Mouse models for extracellular matrix diseases

Mutations of a number of genes encoding for extracellular matrix (ECM) proteins in mice have provided new insights regarding their role during development and disease. Many mouse strains have helped to verify the link between mutation and disease in humans, and others have produced unexpected phenotypes and identified new functions for ECM proteins. Finally, some null mutations in ECM genes provide no phenotypic alterations in mice, confronting the scientific community with a new challenge to search for their functions. This review lists all mouse strains with spontaneous and experimentally induced mutations in ECM genes. The phenotypes of these mice are discussed in comparison with the human diseases.     

Assessment of whole body L-leucine oxidation by noninvasive L-[1-13C]leucine breath tests: a reappraisal in patients with maple syrup urine disease, obligate heterozygotes, and healthy subjects

Suitability of a recently proposed noninvasive L-[13C]leucine breath test for assessment of whole body leucine oxidation in maple syrup urine disease (MSUD) was examined. Oral L-[1-13C]leucine loads (38 micromol/kg body weight) were performed in overnight fasted MSUD patients (n = 6, classical form), obligate heterozygote parents (n = 6), and control subjects (n = 10). Three-hour 13CO2 exhalation kinetics were evaluated using curve fitting procedures. Venous blood was obtained in most cases and analyzed for 13C-labeled plasma metabolites. In control subjects, maximal 13CO2 exhalation was reached at tmax = 55 +/- 18 min. Cumulative 13CO2 output at 3 h amounted to 4.7 +/- 0.7 micromol x (kg body weight)(-1). Estimated total 3CO2 exhalation was 7.2 +/- 1.4 micromol x (kg body weight)(-1) (19.0 +/- 3.6% of the dose). Half of this amount was expired at t1/2 = 130 +/- 18 min. The data show a considerable degree of intersubject variability. Intraindividual variability was comparable, however, when checked in two volunteers. In obligate heterozygotes, 13CO2 kinetics were similar to controls (tmax = 35 +/- 8 min, t1/2 = 95 +/- 16 min). Total 13CO2 output [5.7 +/- 1.4 micromol x (kg body weight)(-1)] tended to be in the lower control range. None of the MSUD patients under study exhibited a significant increase in 13CO2 output after load. Maximal increase of label in plasma 4-methyl-2-oxopentanoate, the physiologic precursor of 13CO2, was 16.1 +/- 3.5 MPE in control subjects. In MSUD, label dilution was increased and correlated with the patients' leucine/4-methyl-2-oxopentanoate plasma levels. Considering the generally high variability of 13CO2 output and the unstable substrate pools in MSUD, we discuss the limitations of whole body leucine oxidation measurements by noninvasive approaches.     

A simple test device for electrometers

Electrometers used in dosimetric instruments need to be checked regularly in order to measure and maintain the prescribed dose to the patient. This paper describes the function of a simple but accurate test device for electrometers. A number of electrometers have been tested and compared with calibrations performed by the Swedish Standard Dosimetry Laboratory (SSDL). The device can be used to test current, resistance, voltage and charge measurements. The charge can be conducted to the electrometer in different ways, and the input resistance of the electrometer can also be determined. The calibration factors obtained by the device are in good agreement with results obtained from calibrations at the SSDL.     

Stereo- and regioselective hydroxylation of alpha-ionone by Streptomyces strains

A total of 215 Streptomyces strains were screened for their capacity to regio- and stereoselectively hydroxylate beta- and/or alpha-ionone to the respective 3-hydroxy derivatives. With beta-ionone as the substrate, 15 strains showed little conversion to 4-hydroxy- and none showed conversion to the 3-hydroxy product as desired. Among these 15 Streptomyces strains, S. fradiae Tü 27, S. arenae Tü 495, S. griseus ATCC 13273, S. violaceoniger Tü 38, and S. antibioticus Tü 4 and Tü 46 converted alpha-ionone to 3-hydroxy-alpha-ionone with significantly higher hydroxylation activity compared to that of beta-ionone. Hydroxylation of racemic alpha-ionone [(6R)-(-)/(6S)-(+)] resulted in the exclusive formation of only the two enantiomers (3R,6R)- and (3S, 6S)-hydroxy-alpha-ionone. Thus, the enzymatic hydroxylation of alpha-ionone by the Streptomyces strains tested proceeds with both high regio- and stereoselectivity.     

Cytotoxicity screening of 23 engineered nanomaterials using a test matrix of ten cell lines and three different assays

Background

The most common causes of granulomatous inflammation are persistent pathogens and poorly-degradable irritating materials. A characteristic pathological reaction to intratracheal instillation, pharyngeal aspiration, or inhalation of carbon nanotubes is formation of epithelioid granulomas accompanied by interstitial fibrosis in the lungs. In the mesothelium, a similar response is induced by high aspect ratio nanomaterials, including asbestos fibers, following intraperitoneal injection. This asbestos-like behaviour of some engineered nanomaterials is a concern for their potential adverse health effects in the lungs and mesothelium. We hypothesize that high aspect ratio nanomaterials will induce epithelioid granulomas in nonadherent macrophages in 3D cultures.

Results

Carbon black particles (Printex 90) and crocidolite asbestos fibers were used as well-characterized reference materials and compared with three commercial samples of multiwalled carbon nanotubes (MWCNTs). Doses were identified in 2D and 3D cultures in order to minimize acute toxicity and to reflect realistic occupational exposures in humans and in previous inhalation studies in rodents. Under serum-free conditions, exposure of nonadherent primary murine bone marrow-derived macrophages to 0.5 μg/ml (0.38 μg/cm²) of crocidolite asbestos fibers or MWCNTs, but not carbon black, induced macrophage differentiation into epithelioid cells and formation of stable aggregates with the characteristic morphology of granulomas. Formation of multinucleated giant cells was also induced by asbestos fibers or MWCNTs in this 3D in vitro model. After 7-14 days, macrophages exposed to high aspect ratio nanomaterials co-expressed proinflammatory (M1) as well as profibrotic (M2) phenotypic markers.

Conclusions

Induction of epithelioid granulomas appears to correlate with high aspect ratio and complex 3D structure of carbon nanotubes, not with their iron content or surface area. This model offers a time- and cost-effective platform to evaluate the potential of engineered high aspect ratio nanomaterials, including carbon nanotubes, nanofibers, nanorods and metallic nanowires, to induce granulomas following inhalation.     

Development of a gas layer to mitigate cavitation damage in liquid mercury spallation targets

Establishment of a gas layer between the flowing liquid and container wall is proposed for mitigating the effects of cavitation in mercury spallation targets. Previous work has shown an order of magnitude decrease in damage for a gas layer developed in a stagnant mercury target for an in-beam experiment. This work is aimed at extending these results to the more complex conditions introduced by a flowing mercury target system. A water-loop has been fabricated to provide initial insights on potential gas injection methods into a flowing liquid. An existing full-scale flow loop designed to simulate the Spallation Neutron Source target system will be used to extend these studies to mercury. A parallel analytical effort is being conducted using computational fluid dynamics (CFD) modeling to provide direction to the experimental effort. Some preliminary simulations of gas injection through a single hole have been completed and show behavior of the models that is qualitatively meaningful.     

Integrale Quantifizierung von Schadstoffmassenflüssen im Grundwasser – Konditionierung der numerischen Inversion der Konzentrationsganglinien von Immissionspumpversuchen

A series of integral pumping tests (IPTs) were conducted at a former gasworks site to quantify the contaminant mass flux and average concentration in groundwater along three control planes. The resulting concentration-time series were analysed numerically with the help of the inversion code CSTREAM and a flow and transport model representing the highly heterogeneous aquifer. Since the control planes cover the entire downstream width of the potentially contaminated area, they allow conclusions to be drawn about the current location and spread of the contaminant plume. Previous evaluations of integral pumping tests could calculate three scenarios concerning the spread of the plume around the IPT well: (i) the plume is located to the right of the well, (ii) to the left, or (iii) is distributed symmetrically around it. To create a more realistic picture of the plume position, a series of direct-push monitoring wells were installed along one control plane. The concentrations found in these wells were included in the numerical analysis to condition the inversion results, and resulted in a more pronounced plume centre and fringe, which supports the development of optimised remediation strategies.     

A new minimum blocking condition including subreflector and quadrupod shadow for the design of dual reflector systems

Earlier minimum blocking conditions (MBC's) for paraboloidal reflector antennas formulated by Hannan and Gillitzer did not take into acount the significant blocking contribution of the shadow of the legs supporting the feed or subreflector. A numerical-integration study to determine conditions under which the combined blocking shadow of the subreflector and its supports are minimized is described. Two types of MBC's are found: one which is based on the sidelobe level-more important in radioastronomy (polarization measurements)-and a second type which is related to the aperture efficiency (directivity)-more important for spectroscopy and telecommunications. Both MBC's are compared and it is shown that they are only slightly different from each other.     

Implementation of the Cell Broadband Engine™ in 65 nm SOI Technology Featuring Dual Power Supply SRAM Arrays Supporting 6 GHz at 1.3 V

The 65 nm cell broadband enginetrade (cell BE) is a multi-core SoC, implemented in a high performance SOI technology featuring a separate dual power supply for SRAM arrays to improve stability and performance using an elevated voltage. A new method is shown to analyze the SRAM cell under application conditions which was used to tune the cell for stability, write-ability and performance. An improved write scheme is shown which widens the overall functional window and allows setting the power/performance point of the arrays independently of the surrounding logic. Hardware measurements demonstrate the advantages of the dual power supply under different aspects.     

Phosphodiesterase profile of human B lymphocytes from normal and atopic donors and the effects of PDE inhibition on B cell proliferation

1. CD19+ B lymphocytes were purified from the peripheral blood of normal and atopic subjects to analyse and compare the phosphodiesterase (PDE) activity profile, PDE mRNA expression and the importance of PDE activity for the regulation of B cell function. 2. The majority of cyclic AMP hydrolyzing activity of human B cells was cytosolic PDE4, followed by cytosolic PDE7-like activity; marginal PDE3 activity was found only in the particulate B cell fraction. PDE1, PDE2 and PDE5 activities were not detected. 3. By cDNA-PCR analysis mRNA of the PDE4 subtypes A, B (splice variant PDE4B2) and D were detected. In addition, a weak signal for PDE3A was found. 4. No differences in PDE activities or mRNA expression of PDE subtypes were found in B cells from either normal or atopic subjects. 5. Stimulation of B lymphocytes with the polyclonal stimulus lipopolysaccharide (LPS) induced a proliferative response in a time- and concentration-dependent manner, which was increased in the presence of interleukin-4 (IL-4). PDE4 inhibitors (rolipram, piclamilast) led to an increase in the cellular cyclic AMP concentration and to an augmentation of proliferation, whereas a PDE3 inhibitor (motapizone) was ineffective, which is in accordance with the PDE profile found. The proliferation enhancing effect of the PDE4 inhibitors was partly mimicked by the cyclic AMP analogues dibutyryl (db) cyclic AMP and 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole-3',5'-cyclic monophosphorothioate, Sp-isomer (dcl-cBIMPS), respectively. However, at concentrations exceeding 100 microM db-cyclic AMP suppressed B lymphocyte proliferation, probably as a result of cytotoxicity. Prostaglandin E2 (PGE2, 1 microM) and forskolin (10 microM) did not affect B cell proliferation, even when given in combination with rolipram. 6. Inhibition of protein kinase A (PKA) by differentially acting selective inhibitors (KT 5720, Rp-8-Br-cyclic AMPS) decreased the proliferative response of control cells and reversed the proliferation enhancing effects of rolipram. 7. Importantly, PDE4 activity in LPS/IL-4-activated B lymphocytes decreased by about 50% compared to unstimulated control values. 8. We conclude that an increase in cyclic AMP, mediated by down-regulation of PDE4 activity, is involved in the stimulation of B cell proliferation in response to LPS/IL-4. B cell proliferation in response to a mitogenic stimulus can be further enhanced by pharmacological elevation of cyclic AMP.