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Biomedical subjects

K Vogel

Publications and source records attributed to K Vogel.

At least 19 recordsLinked to original sources
PubMed2005

Filler features and their effects on wear and degree of conversion of particulate dental resin composites.

Based on the incomplete understanding on how filler features influence the wear resistance and monomer conversion of resin composites, this study sought to evaluate whether materials containing different shapes and combinations of size of filler particles would perform similarly in terms of three-body abrasion and degree of conversion. Twelve experimental monomodal, bimodal or trimodal composites containing either spherical or irregular shaped fillers ranging from 100 to 1500 nm were examined. Wear testings were conducted in the OHSU wear machine (n = 6) and quantified after 10(5) cycles using a profilometer. Degree of conversion (DC) was measured by FTIR spectrometry at the surface of the composites (n = 6). Data sets were analyzed using one-way Anova and Tukey's test at a significance level of 0.05. Filler size and geometry was found to have a significant effect on wear resistance and DC of composites. At specific sizes and combinations, the presence of small filler particles, either spherical or irregular, may aid in enhancing the wear resistance of composites, without compromising the percentage of reacted carbon double bonds.

Composite Resins↗
PubMed2003

Characteristics of fungal phytases from Aspergillus fumigatus and Sartorya fumigata.

Aspergillus fumigatus phytase has previously been identified as a phytase with a series of favourable properties that may be relevant in animal and human nutrition, both for maximising phytic acid degradation and for increasing mineral and amino acid availability. To study the natural variability in amino acid sequence and its impact on the catalytic properties of the enzyme, we cloned and overexpressed the phytase genes and proteins from six new purported A. fumigatus isolates. Five of these phytases displayed < or= 2 amino acid substitutions and had virtually identical stability and catalytic properties when compared with the previously described A. fumigatus ATCC 13073 phytase. In contrast, the phytase from isolate ATCC 32239 ( Sartorya fumigata, the anamorph of which was identified as A. fumigatus) was more divergent (only 86% amino acid sequence identity), had a higher specific activity with phytic acid, and displayed distinct differences in substrate specificity and pH-activity profile. Finally, comparative experiments confirmed the favourable stability and catalytic properties of A. fumigatus phytase.

6-Phytase↗
PubMed2001

Carbon monoxide binding by de novo heme proteins derived from designed combinatorial libraries.

Carbon monoxide binding was studied in a collection of de novo heme proteins derived from combinatorial libraries of sequences designed to fold into 4-helix bundles. The design of the de novo sequences was based on the previously reported "binary code" strategy, in which the patterning of polar and nonpolar amino acids is specified explicitly, but the exact identities of the side chains are varied extensively.(1) The combinatorial mixture of amino acids included histidine and methionine, which ligate heme iron in natural proteins. However, no attempt was made to explicitly design a heme binding site. Nonetheless, as reported previously, approximately half of the binary code proteins bind heme.(2) This collection of novel heme proteins provides a unique opportunity for an unbiased assessment of the functional potentialities of heme proteins that have not been prejudiced either by explicit design or by evolutionary selection. To assess the capabilities of the de novo heme proteins to bind diatomic ligands, we measured the affinity for CO, the kinetics of CO binding and release, and the resonance Raman spectra of the CO complexes for eight de novo heme proteins from two combinatorial libraries. The CO binding affinities for all eight proteins were similar to that of myoglobin, with dissociation constants (K(d)) in the low nanomolar range. The CO association kinetics (k(on)) revealed that the heme environment in all eight of the de novo proteins is partially buried, and the resonance Raman studies indicated that the local environment around the bound CO is devoid of hydrogen-bonding groups. Overall, the CO binding properties of the de novo heme proteins span a narrow range of values near the center of the range observed for diverse families of natural heme proteins. The measured properties of the de novo heme proteins can be considered as a "default" range for CO binding in alpha-helical proteins that have neither been designed to bind heme or CO, nor subjected to genetic selections for heme or CO binding.

Carbon Monoxide↗
PubMed2001

Tumor vascularization in experimental melanomas: correlation between unenhanced and contrast enhanced power Doppler imaging and histological grading.

The purpose of this study was to evaluate the reliability of unenhanced and enhanced power Doppler sonography in visualization of intratumoral angioneogenesis. Thirty-seven malignant melanomas, which had been implanted intra- or subcutaneously in 22 mice, were examined. Various B-mode criteria, power Doppler criteria and spectral Doppler parameters were evaluated before and after IV-application of the d-galactose-based contrast agent Levovist. After sonographic examination, all tumors were analyzed histologically with semiquantitative grading of tumoral vascularization. Unenhanced, in 70% of the tumors, no intratumoral vessels were visible using power Doppler, but only in 11% of the intracutaneous and in 0% of the subcutaneous after injection of the contrast agent. The enhanced mode was definitely superior to unenhanced Doppler in showing the intratumoral vascularity. The intratumoral vascular structure could be sufficiently analyzed in 30% of all tumors by unenhanced Doppler, but in 92% enhanced. The mean percentage vessel area increased about 433% after application of Levovist (intracutaneous: 485%, subcutaneous: 280%). Despite the missing direct correlation between the sonographically and histologically determined grade of tumor vascularization (Pearson's correlation unenhanced 0,356, p <.05/enhanced 0.395, p <.05), the correlation between the percentage vessel area and the histologic grade of vascularization was improved after application of the contrast agent (Pearson's correlation unenhanced 0.347, p <.05/enhanced 0.686, p <.01). We did not find a significant direct correlation between histologically and sonographically determined degree of vascularization. However, the correlation was improved using a d-galactose-based signal-enhancing agent in power Doppler sonography.

Animals↗
PubMed2000

Active site residue 297 of Aspergillus niger phytase critically affects the catalytic properties.

The wild-type phytases from the Aspergillus niger strains NRRL 3135 and T213 display a three-fold difference in specific activity (103 versus 32 U/mg protein), despite only 12 amino acid differences that are distributed all over the sequence of the protein. Of the 12 divergent positions, three are located in or close to the substrate binding site. Site-directed mutagenesis of these residues in A. niger T213 phytase showed that the R297Q mutation (R in T213, Q in NRRL 3135) fully accounts for the differences in catalytic properties observed. Molecular modelling revealed that R297 may directly interact with a phosphate group of phytic acid. The fact that this presumed ionic interaction - causing stronger binding of substrates and products - correlates with a lower specific activity indicates that product (myo-inositol pentakisphosphate) release is the rate-limiting step of the reaction.

6-Phytase↗
PubMed2000

Targeting of SNAP-25 to membranes is mediated by its association with the target SNARE syntaxin.

The docking and fusion of synaptic vesicles with the presynaptic plasma membrane require the interaction of the vesicle-associated membrane protein VAMP with the plasma membrane proteins syntaxin and SNAP-25. Both of these proteins behave as integral membrane proteins, although they are unusual in that they insert into membranes post-translationally. Whereas VAMP and syntaxin possess hydrophobic transmembrane domains, SNAP-25 does not, and it is widely believed that SNAP-25 traffics to and inserts into membranes by post-translational palmitoylation. In pulse-chase biosynthesis studies, we now show that SNAP-25 and syntaxin rapidly bind to each other while still in the cytosol of neuroendocrine and transfected heterologous cells. Cell fractionation studies revealed that cytosolic SNAP-25.syntaxin complexes then traffic to and insert into membranes. Furthermore, the association of SNAP-25 with membranes is dramatically enhanced by syntaxin, and the transmembrane domain of syntaxin is essential for this effect. Surprisingly, despite the importance of the SNAP-25 palmitoylation domain for membrane anchoring at steady state, removal of this domain did not inhibit the initial association of newly synthesized SNAP-25 with membranes in the presence of syntaxin. These data demonstrate that the initial attachment of newly synthesized SNAP-25 to membranes is a consequence of its association with syntaxin and that it is only after syntaxin-mediated membrane tethering that SNAP-25 is palmitoylated.

Animals↗
PubMed1999

SNAP-23 and SNAP-25 are palmitoylated in vivo.

The neuronal presynaptic membrane t-SNARE complex consists of the transmembrane protein syntaxin with the palmitoylated protein SNAP-25. In non-neuronal tissues, SNAP-23 replaces SNAP-25 in the t-SNARE complex, although the mechanism of membrane anchoring of SNAP-23 has not been determined. We now report that like SNAP-25, SNAP-23 is palmitoylated in vivo on one or more cysteine residues present in a central "palmitoylation domain." Interestingly, SNAP-23 is palmitoylated less well than SNAP-25, and in vivo binding studies indicate a correlation between the extent of palmitoylation and the ability of SNAP-23 or SNAP-25 to bind to syntaxin in vivo.

Amino Acid Sequence↗
PubMed1999

Biophysical characterization of fungal phytases (myo-inositol hexakisphosphate phosphohydrolases): molecular size, glycosylation pattern, and engineering of proteolytic resistance.

Phytases (myo-inositol hexakisphosphate phosphohydrolases) are found naturally in plants and microorganisms, particularly fungi. Interest in these enzymes has been stimulated by the fact that phytase supplements increase the availability of phosphorus in pig and poultry feed and thereby reduce environmental pollution due to excess phosphate excretion in areas where there is intensive livestock production. The wild-type phytases from six different fungi, Aspergillus niger, Aspergillus terreus, Aspergillus fumigatus, Emericella nidulans, Myceliophthora thermophila, and Talaromyces thermophilus, were overexpressed in either filamentous fungi or yeasts and purified, and their biophysical properties were compared with those of a phytase from Escherichia coli. All of the phytases examined are monomeric proteins. While E. coli phytase is a nonglycosylated enzyme, the glycosylation patterns of the fungal phytases proved to be highly variable, differing for individual phytases, for a given phytase produced in different expression systems, and for individual batches of a given phytase produced in a particular expression system. Whereas the extents of glycosylation were moderate when the fungal phytases were expressed in filamentous fungi, they were excessive when the phytases were expressed in yeasts. However, the different extents of glycosylation had no effect on the specific activity, the thermostability, or the refolding properties of individual phytases. When expressed in A. niger, several fungal phytases were susceptible to limited proteolysis by proteases present in the culture supernatant. N-terminal sequencing of the fragments revealed that cleavage invariably occurred at exposed loops on the surface of the molecule. Site-directed mutagenesis of A. fumigatus and E. nidulans phytases at the cleavage sites yielded mutants that were considerably more resistant to proteolytic attack. Therefore, engineering of exposed surface loops may be a strategy for improving phytase stability during feed processing and in the digestive tract.

6-Phytase↗
PubMed1999

Leptin antagonizes the insulin-like growth factor-I augmentation of steroidogenesis in granulosa and theca cells of the human ovary.

There is increasing evidence that leptin is a physiological link between obesity and infertility. Although leptin receptors have been demonstrated in human ovaries, there is no information regarding the effects of leptin on cells from developing ovarian follicles. To test the direct effects of leptin on human ovarian cells, granulosa cells (GC) and theca cells were isolated from the ovaries of regularly cycling women. Serum was obtained at the time of surgery, and follicular fluid was aspirated from the follicles before isolation of the ovarian cells. Leptin concentrations were similar in follicular fluid and serum. RT-PCR analysis demonstrated that the long, signaling form of the leptin receptor was expressed in both theca and GC. In cultured GC, leptin had no effect on estradiol production, alone or in the presence of FSH, but caused a concentration-related inhibition of the insulin-like growth factor I (IGF-I) augmentation of FSH-stimulated estradiol production. The effect of leptin was specific, because there was no effect on progesterone production. In cultured theca cells, leptin did not alter androstenedione production, alone or in the presence of LH. Leptin caused a concentration-related inhibition of the IGF-I augmentation of LH-stimulated androstenedione production. These data demonstrate that leptin can directly inhibit IGF-I action in ovarian theca and GC at concentrations commonly present in obese women.

Adult↗
PubMed1999

SPIO-MR imaging versus double-phase spiral CT in detecting malignant lesions of the liver.

PURPOSE: To assess the diagnostic performance of superparamagnetic iron oxide MR (SPIO-MR) imaging compared with double-phase spiral CT in detecting liver metastases and hepatocellular carcinoma. MATERIAL AND METHODS: Thirty-eight patients with a total of 144 malignant lesions of the liver were examined by CT and SPIO-MR. After definition of a gold standard by a panel of experts, the patient images were randomized and presented to a blinded jury of 5 independent observers whose task was to identify as many lesions as possible. The results were tested for statistical significance using multifactorial analysis of variance (alpha=5%). RESULTS: SPIO-MR produced the highest detection rate and was significantly superior (p<0.05) to unenhanced MR imaging and double spiral-phase contrast-enhanced CT (DPS-CECT). Maximum performance in DPS-CECT was obtained during the portal venous contrast phase but was significantly inferior to SPIO-MR imaging. The scores for unenhanced CT and unenhanced MR were significantly lower than for the corresponding enhanced procedures. SPIO-MR imaging produced a higher incidence of false-positive findings (n=39). CONCLUSION: SPIO-MR produced a significantly better detection rate for malignant focal liver lesions compared with double-phase spiral DPS-CECT but was associated with a higher rate of false-positive findings.

Adult↗
PubMed1998

Oral immunization with enterocoated microbeads induces antigen-specific cytolytic T-cell responses.

A cytolytic T-lymphocyte (CTL) response has been generated to a protein antigen via oral administration in an experimental murine system. This was achieved by the formulation of the test antigen ovalbumin (OVA) in a pH-sensitive microsphere particle (microbead) with an acrylic-based coating to avoid its degradation by gastric enzymes. Comparative studies of orally administered enterocoated microbead-OVA particles, versus the more traditional formulation of OVA in adjuvant (DETOX-PC) given subcutaneously as immunogen, were conducted; both elicited comparable responses in the generation of antigen-specific lymphoproliferative and CTL responses. CTL lines generated via oral administration of antigen were shown to be CD3(+), CD4(-), and CD8(+). CTL lysis of OVA peptide-pulsed targets was shown to be inhibited by anti-CD8 antibody. Whereas oral administration of pH-sensitive enterocoated microbeads containing proteins has previously been shown to elicit antibody and lymphoproliferative T-cell responses to protein antigens, this is the first demonstration of the generation of an antigen-specific cytolytic T-cell response via oral administration of a protein immunogen formulated in such microbeads.

Administration, Oral↗
PubMed1998

[MRI with supermagnetic iron particles versus double-spiral CT in identification of malignant liver lesions].

PURPOSE: The purpose of this study was to evaluate the efficacy of MRI with superparamagnetic iron oxide (SPIO) and double-spiral CT in the detection of liver metastases and hepatocellular carcinoma. METHODS: 38 patients with a total of 144 malignant hepatic lesions underwent CT and MRI. A panel of experts defined the gold standard. Five experienced judges performed independently blinded evaluation of the number of detectable lesions. Multifactorial variance analysis was used to determine the statistical significance. RESULTS: SPIO-MRI shows the highest rate of detection and is significantly superior to native MRI and native CT. The highest rate of detection by CT is shown in the portal-venous phase of contrast; nevertheless, the rate is significantly inferior to SPIO-MRI. In general, the native phases of CT and MRI are significantly inferior to the contrast phases of both. SPIO-MRI shows a higher rate of false positive findings. CONCLUSION: The time-consuming and cost-intensive SPIO-MRI significantly increases the rate of detectability for malignant liver lesions compared with double-spiral CT but it also increases the rate of false positive findings.

Adult↗
PubMed1996

Subtilisin-catalyzed religation of proteolyzed hen egg-white lysozyme: investigation of the role of disulfides.

BACKGROUND: The use of proteases to form, instead of break, peptide bonds has expanded the repertoire of techniques available for protein semisynthesis. Several groups have previously reported the use of proteases in aqueous-organic solvents to form single amide bonds within proteins, but low yields and lengthy reaction times make this an impractical approach to protein synthesis. We recently found that proteolyzed triose phosphate isomerase can be re-ligated rapidly and efficiently by subtilisin, in mixed aqueous-organic solvent systems. RESULTS: We now report the use of subtilisin to resynthesize hen egg-white lysozyme from a mixture of its proteolyzed fragments in high yield and with rapid reaction times. This enzymatic religation can also be achieved after reduction of the four disulfide bonds present in lysozyme, with the same efficiency as that observed for the disulfide-containing proteolysis mixture. CONCLUSIONS: For egg-white lysozyme, the subtilisin religation reaction can be used to re-synthesize a proteolyzed protein even after reduction of disulfide bonds. The utility of this reaction in more generalized protein semisynthesis reactions is currently being explored.

Animals↗
PubMed1995

The adolescent child health and illness profile. A population-based measure of health.

This study was designed to test the reliability and validity of an instrument to assess adolescent health status. Reliability and validity were examined by administration to adolescents (ages 11-17 years) in eight schools in two urban areas, one area in Appalachia, and one area in the rural South. Integrity of the domains and subdomains and construct validity were tested in all areas. Test/retest stability, criterion validity, and convergent and discriminant validity were tested in the two urban areas. Iterative testing has resulted in the final form of the CHIP-AE (Child Health and Illness Profile-Adolescent Edition) having 6 domains with 20 subdomains. The domains are Discomfort, Disorders, Satisfaction with Health, Achievement (of age-appropriate social roles), Risks, and Resilience. Tested aspects of reliability and validity have achieved acceptable levels for all retained subdomains. The CHIP-AE in its current form is suitable for assessing the health status of populations and subpopulations of adolescents. Evidence from test-retest stability analyses suggests that the CHIP-AE also can be used to assess changes occurring over time or in response to health services interventions targeted at groups of adolescents.

Adolescent↗
PubMed1994

Protozoa as model systems for the study of cellular responses to altered gravity conditions.

The orientation behavior of Paramecium changed in a similar way after transition to conditions of free-fall in a sounding rocket and after transition to conditions of simulated weightlessness on a fast rotating clinostat. After a period of residual orientation, Paramecium cells distributed themselves randomly 80 s (120 s) after onset of free-fall (simulated weightlessness). Swimming velocity increased significantly; however, the increase was transient and subsided after 3 min in the rocket experiments, while the velocity remained enhanced even during 2 h of rotation on a fast clinostat. Trichocysts were present and without morphological changes in Paramecium cells which had been exposed to a rocket flight, as well as to fast or slow rotation on a clinostat. Regeneration of the oral apparatus of Stentor and morphogenesis of Eufolliculina proceeded normally on the clinostat. The results demonstrate that the clinostat is a useful tool to simulate the conditions of weightlessness on earth and to detect gravisensitive cellular functions.

Animals↗