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Biomedical subjects

G Maier

Publications and source records attributed to G Maier.

At least 19 recordsLinked to original sources
PubMed2006

Magnetic resonance spectroscopy for measuring the biodistribution and in situ in vivo pharmacokinetics of fluorinated compounds: validation using an investigation of liver and heart disposition of tecastemizole.

BACKGROUND AND OBJECTIVE: The study of biodistribution and in situ pharmacokinetics is a challenging, but sometimes very important, aspect of premarketing characterization of drugs. We aimed to develop a non-invasive fluorine magnetic resonance (MR) spectroscopic method for the absolute quantitation of a mono-fluorinated compound and of its metabolites in the heart and liver of healthy subjects for this purpose. METHOD: We used fluorine MR spectroscopy (MRS) at 4 T (Tesla) and external standardization in an open label multiple-dose study. Twenty-three healthy adult subjects were enrolled in the study. The surface coil localized fluorine MR spectrum was monitored in the heart and liver at baseline and after oral administration of multiple doses of tecastemizole. Steady-state measurements were made at set time points that depended upon dose, and washout measurements were made only on subjects in which in vivo fluorine signal was observed. RESULTS AND DISCUSSION: At 4 T, under the given experimental conditions, the method had a lower limit of quantitation (LLOQ) of about 2.6 microm and a limit of detection (LOD) of about 0.3 microm for solution state samples (linewidth approximately 15 Hz). The measurement reproducibility was 6.4% using a 50 microm phantom. The effect of MR operator and spectral analyst on the calculated calibration curve slope was small, with inter-rater correlation coefficients of 0.999 and 0.998 respectively. MR signal from fluorine-containing tecastemizole-related moieties was observed in situ only at day 8 in the liver of three of five subjects dosed at 270 mg/day. The average in situ concentration was estimated to be 58+/-22 microm, with an average test-retest reproducibility of 216%. Extrapolating the in vitro results to human measurements, with an approximate linewidth of 250 Hz, predicts in situ LOD and LLOQ values of approximately 6 and 44 microm respectively. However, the human study had a fluorine MRS LOD of approximately 20 microm. The decrease in sensitivity and the increase in variability of the in vivo, in situ measurements compared with the validation study most likely arose from coil placement and incomplete rephasing of the MR signal by the respiratory phase compensation method. CONCLUSION: The measured concentrations were the lowest ever recorded for a multi-dose exogenous mono-fluorinated compound in the human liver using a validated fluorine MR quantitation method. The proposed non-invasive MR method for studying the biodistribution and in situ pharmacokinetics of mono-fluorinated compounds in the liver and heart should have broader application to the development of non-invasive biomarkers.

Adult↗
PubMed2005

Detection and imaging of atmospheric radio flashes from cosmic ray air showers.

The nature of ultrahigh-energy cosmic rays (UHECRs) at energies >10(20) eV remains a mystery. They are likely to be of extragalactic origin, but should be absorbed within approximately 50 Mpc through interactions with the cosmic microwave background. As there are no sufficiently powerful accelerators within this distance from the Galaxy, explanations for UHECRs range from unusual astrophysical sources to exotic string physics. Also unclear is whether UHECRs consist of protons, heavy nuclei, neutrinos or gamma-rays. To resolve these questions, larger detectors with higher duty cycles and which combine multiple detection techniques are needed. Radio emission from UHECRs, on the other hand, is unaffected by attenuation, has a high duty cycle, gives calorimetric measurements and provides high directional accuracy. Here we report the detection of radio flashes from cosmic-ray air showers using low-cost digital radio receivers. We show that the radiation can be understood in terms of the geosynchrotron effect. Our results show that it should be possible to determine the nature and composition of UHECRs with combined radio and particle detectors, and to detect the ultrahigh-energy neutrinos expected from flavour mixing.

Journal Article↗
PubMed2005

[Computer-assisted arthrodesis of the sacroiliac joint].

Arthrodesis of the sacroiliac joint (SI) usually requires a large surgical exposure using the lateral approach. Computer-assisted surgery based on intraoperative 3D fluoroscopy imaging can reduce the approach to stab incisions. The clinical example shows the insertion of two screws and a cylindrical bone graft to achieve an arthrodesis of the SI joint. The intraoperatively navigated placement of implants and bone graft was performed only by stab incisions.

Adult↗
PubMed2004

Regulation of the voltage gated K+ channel Kv1.3 by the ubiquitin ligase Nedd4-2 and the serum and glucocorticoid inducible kinase SGK1.

The stimulation of cell proliferation by insulin like growth factor IGF-1 has previously been shown to depend on activation of voltage gated K(+) channels. The signaling involved in activation of voltage gated K(+) channel Kv1.3 includes the phosphatidylinositol-3 (PI3) protein kinase, 3-phosphoinositide dependent protein kinase PDK1 and the serum and glucocorticoid inducible kinase SGK1. However, nothing is known about mechanisms mediating the stimulation of Kv1.3 by SGK1. Most recently, SGK1 has been shown to phosphorylate and thus inactivate the ubiquitin ligase Nedd4-2. The present study has been performed to explore whether the regulation of Kv1.3 involves Nedd4-2. To this end Kv1.3 has been expressed in Xenopus oocytes with or without coexpression of Nedd4-2 and/or constitutively active (S422D)SGK1. In oocytes expressing Kv1.3 but not in water injected oocytes, depolarization from a holding potential of -80 mV to +20 mV triggers rapidly inactivating currents typical for Kv1.3. Coexpression of Nedd4-2 decreases, coexpression of (S422D)SGK1 enhances the currents significantly. The effects of either Nedd4-2 or of SGK1 are abrogated by destruction of the respective catalytic subunits ((C938S)Nedd4-2 or (K127N)SGK1). Further experiments revealed that wild type SGK1 and SGK3 and to a lesser extent SGK2 are similarly effective in stimulating Kv1.3 in both, presence and absence of Nedd4-2. It is concluded that Kv1.3 is downregulated by Nedd4-2 and stimulates by SGK1, SGK2, and SGK3. The data thus disclose a novel mechanism of Kv1.3 channel regulation.

Animals↗
PubMed2001

An experimental and theoretical study of the basicity of tetra-tert-butyltetrahedrane.

The gas-phase basicity (GB) of tetra-tert-butyltetrahedrane (tBu4THD) was determined by FT-ICR mass spectrometry and comparison with reference compounds of known basicity. Its GB, 1035+/-10 kJ x mol(-1), makes tetra-tert-butyltetrahedrane one of the strongest bases reported so far. Ab initio calculations [B3LYP/6-31G(d) and B3LYP/6-311 + G(d,p)//6-31G(d)] have been carried out in order to compare the high experimental basicity of tBu4THD with that estimated theoretically. Both B3LYP/6-31G(d) and QCISD(T) calculations were used to determine the reaction path which connects the initial tetrahedrane-ammonium complex with the final products, protonated cyclobutadiene (CBDH+) and ammonia.

Journal Article↗
PubMed2001

Assessment of porcine bone metabolism by dynamic.

UNLABELLED: The aim of this study was to quantify regional bone blood flow and [(18)F]fluoride ion influx with [(18)F]fluoride ion PET and correlate the results with specific static and dynamic indices of bone metabolism in healthy pigs. METHODS: During continuous ventilation (fractional concentration of oxygen in inspired gas = 0.3), dynamic PET scans 120 min in duration were obtained for 9 mini pigs after intravenous injection of 10.0 +/- 1.2 MBq (mean +/- SD) of [(18)F]fluoride ion per kilogram of body weight. Iliac crest bone biopsies were performed immediately before the PET scan to determine static and dynamic indices of bone metabolism (i.e., the mineral apposition rate) by bone histomorphometry. Kinetic rate constants describing influx (K(1)) and efflux (k(2)) of [(18)F]fluoride as well as chemisorption and incorporation of [(18)F]fluoride (k(3)) and reverse transport (k(4)) were determined for 6 vertebral bodies in each animal. Blood flow estimates (f) were derived from K(1) values corrected for the permeability-surface area product using a previously derived correction algorithm. A rate constant describing the net forward transport rate of fluoride (K(i)) and the fluoride volume flux (K(flux)) derived from a 2-tissue-compartment model was calculated and compared with the results of Patlak graphic analysis (K(pat)). RESULTS: A significant correlation was found between mineral apposition rate and K(i) (P < 0.005), K(flux) (P < 0.01), K(pat), K(1), and f (P < 0.05). The values of f, K(i), K(flux), and K(pat) did not correlate significantly with other static or dynamic histomorphometric indices or with age, serum alkaline phosphatase, or parathyroid hormone levels. The values of f and K(i) correlated linearly (y = 0.023 + 0.32x; r(2) = 0.74; P < 0.001). CONCLUSION: PET bone studies using [(18)F]fluoride ion provide quantitative estimates of bone blood flow and metabolic activity that correlate with histomorphometric indices of bone formation in the normal bone tissue of the mini pig. Therefore, it seem reasonable to assume that [(18)F]fluoride ion PET can reduce the number of invasive bone biopsies, thus facilitating follow-up of patients with metabolic bone diseases.

Animals↗
PubMed2000

Ring opening of 1-methylcyclopropene and cyclopropene: matrix infrared spectroscopic identification of 2-butene-1,3-diyl and propene-1,3-diyl

Triplet 2-butene-1,3-diyl (T-11) was generated on irradiation of 1-methylcyclopropene (10) in a bromine-doped xenon matrix and was characterized by means of IR spectroscopy for the first time. Experimental results suggest that triplet propene-1,3-diyl (T-3) is formed from cyclopropene (1) under similar conditions. In accordance with theoretical calculations, the experimental data indicate that the reactions 1-->3 and 10-->11 are the lowest energy ground-state pathways for the ring opening of 1 and 10, respectively.

Journal Article↗
PubMed2000

Isomerizations between nitrosyl halides X-N=O and isonitrosyl halides X-O-N: a matrix-spectroscopic study

Irradiation of nitrosyl bromide BrNO (4) with light of the wavelength lambda = 248 nm and nitrosyl chloride CINO (6) with lambda = 193 nm in an argon matrix at 10 K leads to the corresponding isomers isonitrosyl bromide BrON (5) and isonitrosyl chloride CION (7). Both new compounds 5 and 7 have been identified by comparison of the experimental and calculated (BLYP/6-311 + G*) IR spectra. Nitrosyl fluoride FNO (8) could not be transferred into isonitrosyl fluoride FON (9). The back reactions 5-->4 and 7-->6 can be initiated by UV (lambda > 310 nm), visible or IR light. The retransformation also occurs spontaneously in the matrix at 10 K under exclusion of any UV/Vis or IR radiation. Surprisingly, the reaction rates of these spontaneous back reactions are temperature independent between 8.5 and 25 K. The mechanism of these processes is discussed.

Journal Article↗
PubMed2000

[Gainful employment and women in the 2nd half of life].

The present study analyses gender-specific risks for women in the second half of their career, especially their labour force participation rate, the degree and quality of their vocational qualification, the risk of becoming unemployed, and their transition to retirement. The analysed data show that women obviously are at higher risks to participate in the labourforce throughout their biography. Therefore, strategies to improve the working conditions of female elderly workers should not only consist of age-specific policies, but of general practices in human resource development.

Aged↗
PubMed2000

Effect of Escherichia coli chaperonin GroELS on heterologously expressed human immunodeficiency virus type 1 reverse transcriptase in vivo and in vitro.

The two subunits of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (HIV-1 RT), p66 and p51, were coexpressed in Escherichia coli along with the E. coli chaperonin system GroEL/GroES. Coexpression increases the yield of heterodimeric HIV-1 RT by a factor of 4 to 5 and improves the nucleic acid binding affinity of HIV-1 RT by a factor of 1.6. We have analyzed the reasons for the improvements. The total increase in yield of HIV-1 RT can be attributed to an accumulation of RT subunits in the cells (factor of about 2.8) and an increased growth of the E. coli cells (factor of about 1.4). One reason for the accumulation in the cells is an improved stability of HIV-1 RT subunits toward bacterial proteases. In vitro studies showed that the nucleic acid binding affinity of HIV-1 RT purified from cells that did not coexpress GroELS was stimulated by adding purified GroELS (approx 1.5-fold), whereas HIV-1 RT stemming from cells coexpressing GroELS was stimulated only marginally (approx 1.1-fold). The in vivo as well as the in vitro studies suggest that the chaperonin interacts with HIV-1 RT and therefore affects the folding process of HIV-1 RT.

Cell Division↗
PubMed1999

Temporal coordination between initiation of HIV (+)-strand DNA synthesis and primer removal.

In this study, we have analyzed the interdependence between the polymerase and RNase H active sites of human immunodeficiency virus-1 reverse transcriptase (RT) using an in vitro system that closely mimics the initiation of (+)-strand DNA synthesis. Time course experiments show that RT pauses after addition of the 12th DNA residue, and at this stage the RNase H activity starts to cleave the RNA primer from newly synthesized DNA. Comparison of cleavage profiles obtained with 3'- and 5'-end-labeled primer strands indicates that RT now translocates in the opposite direction, i.e. in the 5' direction of the RNA strand. DNA synthesis resumes again in the 3' direction, after the RNA-DNA junction was efficiently cleaved. Moreover, we further characterized complexes generated before, during, and after position +12, by treating these with Fe2+ to localize the RNase H active site on the DNA template. Initially, when RT binds the RNA/DNA substrate, oxidative strand breaks were seen at a distance of 18 base pairs upstream from the primer terminus, whereas 17 base pairs were observed at later stages when the enzyme binds more and more DNA/DNA. These data show that the initiation of (+)-strand synthesis is accompanied by a conformational change of the polymerase-competent complex.

Base Pairing↗
PubMed1999

Poroelastic finite element analysis of a bone specimen under cyclic loading.

It had been suggested that the fluid embodied in bone lacunar-canalicular porosity may play an important role in bone remodelling [Weinbaum et al., 1994. Journal of Biomechanics 27, 339-360]. In this paper a finite element model of a poroelastic prismatic solid of rectangular cross-section is considered to simulate bone behaviour, precisely as in the previous work by Zhang and Cowin [Zhang and Cowin, 1994. Journal of Mechanical Physics of Solids 42, 1575-1599]. This solid is subject to combined cyclic axial and bending loads at its end. The objectives of the study are: (1) to verify the accuracy of the simplifying hypotheses underlying the analytical solutions established by the above authors; (2) to provide further insight into the behaviour of that solid; (3) to test the advantages in generality and versatility and the computing costs of general-purpose finite element codes in poroelastic analysis. The study is parametric with respect to the fluid leakage coefficient, to the ratio of the bending moment and axial load, and to the ratio of the characteristic relaxation time of the pore pressure over the excitation period. Results show that, for all the cases considered, the pore pressure distribution along the section height of the poroelastic beam exhibits a very good matching with previous analytical results. Stresses transversal with respect to the beam axis (assumed as constant or zero in previous analytical solutions) are evaluated. The analysis pointed out that: (1) the effects due to end-loads with zero resultants practically extinguish within a distance from the beam end almost equal to a typical length of the cross-section; (2) cross-sections remain plane above that distance; (3) the transversal total stresses are three orders of magnitude lower than axial stress.

Adaptation, Physiological↗
PubMed1999

Mixed reconstitution of mutated subunits of HIV-1 reverse transcriptase coexpressed in Escherichia coli - two tags tie it up.

The active form of HIV-1 reverse transcriptase (RT) is a p66/p51 heterodimer, in which the p51 subunit is generated by C-terminal proteolytic cleavage of p66. A well-known problem of p66 recombinant expression is partial cleavage of a 15-kDa peptide from the C-terminus by host proteases that can not be completely suppressed. In order to analyse the contribution of specific residues to a particular function in one distinct subunit, an expression and purification system is required that selects for the combination of the two individual subunits with the desired substitutions. We reconstituted the p66/p51 heterodimer from subunits coexpressed in Escherichia coli as an N-terminal fusion protein of glutathione S-transferase (GST) with p51 and a C-terminally His-tagged p66, respectively. The two-plasmid coexpression system ensures convenience for gene manipulation while degradation is reduced to a minimum, as dimerization protects the protein from further proteolysis. The combination of glutathione-agarose, phenyl-superose and Ni/nitrilotriacetate affinity chromatography allows rapid and selective purification of the desired subunit combination. Truncated forms of p51 are efficiently removed. Mobility-shift assay revealed that the preparations are free of p66 homodimer. In a successful test of the novel expression system, mixed reconstituted RTs with p51 selectively mutated in a putative nucleic acid binding motif (the so called helix clamp) show reduced binding of dsDNA in mobility-shift assays. This indicates the p51 subunit has an active role in DNA binding

Base Sequence↗
PubMed1998

Stimulation of phospholipase C-beta2 by the Rho GTPases Cdc42Hs and Rac1.

Neutrophils contain a soluble guanine-nucleotidebinding protein, made up of two components with molecular masses of 23 and 26 kDa, that mediates stimulation of phospholipase C-beta2 (PLCbeta2). We have identified the two components of the stimulatory heterodimer by amino acid sequencing as a Rho GTPase and the Rho guanine nucleotide dissociation inhibitor LyGDI. Using recombinant Rho GTPases and LyGDI, we demonstrate that PLCbeta2 is stimulated by guanosine 5'-O-(3-thiotriphosphate) (GTP[S])-activated Cdc42HsxLyGDI, but not by RhoAxLyGDI. Stimulation of PLCbeta2, which was also observed for GTP[S]-activated recombinant Rac1, was independent of LyGDI, but required C-terminal processing of Cdc42Hs/Rac1. Cdc42Hs/Rac1 also stimulated PLCbeta2 in a system made up of purified recombinant proteins, suggesting that this function is mediated by direct protein-protein interaction. The Cdc42Hs mutants F37A and Y40C failed to stimulate PLCbeta2, indicating that the Cdc42Hs effector site is involved in this interaction. The results identify PLCbeta2 as a novel effector of the Rho GTPases Cdc42Hs and Rac1, and as the first mammalian effector directly regulated by both heterotrimeric and low-molecular-mass GTP-binding proteins.

Amino Acid Sequence↗
PubMed1998

Crystal structure of an eight-base pair duplex containing the 3'-DNA-RNA-5' junction formed during initiation of minus-strand synthesis of HIV replication.

During initiation of minus-strand synthesis by HIV-1 reverse transcriptase, a 3'-DNA-RNA-5' junction is formed involving the 3'-end of tRNAlys,3. The HIV-RT-associated RNase H cleaves the RNA template strand specifically, opposite the newly synthesized DNA strand. We have determined the crystal structure at 1.9 A resolution of an eight-base pair hybrid duplex representing the junction to identify global or local structural perturbations which may be recognized by HIV-RT RNase H. The junction octamer is in a global A-type conformation throughout. A base pair step with distinct stacking geometry and variable backbone conformation is located next to the main endonucleolytic cleavage site. This base pair step may serve as a recognition site for HIV-RT RNase H.

Base Composition↗
PubMed1998

Synthesis and in vitro efficacy of transferrin conjugates of the anticancer drug chlorambucil.

One strategy for improving the selectivity and toxicity profile of antitumor agents is to design drug carrier systems employing soluble macromolecules or carrier proteins. Thus, five maleimide derivatives of chlorambucil were bound to thiolated human serum transferrin which differ in the stability of the chemical link between drug and spacer. The maleimide ester derivatives 1 and 2 were prepared by reacting 2-hydroxyethylmaleimide or 3-maleimidophenol with the carboxyl group of chlorambucil, and the carboxylic hydrazone derivatives 5-7 were obtained through reaction of 2-maleimidoacetaldehyde, 3-maleimidoacetophenone, or 3-maleimidobenzaldehyde with the carboxylic acid hydrazide derivative of chlorambucil. The alkylating activity of transferrin-bound chlorambucil was determined with the aid of 4-(4-nitrobenzyl)pyridine (NBP) demonstrating that on average 3 equivalents were protein-bound. Evaluation of the cytotoxicity of free chlorambucil and the respective transferrin conjugates in the MCF7 mammary carcinoma and MOLT4 leukemia cell line employing a propidium iodide fluorescence assay demonstrated that the conjugates in which chlorambucil was bound to transferrin through non-acid-sensitive linkers, i.e., an ester or benzaldehyde carboxylic hydrazone bond, were not, on the whole, as active as chlorambucil. In contrast, the two conjugates in which chlorambucil was bound to transferrin through acid-sensitive carboxylic hydrazone bonds were as active as or more active than chlorambucil in both cell lines. Especially, the conjugate in which chlorambucil was bound to transferrin through an acetaldehyde carboxylic hydrazone bond exhibited IC50 values which were approximately 3-18-fold lower than those of chlorambucil. Preliminary toxicity studies in mice showed that this conjugate can be administered at higher doses in comparison to unbound chlorambucil. The structure-activity relationships of the transferrin conjugates are discussed with respect to their pH-dependent acid sensitivity, their serum stability, and their cytotoxicity.

Animals↗
PubMed1998

Localization of the active site of HIV-1 reverse transcriptase-associated RNase H domain on a DNA template using site-specific generated hydroxyl radicals.

Reverse transcriptase (RT)-associated ribonuclease H (RNase H) can cleave both the RNA template of DNA/RNA hybrids as well as double-stranded (ds) RNA. This report shows that human immunodeficiency virus (HIV)-RT can also cleave the template strand of dsDNA when Mg2+ is replaced by Fe2+ in the RNase H active site of HIV-RT. The cleavage mechanisms as well as the positions of the cut vary depending on whether RNA or DNA is used. While DNA is cleaved 17 base positions upstream of the primer 3'-end, RNA is cleaved 18 base positions upstream. Competition experiments show that Fe2+ replaces the catalytically active Mg2+ of RT-associated RNase H. The bound Fe2+ is the source of locally generated OH-radicals that cleave the most proximate base in the DNA. Electrophoretic mobility studies of the cleaved fragments suggest that DNA is cleaved by an oxidative mechanism, while RNA is cleaved by an enzymatic mechanism which is indistinguishable from the Mg2+-dependent cleavage. The Fe2+-dependent cuts can be used to trace the active site of RT-associated RNase H on dsDNA as well as on dsRNA and DNA/RNA hybrids. The observed 1 base difference in the cleavage positions on DNA and RNA templates can be attributed to conformational differences of the bound nucleic acids. We suggest that the lower pitch of dsRNA and DNA/RNA hybrids compared with dsDNA permits accommodation of an additional base pair in the region between the primer 3'-end and the Fe2+-dependent cleavage position at the RNase H active site.

Binding Sites↗