligand (BM 17.0744; 25-38 mg x kg(-1) x day(-1)), commencing at 8 wk of age, on ex vivo cardiac function and metabolism was determined. Elevated plasma concentrations of glucose, fatty acids, and triacylglycerol (34.0 +/- 3.6, 2.0 +/- 0.4, and 0.9 +/- 0.1 mM, respectively) were reduced to normal after treatment with BM 17.0744 (10.8 +/- 0.6, 1.1 +/- 0.1, and 0.6 +/- 0.1 mM). Plasma insulin was also reduced significantly in treated compared with untreated db/db mice. Chronic treatment of db/db mice with the PPAR-alpha agonist resulted in a 50% reduction in rates of fatty acid oxidation, with a concomitant increase in glycolysis (1.7-fold) and glucose oxidation (2.3- fold). Correction of the diabetes-induced abnormalities in systemic and cardiac metabolism after BM 17.0744 treatment did not, however, improve left ventricular contractile function.