Intima-Media Thickness Does Not Differ between Two Common Carotid Artery Segments in Children

Carotid intima-media thickness (cIMT) is a surrogate marker of early atherosclerotic changes in children. cIMT-studies are hard to compare, due to variations in ultrasound protocols, especially regarding the common carotid artery (CCA) segment measured in relation to the bulb. This study’s purpose was therefore to compare two distinct CCA segments in children, to see if cIMT values differ substantially according to the site of measurement. cIMT was assessed after power calculation in 30 children (15 girls) aged 8–17, using B-Mode ultrasound (5–13 MHz) at two CCA locations. The first measurement was performed over a distance of 1 cm immediately after the bulb (A), the second 1cm proximal the bulb (B) over the same distance of 1cm length. Means of end-diastolic far wall cIMT were compared between measurement A and B. cIMT in 30 participants was 0.51±0.06 mm for measurement A and 0.51±0.05 mm for measurement B. Results did not differ significantly (p = .947) over a distance of 2 cm after the bulb. According to our results, studies measuring CCA IMT within the first 2 cm, either close to the bulb or further proximal, can be compared. This will improve interpretation of data and application of reference values.     

An Estimation Procedure for the Joint Distribution of Spatial Direction and Thickness of Flat Bodies Using Vertical Sections. Part II: An application in soil micromorphology

In soil micromorphology fissures are considered in vertical sections. To get information about the properties of the soil the joint distribution of spatial direction and width of these fissures is of interest. The fissures are mathematically generalized to flat bodies which are defined as stationary weighted surface processes with the weight “thickness”. In a typical point of the surface process suitable, joint parametric distributions of direction and thickness are assumed. The parameters have to be estimated from measurements on vertical sections which are taken from the soil. On these sections only a visible thickness and a visible angle can be observed. The joint distribution of these variables can be expressed by the joint distribution of spatial direction and thickness with the same parameters and in this indirect way the parameters can be estimated. The paper describes how to randomize the vertical section and how to measure the visible variables on the sections. The Chi-Square method is proposed for the parameter estimation. Further it is discussed how to derive good starting values for the numerical procedure. All this is demonstrated in a simulation study using the Bingham-Mardia distribution for the direction and the lognormal distribution for the thickness including a way to correlate the mean thickness and the direction. Finally an application in soil micromorphology is demonstrated for one soil horizon.     

Glioma IL13Rα2 Is Associated with Mesenchymal Signature Gene Expression and Poor Patient Prognosis

A major challenge for successful immunotherapy against glioma is the identification and characterization of validated targets. We have taken a bioinformatics approach towards understanding the biological context of IL-13 receptor α2 (IL13Rα2) expression in brain tumors, and its functional significance for patient survival. Querying multiple gene expression databases, we show that IL13Rα2 expression increases with glioma malignancy grade, and expression for high-grade tumors is bimodal, with approximately 58% of WHO grade IV gliomas over-expressing this receptor. By several measures, IL13Rα2 expression in patient samples and low-passage primary glioma lines most consistently correlates with the expression of signature genes defining mesenchymal subclass tumors and negatively correlates with proneural signature genes as defined by two studies. Positive associations were also noted with proliferative signature genes, whereas no consistent associations were found with either classical or neural signature genes. Probing the potential functional consequences of this mesenchymal association through IPA analysis suggests that IL13Rα2 expression is associated with activation of proinflammatory and immune pathways characteristic of mesenchymal subclass tumors. In addition, survival analyses indicate that IL13Rα2 over-expression is associated with poor patient prognosis, a single gene correlation ranking IL13Rα2 in the top ~1% of total gene expression probes with regard to survival association with WHO IV gliomas. This study better defines the functional consequences of IL13Rα2 expression by demonstrating association with mesenchymal signature gene expression and poor patient prognosis. It thus highlights the utility of IL13Rα2 as a therapeutic target, and helps define patient populations most likely to respond to immunotherapy in present and future clinical trials.     

Transformation of toluene and benzene by mixed methanogenic cultures

The aromatic hydrocarbons toluene and benzene were anaerobically transformed by mixed methanogenic cultures derived from ferulic acid-degrading sewage sludge enrichments. In most experiments, toluene or benzene was the only semicontinuously supplied carbon and energy source in the defined mineral medium. No exogenous electron acceptors other than CO2 were present. The cultures were fed 1.5 to 30 mM unlabeled or 14C-labeled aromatic substrates (ring-labeled toluene and benzene or methyl-labeled toluene). Gas production from unlabeled substrates and 14C activity distribution in products from the labeled substrates were monitored over a period of 60 days. At least 50% of the substrates were converted to CO2 and methane (greater than 60%). A high percentage of 14CO2 was recovered from the methyl group-labeled toluene, suggesting nearly complete conversion of the methyl group to CO2 and not to methane. However, a low percentage of 14CO2 was produced from ring-labeled toluene or from benzene, indicating incomplete conversion of the ring carbon to CO2. Anaerobic transformation pathways for unlabeled toluene and benzene were studied with the help of gas chromatography-mass spectrometry. The intermediates detected are consistent with both toluene and benzene degradation via initial oxidation by ring hydroxylation or methyl oxidation (toluene), which would result in the production of phenol, cresols, or aromatic alcohol. Additional reactions, such as demethylation and ring reduction, are also possible. Tentative transformation sequences based upon the intermediates detected are discussed.     

Macrophages from endotoxin-hyporesponsive (Lpsd) C3H/HeJ mice are permissive for vesicular stomatitis virus because of reduced levels of endogenous interferon: possible mechanism for natural resistance to virus infection.

The C3H/HeJ mouse strain bears an autosomal gene defect, Lpsd, which results in a greatly diminished capacity to respond to endotoxin, the ubiquitous lipopolysaccharide derived from the cell walls of gram-negative bacteria. These mice also exhibit greater susceptibility to a variety of viral and bacterial infections than syngeneic, fully lipopolysaccharide-responsive (Lpsn) mouse strains and possess macrophages with defects in differentiation which are reversed by treatment with exogenous interferon (IFN). To test directly the hypothesis that C3H/HeJ macrophages are deficient in endogenous IFN levels, macrophages from C3H/HeJ (Lpsd) and C3H/OuJ (Lpsn) mice were compared for sensitivity to vesicular stomatitis virus. At a multiplicity of infection of 0.1, C3H/OuJ macrophages were completely refractory to infection, whereas C3H/HeJ macrophages were permissive for replication, and infection resulted in 100% cytopathic effect. These findings were confirmed with a second inbred Lpsn and Lpsd strain pair. Levels of 2',5'-oligoadenylate synthetase were significantly higher in Lpsn cells. C3H/HeJ macrophages, derived from bone marrow precursors under the influence of macrophage colony-stimulating factor, shown previously to induce IFN in macrophages, were as refractory as C3H/OuJ macrophages. Exposure of nonpermissive macrophages to anti-IFN-alpha/beta antibody prior to infection rendered cells permissive. Our findings suggest that endotoxin provides a primary stimulus for the maintenance of normal macrophage differentiation and innate resistance via the induction of endogenous IFN by macrophages.     

Spontaneous and cyclophosphamide-induced sister-chromatid exchanges in diploid and endoreduplicated tetraploid metaphases of preimplantation mouse embryos

Endoreduplicated tetraploid metaphases could for the first time be induced in preimplantation mouse embryos by culture in the suboptimum medium MEM. In such endomitoses sister-chromatid exchange (SCE) frequency was approximately the same during the first and the second cell cycle. However, when morulae and blastocysts were cultured in the presence of cyclophosphamide metabolites SCE frequency was increased predominantly during the second cell cycle. Compared to diploid metaphases a decreased SCE frequency was found under both conditions of endomitoses induction, which may be related to DNA-repair processes.     

The secondary structure of bacteriorhodopsin determined by Raman and circular dichroism spectroscopy

The secondary structure of bacterio-opsin (BO), the retinal free protein-component of bacteriorhodopsin (BR), has been determined by Raman spectroscopy. Additional circular dichroism (CD) measurements have revealed only negligible conformational differences between BO in apomembranes and BR in purple membranes. Therefore, the secondary structure of BR was derived from the Raman data of BO. The protein conformation was determined to consist of 72-82% helices, 2-11% beta-strands, and 11-17% beta-turns. Only about half of the helical structures correspond to alpha 1-helices, the other half possess non-alpha 1-helical structures. According to the analysis of the Raman data, the derived secondary structure of BR was obtained with high reliability for all structure classes which can be distinguished by this method within the given uncertainty range. This is a remarkable difference from recently published secondary structural data derived from CD measurements where the helix content was reported to be between 50 and 80%. The inherent experimental and methodological uncertainties of the CD-technique leading to such a range of variation are critically discussed in comparison to the method of Raman spectroscopy. The combined application of Raman and CD spectroscopy, as performed here, is demonstrated to be a substantial improvement in the secondary structure determination of retinal-containing membrane proteins. On the basis of our results, some of the recently proposed structural models of BR with a beta-strand content of more than 11% can be ruled out.     

Interleukin 2 and concanavalin A stimulate interferon-gamma production in a murine cytolytic T cell clone by different pathways

We identified a variant murine cytolytic T lymphocyte (CTL) clone which, in contrast to the parent clone and all other murine T cell populations tested, was found to have acquired spontaneously the ability to produce interferon-gamma (IFN-gamma) in response to recombinant interleukin 2 (rIL-2). IFN-gamma production in response to concanavalin A (Con A), which was characteristic of all T cell populations tested, was preserved in this variant. The IFN produced by the variant in response to either stimulus was active in both a macrophage-activating factor assay and an anti-viral assay. Both activities induced by either stimulus could be blocked by monoclonal anti-IFN-gamma antibodies. Upon Northern blot analysis using an IFN-gamma-specific cDNA probe, the IFN-gamma RNA isolated from variant cells stimulated with Con A or IL-2 were found to migrate equivalently. The unusual pattern of responsiveness in this variant CTL was exploited to compare the mechanisms involved in induction of IFN-gamma production by Con A or IL-2. Striking differences were observed. Unlike IFN-gamma production induced by Con A, IFN-gamma production induced by IL-2 was not accompanied by an elevation of intracellular Ca2+ levels, did not require physiologic extracellular Ca2+ levels, and was not inhibited by the immunosuppressive agent cyclosporin A. Thus, in this variant CTL clone, conditions that have ordinarily been associated in an obligate manner with lymphokine gene expression were found instead to be related to the specific mode of stimulation.     

Species specificity of anti-acetylcholine receptor antibodies elicited by synthetic peptides

Two peptides corresponding to amino acid residues 351-368 of the alpha-subunits of Torpedo and human acetylcholine receptor (AChR) were synthesized. These peptides contain a segment (residues 355-364) which displays the greatest variability in amino acid sequence between the two species. Antibodies elicited against the two peptides cross-reacted with the respective native AChRs and were shown to be species specific by radioimmunoassay, immunoblotting, and immunofluorescence microscopy. Thus, antibodies against the Torpedo peptide cross-reacted with Torpedo AChR but did not bind to mammalian or chicken AChR. Antibodies against the human peptide proved to be specific probes for mammalian muscle AChR. They cross-reacted with mammalian AChR (human, calf, mouse, and rat) but not with Torpedo or chicken AChR. These antibodies were also shown to react preferentially with the extrajunctional form of muscle AChR, as compared to their reactivity with junctional muscle AChR. In immunofluorescence experiments, the anti-human peptide antibody stained AChR aggregates in sectioned or ethanol-permeabilized rat and mouse myotubes grown in culture but did not stain living myotubes. This indicates that the sequence 351-368 of the alpha-subunit of mammalian AChR is on the cytoplasmic face of muscle cell membranes, as predicted theoretically.