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PubMed2003

Oxidative stress as a therapeutic target in diabetes: revisiting the controversy.

Oxidative stress has been repetitively shown to be a hallmark of many diseases linked with metabolic or vascular disorders. Therefore diabetes represents an ideal candidate for studying the consequences of oxidative stress and its treatment. Indeed diabetes constitutes a multiple source of free radicals, starting very early in the disease process and worsening over the course of disease. In view of the typical characteristics of diabetes, oxidative stress is expected to have a double impact, on both metabolic and vascular functions. It is therefore particularly disappointing to note the dramatic failure of clinical trials with antioxidants, although it must be pointed out that such studies have not been performed with only diabetic patients. This review describes the many different aspects of oxidative stress in diabetes and proposes possible explanations for the apparent lack of efficacy of antioxidant treatments in patients. Some verifications seem warranted before a definitive conclusion can be drawn about the validity of this therapeutic concept.

Antioxidants↗

PubMed2006

Identification of sites of mannose 6-phosphorylation on lysosomal proteins.

Most newly synthesized soluble lysosomal proteins contain mannose 6-phosphate (Man-6-P), a specific carbohydrate modification that is recognized by Man-6-P receptors (MPRs) that direct targeting to the lysosome. A number of proteomic studies have focused on lysosomal proteins, exploiting the fact that Man-6-P-containing forms can be purified by affinity chromatography on immobilized MPRs. These studies have identified many known lysosomal proteins as well as many proteins not previously classified as lysosomal. The latter are of considerable biological interest with potential implications for lysosomal function and as candidates for lysosomal storage diseases of unknown etiology. However, a significant problem in interpreting the biological relevance of such proteins has been in distinguishing true Man-6-P glycoproteins from simple contaminants and from proteins associated with true Man-6-P glycoproteins (e.g. protease inhibitors and lectins). In this report, we describe a mass spectrometric approach to the verification of Man-6-phosphorylation based upon LC-MS of MPR-purified proteolytic glycopeptides. This provided a useful tool in validating novel MPR-purified proteins as true Man-6-P glycoproteins and also allowed identification of low abundance components not observed in the analysis of the total Man-6-P glycoprotein mixture. In addition, this approach allowed the global mapping of 99 Man-6-phosphorylation sites from 44 known lysosomal proteins purified from mouse and human brain. This information is likely to provide useful insights into protein determinants for this modification and may be of significant value in protein engineering approaches designed to optimize protein delivery to the lysosome in therapeutic applications such as gene and enzyme replacement therapies.

Amino Acid Sequence↗

PubMed2005

In silico and wet-bench identification of nuclear matrix attachment regions.

Chromatin loops are tethered at discrete regions that are approx 100-1000 bp in length. These regions of attachment serve as specific sequence landmarks, anchoring the DNA to the fibers of the chromosomal scaffold. It has been estimated that our genome contains 70,000 nuclear matrix attachment sites that serve as a dynamic nuclear organizer in both the interphase and metaphase cell. Approximately 30,000-40,000 matrix attachment regions (MARs) serve as origins of replication. MARs can also be associated with chromosomal segments densely populated with transcription factor-binding sites. This may facilitate transcription that is initiated within the region of the chromosome coincident with the surface of the nuclear matrix. Assuming an average somatic loop size of 100 kb, it is reasonable to propose that each cell utilizes 30,000 MARs to anchor each of the approx 20,000 active genic domains. This is sufficient to encompass the 30,000 functional genes in our genome that exist as members of single or multigenic families, each constituting a single chromatin domain. With the sequencing phase of various genome projects complete, in silico tools are being developed to identify the long-range control elements that modulate gene expression. This information is necessary to specifically target the time-intensive wet-bench verification and expression experiments that will provide a unified understanding of gene regulation. In this chapter we review some of the in silico strategies that are currently available and a new in vivo method based on the real-time polymerase chain reaction, to assess regions of matrix association.

Actins↗

PubMed1983

Quality assurance operations: good clinical practices.

The pharmaceutical industry sponsor bears the ultimate responsibility for the verification of all clinical data submitted to the FDA in support of a New Drug Application (NDA). The author provides an overview of a systematic approach to providing quality assurance for all clinical data collected in the course of human trials with research compounds. This systemic approach includes such features as: Standard Operating Procedures Manuals, the development of a quality assurance function within the organization, detailed documentation for all monitoring activities, careful assessment of all clinical research data collected, and an emphasis on preventive measures that can be introduced to assure the quality of subsequent research data.

Clinical Trials as Topic↗

PubMed1989

[The structure and function of an automated system for predicting neonatal body weight (the Sinodeta P/1)].

The structure and function of an automated nosometric system, created by the authors, for prognosis of the risk of delivery of a child with low weight is described. The system is designed for health services, which give direction to parturient (public health sectors) and obstetric wards without technique and specialists for management of such children. The summarized extract is made on 883 cases, but verification--on 463 cases. The total effectiveness of the system is 82.1% against 66.25% of exact results during intuitive prognosis.

Algorithms↗

PubMed1995

Echo-planar MR cerebral blood volume mapping of gliomas. Clinical utility.

Neovascularization is a common phenomenon in gliomas. MR imaging cerebral blood volume (CBV) mapping utilizes ultrasfast echo-planar imaging and simultaneous use of gadolinium-based contrast material. To determine the utility of MR CBV mapping in the clinical evaluation of gliomas, we followed 15 patients with serial studies. This technique provided functional information that was not evident with conventional CT or MR imaging. Low-grade tumors demonstrated homogeneously low CBV, while high-grade tumors often showed areas of both high and low CBV. The maximum tumor CBV/white matter ratio was compared between low- (n = 3) and high-grade gliomas (n = 5) in patients without previous treatment and with histologic verification (n = 8) and was significantly higher in high-grade gliomas (p < 0.01). High CBV foci in nonenhancing tumor areas were present in 2 cases. The distinction between radiation necrosis and active tumor could be made correctly in 3 of 4 cases. The information provided by MR CBV mapping has the potential to be an adjunct in the clinical care of glioma patients.

Adolescent↗

PubMed1994

Methods for fully automated verification of patient set-up in external beam radiotherapy with polygon shaped fields.

Methods have been developed to perform fully automated comparisons of the radiotherapy simulator and portal images in the most common cases where the field edges consist of straight lines. The field defining wires or rods in the simulator image, and the edges of the portal image, are localized by means of a truncated Radon transform. Edges are enhanced with the use of a generalized Laplacian operator. From the detected field outlines, a geometrical mapping function is determined that rotates, scales, and translates one image with respect to the other. A subsequent match of field shapes is executed. If satisfactory agreement is established, the anatomical structures of the simulator and portal images are compared by correlating a new pair of images that only contain the intensity ridges representing bone outlines. The correlation determines the rotation and translation that must be applied to align the anatomical structures. With the simulator image already processed, the remaining automatic processing takes approximately 2 min on a SUN Sparcstation 2. By use of a priori knowledge of the fields, the computation time needed after acquisition of the portal image may be reduced to about 40 s for 512 x 512 images and about 12 s for 256 x 256 images.

Algorithms↗

PubMed2000

A variable resolution x-ray detector for computed tomography: I. Theoretical basis and experimental verification.

A computed tomography imaging technique called variable resolution x-ray (VRX) detection provides detector resolution ranging from that of clinical body scanning to that of microscopy (1 cy/mm to 100 cy/mm). The VRX detection technique is based on a new principle denoted as "projective compression" that allows the detector resolution element to scale proportionally to the image field size. Two classes of VRX detector geometry are considered. Theoretical aspects related to x-ray physics and data sampling are presented. Measured resolution parameters (line-spread function and modulation-transfer function) are presented and discussed. A VRX image that resolves a pair of 50 micron tungsten hairs spaced 30 microns apart is shown.

Algorithms↗

PubMed2005

Spatiotemporal image correlation spectroscopy (STICS) theory, verification, and application to protein velocity mapping in living CHO cells.

We introduce a new extension of image correlation spectroscopy (ICS) and image cross-correlation spectroscopy (ICCS) that relies on complete analysis of both the temporal and spatial correlation lags for intensity fluctuations from a laser-scanning microscopy image series. This new approach allows measurement of both diffusion coefficients and velocity vectors (magnitude and direction) for fluorescently labeled membrane proteins in living cells through monitoring of the time evolution of the full space-time correlation function. By using filtering in Fourier space to remove frequencies associated with immobile components, we are able to measure the protein transport even in the presence of a large fraction (>90%) of immobile species. We present the background theory, computer simulations, and analysis of measurements on fluorescent microspheres to demonstrate proof of principle, capabilities, and limitations of the method. We demonstrate mapping of flow vectors for mixed samples containing fluorescent microspheres with different emission wavelengths using space time image cross-correlation. We also present results from two-photon laser-scanning microscopy studies of alpha-actinin/enhanced green fluorescent protein fusion constructs at the basal membrane of living CHO cells. Using space-time image correlation spectroscopy (STICS), we are able to measure protein fluxes with magnitudes of mum/min from retracting lamellar regions and protrusions for adherent cells. We also demonstrate the measurement of correlated directed flows (magnitudes of mum/min) and diffusion of interacting alpha5 integrin/enhanced cyan fluorescent protein and alpha-actinin/enhanced yellow fluorescent protein within living CHO cells. The STICS method permits us to generate complete transport maps of proteins within subregions of the basal membrane even if the protein concentration is too high to perform single particle tracking measurements.

Actinin↗

PubMed1983

Cytotoxic T-lymphocytes. How do they function?

The central theme of this work has been the roles of the CTL receptor and of MHC-proteins in CTL recognition and lysis. A major conclusion that may be deduced from the work presented here is that one CTL receptor is responsible for both target cell recognition and lysis. Although their function as recognitive structures is well established, involvement of MHC-proteins in the events that follow recognition has not been investigated in detail. We have proposed that MHC-proteins are molecular mediators whereby CTL receptors transmit signals ultimately leading to lysis of the target cell. I see future work on CTL-mediated lysis proceeding in the following directions: 1. Verification and analysis of the precise role of MHC proteins in CTL recognition and lysis by use of cell and vesicle systems of defined composition and structure. 2. Study of CTL-mediated 'lethal hit' in systems enabling analysis of early events (millisecond level) preceding lysis. 3. Grafting of CTL receptor(s) activity onto naive cells, using liposomes or other vehicles, and 4. Production of idiotypic reagents such as monoclonal antibodies specific for the combining site/effector mechanism of CTL.

Animals↗

PubMed1987

[Presentation of a survey scale for neuro-rehabilitation--with a general discussion of current rehabilitation scales].

Presented is a newly developed rehabilitation survey scale for the field of neuro-rehabilitation, in which levels of functioning relative to ten specific and four global functions are documented at set intervalls by means of a five-grade rating scale. Additionally, numerous symptoms or associated phenomena are recorded ungraded. Used in the clinical setting, this scale affords a quick overview of the rehabilitation profile, with relatively little time and effort being required. Scientific evaluation may uncover information on the impact the disorders of specific functions have on global functions, which is relevant primarily in the social context. Also, the scale may be applied in the framework of larger-scale comparative studies. Dealt with in an extensive discussion, our newly presented scale may be accorded a number of advantages over currently used scales, which warrant its introduction. It is the outcome of many years of developmental work and pertaining reliability verifications.

Activities of Daily Living↗

PubMed1992

Quantitative single photon emission tomography: verification for sources in an elliptical water phantom.

Accurate absorbed dose calculations are important for a proper dose planning in internal radionuclide therapy. The activity distribution must be measured and the target volume defined. This can be done with single photon emission tomography (SPET) if proper attenuation and scatter correction are employed. This study investigated the calculation of the activity and the volume of different spherical sources. These two parameters are essential for a proper dose calculation. The scatter and attenuation correction method is based on spatially variant scatter functions and density maps. The volume calculation method is based on obtaining a threshold from a grey-level histogram. Both point sources and spheres of different diameters containing technetium-99m were placed in different locations in an elliptical water phantom and imaged by SPET. The activity and the volume of the spheres were calculated from the SPET images and compared with known activities. Results show a quantification of activity within 10% for most of the sources. Important influences on the quantification are (a) the presence of artefacts due to improper reconstruction and (b) the finite spatial resolution which affects the total number of counts within the determined volume.

Humans↗

PubMed1985

Molecular cloning of alpha-amylase genes from Drosophila melanogaster. II. Clone organization and verification.

Restriction maps of an alpha-amylase structural gene clone, lambda Dm65, and of four putative alpha-amylase pseudogene clones are presented. Two alpha-amylase structural genes, inverted with respect to each other, are contained in lambda Dm65. Subregions of internal DNA sequence homology within lambda Dm65 and of cross-homology between the presumptive pseudogene clones and lambda Dm65 were determined. Subregions of cross-homology between the Drosophila clones and the mouse alpha-amylase cDNA clone, pMSa104, were also determined. The presence of functional alpha-amylase structural genes in lambda Dm65 was verified by injection of appropriate subclones into the germinal vesicle of Xenopus oocytes, followed by incubation of the oocytes under conditions that allowed coupled transcription and translation of injected genes to occur. Subclones of the 3.8- and 5.6-kb EcoRI fragments of lambda Dm65 were shown to code for alpha-amylase isozymes 1 and 3, respectively, of Drosophila melanogaster Canton-S. Both subclones are homologous to RNA of a size sufficient to accommodate the alpha-amylase-coding information. No RNA species homologous to other subcloned EcoRI fragments of lambda Dm65 was detected.

Animals↗

PubMed2006

Huntingtin interacts with the receptor sorting family protein GASP2.

Protein interaction networks are useful resources for the functional annotation of proteins. Recently, we have generated a highly connected protein-protein interaction network for Huntington's disease (HD) by automated yeast two-hybrid (Y2H) screening (Goehler et al., 2004). The network included several novel direct interaction partners for the disease protein huntingtin (htt). Some of these interactions, however, have not been validated by independent methods. Here we describe the verification of the interaction between htt and GASP2 (G protein-coupled receptor associated sorting protein 2), a protein involved in membrane receptor degradation. Using membrane-based and classical coimmunoprecipitation assays we demonstrate that htt and GASP2 form a complex in cotransfected mammalian cells. Moreover, we show that the two proteins colocalize in SH-SY5Y cells, raising the possibility that htt and GASP2 interact in neurons. As the GASP protein family plays a role in G protein-coupled receptor sorting, our data suggest that htt might influence receptor trafficking via the interaction with GASP2.

Animals↗

PubMed2006

Effects of age and mild cognitive impairment on direct and indirect access to arithmetic knowledge.

The present study aimed at investigating age-related changes and mild cognitive impairment (MCI) related effects in simple arithmetic. To pursue this goal, MCI patients, healthy old adults and young adults performed three computerised tasks. The production (e.g., 3 x 4=?) and the verification task (3 x 4 12?) evaluated direct access to multiplication knowledge, the number-matching task (3 x 4 34?, 'do 3 x 4 and 34 have the same digits?') tested indirect access. In verification and number-matching, interference from related distractors (e.g., 3 x 4 followed by 16) relative to unrelated distractors (17) reflects access to stored fact representations as well as efficiency of inhibition processes. Results indicated that, compared to young adults, MCI and healthy old adults were slower in responding across tasks. In production and verification, analyses of individual latency regression slopes and intercepts suggested that these age effects were related to differences at peripheral processing stages (e.g., encoding) rather than at the central (arithmetic retrieval) stage. Differences between MCI and healthy elderly emerged only in the number-matching task. While in verification effects were comparable between groups, in number-matching MCI patients were more susceptible to interference from irrelevant information than healthy old participants. Overall, the present findings indicate that aging has a general effect on peripheral processing speed, but not on arithmetic memory retrieval. Parietal cortico-subcortical circuits mediating arithmetic fact retrieval (Dehaene, S., & Cohen, L. (1995). Towards an anatomical and functional model of number processing. Mathematical Cognition, 1, 83-120; Dehaene, S., & Cohen, L. (1997). Cerebral pathways for calculation: Double dissociation between rote verbal and quantitative knowledge of arithmetic. Cortex, 33, 219-250) thus seem to be preserved in normal aging and MCI. In contrast, MCI patients show enhanced interference in number-matching. This task-specific lack of inhibition may point to dysfunctional frontal cortico-subcortical networks in MCI.

Adult↗

PubMed1979

Flavin-dependent substrate photo-oxidation as a chemical model of dehydrogenase action.

As a model of flavin-dependent biological dehydrogenation, flavin-sensitized photodehydrogenation and photodecarboxylation were studied by variation of substrate, flavin, pH and solvent. Evidence for the following rules is given. (1) When the reactive site of a photosubstrate is an alpha-carbon atom of the type CH-CO2-, decarboxylation is preferred over dehydrogenation, whereas the reverse is true for the neutral CH-CO2H. (2) Consequently these reactions do not exhibit a measurable isotope effect with C2H-CO2-, in contrast with the findings by Penzer, Radda, Taylor & Taylor [(1970) Vitam. Horm. (N.Y.) 28, 441--466], which could not be reproduced. When the substate does not contain a carboxylate group, isotope effects occur, in verification of previous reports, e.g. for benzyl alcohol C6H5-C2H20H. (3) The mechanism of flavin-sensitized substrate photodecarboxylation is assumed to consist in a primary carbanion fixation at the flavin nucleus (position 4a, 5 or 8) with concomitant liberation of CO2. This step is followed by rapid fragmentation of the adduct CH-Fl-red., provided that the substrate contains a functional and electron-donating group X, e.g. X = OH, OCH3 or NH2 (but not NH3+ !) in X CH-CO2-. (4) The minimal requirement for flavin-sensitized C-H dehydrogenation is the presence of a hydroxyl group. For example, methanol as substrate and solvent is dehydrogenated at pH sufficiently alkaline for detection of the presence of the active species CH3O-, whereas at more acidic pH substrate dehydrogenation is competing with flavin autophotolysis, which depends on the substituents in the flavin nucleus.

Carboxylic Acids↗

PubMed2001

Observation of direct dissociative ionization in molecular hydrogen.

Direct dissociative ionization is the simplest three-body breakup process in H2. We describe the experimental verification of direct dissociative ionization to the repulsive A2Sigma(+)(u) state by resolving the kinetic energy and angular distributions of the formed protons. A (2+1) resonant enhanced multiphoton ionization process via the isotropic E,F (1)Sigma(+)(g)(v = 6,J = 0) level is employed. The structure in the kinetic energy spectrum is well described by a projection of the vibrational wave function of the E,F (1)Sigma(+)(g)(v = 6,J = 0) state onto the repulsive ionic state. The electronic character of the ionization continuum is revealed by the proton angular distribution.

Journal Article↗

PubMedDate not supplied

[Value of injection hepato-lymphography during percutaneous transhepatic cholangiography in patients with cholestasis].

Injection hepatography (IH) was made in 278 patients with cholestasis to study the drainage function of the liver. In 208 cases. IH was performed as a test during percutaneous transhepatic cholangiography (PTHC). The hepatic lymph pathways were imaged in 167 (60%) patients. Images of the biliary tract were obtained in 245 (88.1%) patients with cholestasis, it being not dilated in 34 (12.2%) patients. The fact that hepatolymphography may be performed during PTHC as an independent test permits verification of hepatic lymph circulatory disorders that are an index of the rate of inflammation in the organ.

Adult↗