Search PubMed⌕ Search

Biomedical subjects

M Schaefer

Publications and source records attributed to M Schaefer.

At least 55 records · Page 3Linked to original sources
PubMed2000

Physical, chemical, and biological evaluations of P760: a new gadolinium complex characterized by a low rate of interstitial diffusion.

An original gadolinium chelate, termed P760, which diffuses through the vascular endothelium but at a much lower rate than nonspecific agents (NSA), is described. P760 is a gadolinium macrocyclic compound based on a DOTA structure that is substituted by hydrophilic bulky groups branched on the amino-carboxylic residues. The molecular weight is 5293, and the molecular volume, measured by light scattering, is 30 times higher (11.5 nm3) than that of gadolinium (Gd)-DOTA (0.38 nm3). The increase in molecular volume and weight has two consequences: a) higher relaxivity (r1; 24.7 mM-1.s-1 compared with 3.4 mM-1.s-1 for Gd-DOTA at 20 Mhz, 37 degrees C); and b) a lengthening of its transport rate through the endothelium. P760 presents a peculiar pharmacokinetic profile: at early times post injection, the blood concentrations are higher than those of Gd-DOTA, but after 20 minutes, the blood concentrations are equal for the two compounds. The body clearances of the products are identical (i.e., glomerular filtration rate). P760 molecules are large enough to have a restricted diffusion through the endothelium but, conversely, small enough to pass freely through the glomerular membrane. This limited extravasation has been observed in rabbits by magnetic resonance angiography or in investigations of tumor permeability. Further experimental imaging studies are needed to define the clinical interest of such properties.

Animals↗
PubMed2000

Transient receptor potential channels as molecular substrates of receptor-mediated cation entry.

Calcium is a versatile multitarget intracellular second messenger in eukaryotic cells. In addition to calcium release from intracellular stores and influx via voltage- or ligand-operated channels, agonist-induced calcium entry constitutes one of the main pathways by which cytosolic calcium is elevated. Receptor-stimulated currents are initiated in response to agonist binding to G-protein-coupled receptors and to receptor tyrosine kinases. Within the past few years our knowledge about the molecular identity of receptor-stimulated channels has expanded substantially. Drosophila melanogaster visual transduction channels associated with the transient receptor potential (trp) and the trp-like (trpl) mutant visual phenotypes were the first members of this category of channels to be identified at the molecular level. Since then an entire mammalian gene family of TRP homologues has been discovered by homology cloning. Only now are we beginning to fully understand the functional roles of TRP channels in mammalian cells. We review recent findings in TRP channel research and discuss the role of these proteins for receptor-activated cation entry.

Animals↗
PubMed2000

Evaluation of the impact of pharmacist's advice giving on the outcomes of self-medication in patients suffering from dyspepsia.

The aim of this study was to investigate the outcomes of self-medication in patients suffering from dyspepsia by comparing changes in the Health related Quality of Life before and after self-medication of dyspeptic disorders. Another study objective was the quantitative and qualitative analysis of the pharmacist's advice-giving to patients with dyspepsia. Therefore the impact of the counselling by the pharmacist on the patient's health outcomes was surveyed and compared between study and control pharmacies. Moreover, the study analysed the influence of a special training on the services provided by the pharmacies with regard to self-medication. A beneficial effect of self-medication on the HRQoL of patients with dyspepsia on a weekly basis has been detected in the study. There is evidence that advice-giving and counselling by the pharmacists in self-medication have a measurable impact on self-medication outcomes. Moreover, the study reveals that patients value the information provided by the pharmacist. Pharmacists gathered the relevant and comprehensive information from the patients having dyspeptic symptoms and provided advice concerning OTC-drugs. Moreover, pharmacists frequently discussed the relevance of factors aggravating dyspeptic disorders such as lifestyle, drinking, smoking, and manner of nutrition with the patient. Training programs and treatment guidelines for the pharmacist seem to obtain a positive effect on his performance. The findings of the study substantiate the value of a pharmacist-controlled self-medication. The study results suggest that the quality of primary health care in self-medication would improve if pharmacists' involvement were even more intense.

Dyspepsia↗
PubMed2000

Steroidal sigma receptor ligands affect signaling pathways in human spermatozoa.

In human spermatozoa, Ca(2+) entry is stimulated by progesterone or prostaglandin E(1) (PGE(1)). The regulation of cation currents by progestins involves sigma receptors, and sigma binding sites are abundant in testis. We examined the effects of sigma ligands on human spermatozoa. Ca(2+) entry induced by progesterone or PGE(1) was not altered by the sigma ligands haloperidol and ditolylguanidine. However, the steroidal sigma ligands RU 3117 and RU 1968 had distinct effects. Stimulation by RU 3117 resulted in activation and homologous desensitization of the sperm progesterone receptor but not of the PGE(1) receptor. Because haloperidol and ditolylguanidine did not affect RU 3117 and progesterone actions in spermatozoa, we conclude that sigma receptors are not involved. However, RU 1968 potently inhibited both the progesterone- and PGE(1)-induced Ca(2+) entry and acrosome reaction. At higher concentrations, RU 1968 also inhibited hormonal Ca(2+) signaling in fibroblasts. Despite suppression of Ca(2+) mobilization, inhibition of phospholipase C by RU 1968 was not observed. Furthermore, RU 1968 did not impair the binding of inositol-1,4,5-trisphosphate to its endoplasmic reticulum receptor. Because RU 1968 preferentially inhibits signaling pathways in spermatozoa, the future development of more selective drugs structurally related to RU 1968 may be a novel approach for pharmacological contraception.

Acrosome Reaction↗
PubMed2000

Increased expression and immunogenicity of sequence-modified human immunodeficiency virus type 1 gag gene.

A major challenge for the next generation of human immunodeficiency virus (HIV) vaccines is the induction of potent, broad, and durable cellular immune responses. The structural protein Gag is highly conserved among the HIV type 1 (HIV-1) gene products and is believed to be an important target for the host cell-mediated immune control of the virus during natural infection. Expression of Gag proteins for vaccines has been hampered by the fact that its expression is dependent on the HIV Rev protein and the Rev-responsive element, the latter located on the env transcript. Moreover, the HIV genome employs suboptimal codon usage, which further contributes to the low expression efficiency of viral proteins. In order to achieve high-level Rev-independent expression of the Gag protein, the sequences encoding HIV-1(SF2) p55(Gag) were modified extensively. First, the viral codons were changed to conform to the codon usage of highly expressed human genes, and second, the residual inhibitory sequences were removed. The resulting modified gag gene showed increases in p55(Gag) protein expression to levels that ranged from 322- to 966-fold greater than that for the native gene after transient expression of 293 cells. Additional constructs that contained the modified gag in combination with modified protease coding sequences were made, and these showed high-level Rev-independent expression of p55(Gag) and its cleavage products. Density gradient analysis and electron microscopy further demonstrated that the modified gag and gag protease genes efficiently expressed particles with the density and morphology expected for HIV virus-like particles. Mice immunized with DNA plasmids containing the modified gag showed Gag-specific antibody and CD8(+) cytotoxic T-lymphocyte (CTL) responses that were inducible at doses of input DNA 100-fold lower than those associated with plasmids containing the native gag gene. Most importantly, four of four rhesus monkeys that received two or three immunizations with modified gag plasmid DNA demonstrated substantial Gag-specific CTL responses. These results highlight the useful application of modified gag expression cassettes for increasing the potency of DNA and other gene delivery vaccine approaches against HIV.

AIDS Vaccines↗
PubMed2000

Hepatic iron overload in aceruloplasminaemia.

We report the case of a 52 year old male with diabetes mellitus and long standing evidence of hepatic iron excess. Initially considered to have haemochromatosis, this patient was reevaluated when hepatic iron stores were found to be unaffected by a prolonged course of weekly phlebotomy. The development of neurological disease prompted diagnostic consideration of aceruloplasminaemia, which we confirmed by demonstration of a novel frameshift mutation in the ceruloplasmin gene. Our inability to resolve the patient's iron overload by regular phlebotomy is consistent with recent animal studies indicating an essential role for ceruloplasmin in cellular iron efflux. Evaluation of this case underscores the clinical relevance of aceruloplasminaemia in the differential diagnosis of hepatic iron overload and provides insight into the pathogenetic mechanisms of hepatocellular iron storage and efflux.

Ceruloplasmin↗
PubMed2000

Relative potency of cellular and humoral immune responses induced by DNA vaccination.

DNA vaccines can prime broad-based immune responses in small animal models. In the present study, we sought to evaluate the relative ability of DNA vaccines to induce humoral and cellular immune responses. Using a DNA vaccine encoding HIV gag in mice, we observed that CD8+ T cell responses were primed more readily than were antibody responses, particularly at low doses of DNA. These CD8+ T cell responses were detected in spleen cells, as well as at local sites such as the lung and draining lymph nodes. The potency of the HIV gag DNA vaccine used was sufficient to prime strong CTL responses in macaques, but only low to undetectable antibody responses. Therefore, DNA vaccines appear able to prime strong, broad CTL but only modest antibody responses. These results may have implications on the development of vaccines against infectious diseases where both CTL and antibody responses are desired, such as HIV.

AIDS Vaccines↗
PubMed2000

Two different mechanisms for activation of cyclic PIP synthase: by a G protein or by protein tyrosine phosphorylation.

The biosynthesis of the functional, endogenous cyclic AMP antagonist, prostaglandylinositol cyclic phosphate (cyclic PIP) is performed by the plasma membrane-bound enzyme cyclic PIP synthase, which combines prostaglandin E (PGE) and activated inositol phosphate (n-IP) to cyclic PIP. The Km values of the enzyme for the substrates PGE and n-IP are in the micromolar range. The plasma membrane-bound synthase is activated by fluoride, by the stable GTP analog GMP-PNP, by protamine or biguanide, by noradrenaline, and by insulin. The activation by protamine or biguanide and fluoride (10 mM) is additive, which may indicate the presence of two different types of enzyme, comparable to phospholipase Cbeta and phospholipase Cgamma. Plasma membrane-bound cyclic PIP synthase is inhibited by the protein tyrosine kinase inhibitor tyrphostin B46 with an IC50 of 1.7 microM. However, the solubilized and gel-filtrated enzyme is no longer inhibited by tyrphostin, indicating that the activity of cyclic PIP synthase is connected with the activity of a membrane-bound protein tyrosine kinase. Cyclic PIP synthase activity of freshly prepared plasma membranes is unstable. Upon freezing and rethawing of liver plasma membranes, this instability is increased about 2-fold. Protein tyrosine phosphatase inhibitors [vanadate, fluoride (50-100 mM)] stabilize the enzyme activity, but protease inhibitors do not, indicating that inactivation of the enzyme is connected with protein tyrosine dephosphorylation. Cyclic PIP synthase is present in all tissues tested, like brain, heart, intestine, kidney, liver, lung, skeletal muscle, spleen, and testis. Apart from liver, cyclic PIP synthase activity in most tissues is rather low, but it can be increased up to 5-fold when protein tyrosine phosphatase inhibitors like vanadate are present in the homogenization buffer. Preincubation of cyclic PIP synthase of liver plasma membranes with the tyrosine kinase src kinase causes a 2-fold increase of cyclic PIP synthase activity, though this is certainly not the physiological role played by src kinase in intact cells. The data indicate that cyclic PIP synthase can be activated by two separate mechanisms: by a G protein or by protein tyrosine phosphorylation.

Adenosine Triphosphate↗
PubMed1999

Bazooka recruits Inscuteable to orient asymmetric cell divisions in Drosophila neuroblasts.

Asymmetric cell divisions can be generated by the segregation of determinants into one of the two daughter cells. In Drosophila, neuroblasts divide asymmetrically along the apical-basal axis shortly after their delamination from the neuroectodermal epithelium. Several proteins, including Numb and Miranda, segregate into the basal daughter cell and are needed for the determination of its correct cell fate. Both the apical-basal orientation of the mitotic spindle and the localization of Numb and Miranda to the basal cell cortex are directed by Inscuteable, a protein that localizes to the apical cell cortex before and during neuroblast mitosis. Here we show that the apical localizaton of Inscuteable requires Bazooka, a protein containing a PDZ domain that is essential for apical-basal polarity in epithelial cells. Bazooka localizes with Inscuteable in neuroblasts and binds to the Inscuteable localization domain in vitro and in vivo. In embryos lacking both maternal and zygotic bazooka function, Inscuteable no longer localizes asymmetrically in neuroblasts and is instead uniformly distributed in the cytoplasm. Mitotic spindles in neuroblasts are misoriented in these embryos, and the proteins Numb and Miranda fail to localize asymmetrically in metaphase. Our results suggest that direct binding to Bazooka mediates the asymmetric localization of Inscuteable and connects the asymmetric division of neuroblasts to the axis of epithelial apical-basal polarity.

Animals↗
PubMed1999

Interaction of the copper chaperone HAH1 with the Wilson disease protein is essential for copper homeostasis.

The delivery of copper to specific sites within the cell is mediated by distinct intracellular carrier proteins termed copper chaperones. Previous studies in Saccharomyces cerevisiae suggested that the human copper chaperone HAH1 may play a role in copper trafficking to the secretory pathway of the cell. In this current study, HAH1 was detected in lysates from multiple human cell lines and tissues as a single-chain protein distributed throughout the cytoplasm and nucleus. Studies with a glutathione S-transferase-HAH1 fusion protein demonstrated direct protein-protein interaction between HAH1 and the Wilson disease protein, which required the cysteine copper ligands in the amino terminus of HAH1. Consistent with these in vitro observations, coimmunoprecipitation experiments revealed that HAH1 interacts with both the Wilson and Menkes proteins in vivo and that this interaction depends on available copper. When these studies were repeated utilizing three disease-associated mutations in the amino terminus of the Wilson protein, a marked diminution in HAH1 interaction was observed, suggesting that impaired copper delivery by HAH1 constitutes the molecular basis of Wilson disease in patients harboring these mutations. Taken together, these data provide a mechanism for the function of HAH1 as a copper chaperone in mammalian cells and demonstrate that this protein is essential for copper homeostasis.

Adenosine Triphosphatases↗
PubMed1999

Direct activation of human TRPC6 and TRPC3 channels by diacylglycerol.

Eukaryotic cells respond to many hormones and neurotransmitters with increased activity of the enzyme phospholipase C and a subsequent rise in the concentration of intracellular free calcium ([Ca2+]i). The increase in [Ca2+]i occurs as a result of the release of Ca2+ from intracellular stores and an influx of Ca2+ through the plasma membrane; this influx of Ca2+ may or may not be store-dependent. Drosophila transient receptor potential (TRP) proteins and some mammalian homologues (TRPC proteins) are thought to mediate capacitative Ca2+ entry. Here we describe the molecular mechanism of store-depletion-independent activation of a subfamily of mammalian TRPC channels. We find that hTRPC6 is a non-selective cation channel that is activated by diacylglycerol in a membrane-delimited fashion, independently of protein kinases C activated by diacylglycerol. Although hTRPC3, the closest structural relative of hTRPC6, is activated in the same way, TRPCs 1, 4 and 5 and the vanilloid receptor subtype 1 are unresponsive to the lipid mediator. Thus, hTRPC3 and hTRPC6 represent the first members of a new functional family of second-messenger-operated cation channels, which are activated by diacylglycerol.

Animals↗
PubMed1999

P760 and P775: MRI contrast agents characterized by new pharmacokinetic properties.

RATIONALE AND OBJECTIVES: In this paper we discuss novel MR imaging blood pool agents characterized by new pharmacokinetic properties. METHODS: The pharmacokinetics of the products were studied in a rabbit model. The potential of these new products was demonstrated in experimental MR imaging. RESULTS AND CONCLUSION: Three main classes of blood pool agents have been defined and characterized according to their pharmacokinetic properties: low diffusion agents, rapid clearance blood pool agents, slow clearance blood pool agents. Each kind of blood pool agent is expected to have different diagnostic applications.

Animals↗
PubMed1999

Localization of the Wilson's disease protein in human liver.

BACKGROUND & AIMS: Wilson's disease is an autosomal-recessive disorder of copper metabolism that results from the absence or dysfunction of a copper-transporting P-type adenosine triphosphatase that leads to impaired biliary copper excretion and disturbed holoceruloplasmin synthesis. To gain further insight into the role of the Wilson's disease protein in hepatic copper handling, its localization in human liver was investigated. METHODS: By use of a specific antibody, localization of the Wilson's disease protein was studied in liver membrane fractions and liver sections by immunoblotting, immunohistochemistry, and double-label confocal scanning laser microscopy. RESULTS: The 165-kilodalton protein, found by immunoblotting, was most abundant mainly in isolated plasma membrane fractions enriched in canalicular domains. Immunohistochemistry revealed intracellular punctuate staining of hepatocytes in certain regions of the liver, whereas a canalicular membrane staining pattern was observed in other regions. Double-labeling studies showed that in the latter regions the transporter is present mainly in vesicular structures just underneath the canalicular membrane that are positive for markers of the trans-Golgi network. A weak staining of the canalicular membrane, identified by staining for P-glycoprotein, was observed. CONCLUSIONS: These results show that in human liver the Wilson's disease protein is predominantly present in trans-Golgi vesicles in the pericanalicular area, whereas relatively small amounts of the protein appear to localize to the canalicular membrane, consistent with a dual function of the protein in holoceruloplasmin synthesis and biliary copper excretion.

Adenosine Triphosphatases↗
PubMed1999

Macromolecular contrast agents for magnetic resonance imaging. Influence of polymer content in ligand on the paramagnetic properties.

Macromolecular conjugates of Gd3+-diethylenetriaminepentaacetic acid with dextran were synthesized from dextran 40 (about 40 kg/mol). Diethylenetriaminepentaacetic acid (DTPA) was coupled to aminated dextran by means of a watersoluble carbodiimide and macromolecular conjugates containing DTPA ratios as high as 1.25 mmol/g of polymer were obtained. First, it was found that the polymer had a favourable influence on relaxivity, as at 20 MHz, the r1 longitudinal relaxivity of the Gd3+-complexed macromolecular conjugates was 2 to 4 times as great as that of free GdDTPA2-, depending on the DTPA content. Second, r1 greatly increased with the increase in the conjugate DTPA content, from 7.4 to 15.9 mM-1s-1 for an increase in the DTPA content from 0.36 to 0.96 mmol/g. Further increase in the ligand content had no more effect on relaxivity.

Chemical Phenomena↗
PubMed1999

Certification of highly complex safety-related systems.

The BIA has now 15 years of experience with the certification of complex electronic systems for safety-related applications in the machinery sector. Using the example of machining centres this presentation will show the systematic procedure for verifying and validating control systems using Application Specific Integrated Circuits (ASICs) and microcomputers for safety functions. One section will describe the control structure of machining centres with control systems using "integrated safety." A diverse redundant architecture combined with crossmonitoring and forced dynamization is explained. In the main section the steps of the systematic certification procedure are explained showing some results of the certification of drilling machines. Specification reviews, design reviews with test case specification, statistical analysis, and walk-throughs are the analytical measures in the testing process. Systematic tests based on the test case specification, Electro Magnetic Interference (EMI), and environmental testing, and site acceptance tests on the machines are the testing measures for validation. A complex software driven system is always undergoing modification. Most of the changes are not safety-relevant but this has to be proven. A systematic procedure for certifying software modifications is presented in the last section of the paper.

Electronics↗
PubMed1999

Genetic disorders of membrane transport. IV. Wilson's disease and Menkes disease.

Copper is an essential transition metal that permits the facile transfer of electrons in a series of critical biochemical pathways. Menkes disease and Wilson's disease are inherited disorders of copper metabolism resulting from the absence or dysfunction of homologous copper-transporting ATPases that reside in the trans-Golgi network of all cells. Despite striking differences in the clinical presentation of these two diseases, the respective ATPases function in precisely the same manner within the cell and the unique clinical features of each disease are entirely the result of the tissue-specific expression of each protein. Elucidation of the basic defect in these rare genetic disorders has provided a valuable heuristic paradigm for understanding the mechanisms of cellular copper homeostasis.

Adenosine Triphosphatases↗