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Biomedical subjects

J Meyer

Publications and source records attributed to J Meyer.

At least 307 records · Page 17Linked to original sources
PubMed1997

Atomic resolution (0.94 A) structure of Clostridium acidurici ferredoxin. Detailed geometry of [4Fe-4S] clusters in a protein.

The crystal structure of the 2[4Fe-4S] ferredoxin from Clostridium acidurici has been solved using X-ray diffraction data extending to atomic resolution, 0.94 A, recorded at 100 K. The model was refined with anisotropic representation of atomic displacement parameters for all non-hydrogen atoms and with hydrogens riding on their parent atoms. Stereochemical restraints were applied to the protein chain but not to the iron-sulfur clusters. The final R factor is 10.03 % for all data. Inversion of the final least-squares matrix allowed direct estimation of the errors of individual parameters. The estimated errors in positions for protein main chain atoms are below 0.02 A and about 0.003 A for the heavier [4Fe-4S] cluster atoms. Significant differences between the stereochemistry of the two clusters and distortion of both of them from ideal Td tetrahedral symmetry can be defined in detail at this level of accuracy. Regions of alternative conformations include not only protein side chains but also two regions of the main chain. One such region is the loop of residues 25-29, which was highly disordered in the room temperature structure.

Bacterial Proteins↗
PubMed1997

Assembly of a [2Fe-2S]2+ cluster in a molecular variant of Clostridium pasteurianum rubredoxin.

The rubredoxin from Clostridium pasteurianum contains a single iron atom bound to the polypeptide chain by cysteines 6, 9, 39, and 42. The C42A variant of this protein has been prepared by site-directed mutagenesis and heterologous expression of the gene in Escherichia coli. The mutated protein was found to contain an unexpected chromophore that has been characterized by a variety of techniques. UV-visible absorption and resonance Raman spectra were strongly reminiscent of those of [2Fe-2S] proteins. Mössbauer spectra of the oxidized chromophore isolated in oxygen-free conditions indicated low-temperature diamagnetism resulting from antiferromagnetically coupled high-spin ferric ions. Analysis of X-ray absorption fine structure spectra yielded an Fe-Fe distance of 2.68 A. Colorimetric assays of iron and inorganic sulfide showed that the two elements are present in a 1:1 ratio. Electrospray-ionization mass spectra displayed a major component at M = 6190 Da, i.e. the molecular mass of the C42A apoprotein plus two atomic masses of iron and two atomic masses of sulfur. Taken together, these data show that a mere point mutation allows the stabilization of a binuclear [2Fe-2S] cluster in a protein that normally accommodates a mononuclear Fe(Scys)4 site. Assembly of a [2Fe-2S] cluster may occur because rubredoxin assumes a similar fold around its metal center as the [2Fe-2S] Rieske protein. Alternatively, a more extensive structural rearrangement of the polypeptide chain of the C42A rubredoxin variant may be considered as well.

Binding Sites↗
PubMed1997

The HIV transactivator TAT binds to the CDK-activating kinase and activates the phosphorylation of the carboxy-terminal domain of RNA polymerase II.

The human immunodeficiency virus encodes the transcriptional transactivator Tat, which binds to the transactivation response (TAR) RNA stem-loop in the viral long terminal repeat (LTR) and increases rates of elongation rather than initiation of transcription by RNA polymerase II (Pol II). In this study, we demonstrate that Tat binds directly to the cyclin-dependent kinase 7 (CDK7), which leads to productive interactions between Tat and the CDK-activating kinase (CAK) complex and between Tat and TFIIH. Tat activates the phosphorylation of the carboxy-terminal domain (CTD) of Pol II by CAK in vitro. The ability of CAK to phosphorylate the CTD can be inhibited specifically by a CDK7 pseudosubstrate peptide that also inhibits transcriptional activation by Tat in vitro and in vivo. We conclude that the phosphorylation of the CTD by CAK is essential for Tat transactivation. Our data identify a cellular protein that interacts with the activation domain of Tat, demonstrate that this interaction is critical for the function of Tat, and provide a mechanism by which Tat increases the processivity of Pol II.

Animals↗
PubMed1997

Specific interaction of the [2Fe-2S] ferredoxin from Clostridium pasteurianum with the nitrogenase MoFe protein.

Putative physiological partners of the [2Fe-2S] ferredoxin from Clostridium pasteurianum have been searched by running crude soluble extracts of this bacterium through an affinity column to which the [2Fe-2S] ferredoxin had been covalently bound. Subsequent washing of the column with buffers of increasing ionic strength revealed a strong and specific interaction of the ferredoxin with the MoFe protein of nitrogenase. This interaction was further investigated by performing cross-linking experiments with mixtures of the two purified proteins in solution. Analysis of the reactions by SDS-polyacrylamide gel electrophoresis evidenced only two covalently linked products. These were identified by N-terminal sequencing as the alpha and beta subunits of the MoFe protein, each cross-linked to a single polypeptide chain of the ferredoxin. This result, taking into account the dimeric structure of the ferredoxin in solution, strongly suggests an interaction of the ferredoxin with the MoFe protein at a site contributed to by both subunits of the MoFe protein. The ionic strength dependence of the interaction evidenced by affinity chromatography was confirmed in the cross-linking reactions, and its specificity was assessed by showing that no cross-linking occurred when the [2Fe-2S] C. pasteurianum ferredoxin was denatured or replaced by spinach ferredoxin or by clostridial rubredoxin, or when the MoFe protein from C. pasteurianum was either inactivated or replaced by its counterpart from Azotobacter vinelandii. It has also been observed that the ferredoxin inhibits cross-linking between the nitrogenase Fe protein and the MoFe protein, which suggests overlapping binding sites of the ferredoxin and of the Fe protein on the MoFe protein. Cross-linking experiments implementing a number of molecular variants of the [2Fe-2S] C. pasteurianum ferredoxin demonstrated that glutamate residues 31, 34, and 38 are important contributors to the interaction with the MoFe protein.

Chromatography, Affinity↗
PubMed1997

Haemodynamic effects of dopexamine and nitric oxide synthase inhibition in healthy and endotoxaemic sheep.

Chronically instrumented awake healthy sheep (n = 6) received the synthetic catecholamine, dopexamine, during or without a background infusion of the nitric oxide synthase inhibitor. L-nitro-arginine-methylester (L-NAME). Three days later, hypotensive-hyperdynamic circulation was induced and maintained by continuous infusion of Salmonella typhosa endotoxin (10 ng/kg per min). After 24 h of continuous endotoxin infusion, the dopexamine L-NAME protocol was repeated. In healthy and endotoxaemic animals with and without nitric oxide synthase inhibition dopexamine caused the same haemodynamic changes: heart rate and cardiac output increased, mean arterial pressure and systemic vascular resistance decreased. L-NAME infusion induced normalisation of the hypotonic-hyperdynamic circulation in endotoxaemic animals. Dopexamine reduced some adverse effects of L-NAME treatment, like increased pulmonary vascular resistance and decreased oxygen delivery. In conclusion the haemodynamic effects of dopexamine are independent of the amount of nitric oxide production. Dopexamine may attenuate some of the adverse effects of nitric oxide synthase inhibition.

Adrenergic beta-Agonists↗
PubMed1997

Prostacyclin receptor desensitization is a reversible phenomenon in human platelets.

BACKGROUND: Long-term exposure of platelets to endogenous or exogenous prostacyclin or its analogues might result in desensitization of the platelet prostacyclin receptor in vitro and in vivo accompanied by a loss in receptor density on the platelet surface and a reduced sensitivity toward the inhibitory effects of prostacyclins. However, the reversibility of this process in platelets has not yet been investigated. METHODS AND RESULTS: Human platelets desensitized by the chemically stable prostacyclin analogue iloprost showed a significant reduction in [3H]-iloprost binding sites that was reversed by saponin permeabilization. This indicates functionally active internalized prostacyclin receptors. To assess whether the internalized prostacyclin receptors recycle to the cell surface after withdrawal of the agonist, iloprost sensitivity and prostacyclin receptor binding properties of iloprost (30 nmol/L, 2 hours) desensitized platelets incubated in iloprost-free autologous plasma were investigated. While desensitized platelets showed a significant increase in IC50 for iloprost inhibition of thrombin-induced platelet aggregation, serotonin release, and p-selectin expression and a reduced iloprost-stimulated cAMP formation, platelet iloprost sensitivity was restored 3 hours after iloprost withdrawal. In addition, the significant reduction in Bmax and the increase in K(D) of prostacyclin receptors in desensitized platelets as revealed by [3H]-iloprost binding studies also returned to the initial values. CONCLUSIONS: These results indicate that prostacyclin receptors internalized during short-term desensitization are not degraded but can be recycled rapidly to the platelet surface in a functionally active form after withdrawal of the agonist.

Blood Platelets↗
PubMed1997

Regulation of glucose transport, and glucose transporters expression and trafficking in the heart: studies in cardiac myocytes.

Cardiac muscle is characterized by a high rate of glucose consumption. In the absence of insulin, glucose transport into cardiomyocytes limits the rate of glucose utilization and therefore it is important to understand the regulation of glucose transporters. Cardiac muscle cells express 2 distinct glucose transporters, GLUT4 and GLUT1; although GLUT4 is quantitatively the more important glucose transporter expressed in heart, GLUT1 is also expressed at a substantial level. In isolated rat cardiomyocytes, insulin acutely stimulates glucose transport and translocates both GLUT4 and GLUT1 from an intracellular site to the cell surface. Recent evidence indicates the existence of at least 2 distinct intracellular membrane populations enriched in GLUT4 with a different protein composition. Elucidation of the intracellular location of these 2 GLUT4 vesicle pools in cardiac myocytes, their role in GLUT4 trafficking, and their relation to insulin-induced GLUT4 translocation needs to be addressed.

Animals↗
PubMed1997

Promoting action research in healthcare settings.

To coincide with the first annual Action Research conference taking place in London this week, this article promotes the idea of using action research for health care exploration and change. It also introduces the work of the Healthcare Subgroup of the Collaborative Action Research Network, the group's aims and how practitioners can make contact and be involved with the development. The article also describes the outcomes of a research study that underpin the philosophy, process and potential benefits of the network as a means to promote practice development and change in client care.

Clinical Nursing Research↗
PubMed1997

Feasibility and sensing thresholds of temporary single-lead VDD pacing in intensive care.

Long-term pacemaker treatment of patients with a high-degree atrioventricular (AV) block routinely uses AV synchronous pacing because of its hemodynamic advantages compared with VVI pacing. In cases of temporary pacing, however, the limitations inherent in attempting to reliably position a temporary atrial lead generally influences the use of VVI pacing. We therefore tested the use of temporary single-lead VDD pacing, making AV pacing possible with only 1 lead, in 22 patients requiring temporary pacing due to a high-degree AV block. AV synchronous stimulation was achieved in all patients using a quadripolar lead with an atrial dipole with the atrial rings spaced 30 mm apart. During follow-up (14.1 +/- 12.5 hours) intermittent undersensing was detected in 4 of the 22 patients. We initially evaluated the atrial sensing threshold by decreasing the pacemaker device sensitivity stepwise in all patients (1.54 +/- 1.08 mV; n = 22). In 15 patients an intra-atrial electrocardiogram was recorded. During the breathing cycle, the maximum P-wave amplitude was significantly different from the minimum P-wave amplitude (2.19 +/- 1.00 mV vs 1.25 +/- 0.65 mV, p < 0.005). The atrial signal detected by the single lead was further analyzed in 10 of 15 patients using various filters. A mean signal loss of 45% was observed, increasing the lower bandpass frequency from 0.1 to 40 Hz (1.73 +/- 0.71 mV vs 0.92 +/- 0.51 mV, p < 0.02). Lowering the upper filter range from 1,000 Hz down to 100 Hz did not significantly influence the atrial signal (1.73 +/- 0.71 mV vs 1.61 +/- 0.75 mV, NS). Single-lead VDD pacing, even on a temporary basis, is a reliable means of achieving AV synchronous pacing. Due to the floating atrial dipole, the system is characterized by a high degree of variability in the atrial signal with intermittent lower values. A significant signal loss must be expected when a lower bandpass frequency of 40 Hz is used.

Aged↗
PubMed1997

High-performance liquid chromatographic determination of nitric oxide synthase-related arginine derivatives in vitro and in vivo.

In this paper we present a sensitive and reproducible method for the extraction and quantification of the nitric oxide (NO) synthase (NOS)-related basic amino acids L-hydroxyarginine (L-NHA), L-arginine (L-Arg), L-monomethylarginine (L-NMA), and L-dimethylarginine (L-NDA) in human serum samples by high-performance liquid chromatography (HPLC) analysis. We demonstrate that the serum level of L-NHA can be used as a sensitive and highly specific index of a systemic increase in NOS activity in vivo whose serum concentration, unlike that of the NO degradation products nitrite and/or nitrate, is not influenced by dietary intake. First, we measured L-NHA formation by a recombinant NOS preparation and by lipopolysaccharide-stimulated alveolar macrophages to demonstrate that this amino acid is produced by NOS in vitro. HPLC determination of L-NHA in human serum, however, proved to be difficult due to the presence of amino acids interfering with its detection. Therefore, we developed a clean-up procedure for the extraction of basic amino acids from these serum samples by using a cation-exchange cartridge. The isolated amino acids were subjected to precolumn derivatization with o-pthaldialdehyde and analyzed using a short reversed-phase column which allowed the baseline separation of L-NHA, L-Arg, L-NMA, and L-NDA within 16 min. By using this technique, the average concentrations of L-NHA, L-Arg, L-NMA, and L-NDA in the serum of healthy human subjects were determined to be 9.1, 96.1, 0.1, and 0.4 microM, respectively.

Animals↗
PubMed1997

Comparison of saruplase and alteplase in acute myocardial infarction. SESAM Study Group. The Study in Europe with Saruplase and Alteplase in Myocardial Infarction.

Four hundred seventy-three patients with acute myocardial infarction (AMI) were treated with either saruplase (80 mg/hour, n = 236) or alteplase (100 mg every 3 hours, n = 237). Comedication included heparin and acetylsalicylic acid. Angiography was performed at 45 and 60 minutes after the start of thrombolytic therapy. When flow was insufficient, angiography was repeated at 90 minutes. Coronary angioplasty was then performed if Thrombolysis In Myocardial Infarction (TIMI) trial 0 to 1 flow was seen. Control angiography was at 24 to 40 hours. Baseline characteristics were similar. Angiography showed comparable and remarkably high early patency rates (TIMI 2 or 3 flow) in both treatment groups: at 45 minutes, 74.6% versus 68.9% (p = 0.22); and at 60 minutes 79.9% versus 75.3% (p = 0.26). Patency rates at 90 minutes before additional interventions were also comparable (79.9% and 81.4%). Angiographic reocclusion rates were not significantly different: 1.2% versus 2.4% (p = 0.68). After rescue angioplasty, angiographic reocclusion rates of 22.0% and 15.0% were observed. Safety data were similar for both groups. Thus, (1) early patency rates were high for saruplase and alteplase treatment, (2) reocclusion rates for both drugs were remarkably low, and (3) complication rates were similar. Thus, saruplase seems to be as safe and effective as alteplase.

Aged↗
PubMed1997

Intracoronary application of C1 esterase inhibitor improves cardiac function and reduces myocardial necrosis in an experimental model of ischemia and reperfusion.

BACKGROUND: Myocardial injury from ischemia can be aggravated by reperfusion of the jeopardized area. The precise underlying mechanisms have not been clearly defined, but proinflammatory events, including complement activation, leukocyte adhesion, and infiltration and release of diverse mediators, probably play important roles. The present study addresses the possibility of reducing reperfusion damage by the application of C1 esterase inhibitor (C1-INH). METHODS AND RESULTS: Cardioprotection by C1-INH 20 IU/kg IC was examined in a pig model with 60 minutes of coronary occlusion, followed by 120 minutes of reperfusion. C1-INH was administered during the first 5 minutes of coronary reperfusion Compared with the NaCl controls, C1-INH reduced myocardial injury (48.8 +/- 7.8% versus 73.4 +/- 4.0% necrosis of area at risk, P < or = .018). C1-INH treatment significantly reduced circulating C3a and slightly attenuated C5a plasma concentrations. Myocardial protection was accompanied by reduced plasma concentration of creatine kinase and troponin-T. C1-INH had no effect on global hemodynamic parameters, but local myocardial contractility was markedly improved in the ischemic zone. In the short-axis view, 137 degrees of the anteroseptal region showed significantly improved wall motion at early and 29 degrees at late reperfusion with C1-INH treatment. CONCLUSIONS: C1-INH significantly protects ischemic tissue from reperfusion damage, reduces myocardial necrosis, and improves local cardiac function.

Anaphylatoxins↗
PubMed1997

Effect of antibiotic treatment on vegetation size and complication rate in infective endocarditis.

BACKGROUND: Infective endocarditis is associated with significant morbidity and mortality, with valvular destruction, and with congestive heart failure. Embolic events are more common in patients with echocardiographically discernible vegetations, especially when vegetations are > 10 mm in diameter. HYPOTHESIS: The objective of the study was to follow vegetation morphology during native valve endocarditis, to compare it with the clinical course and antibiotic treatment chosen, and to evaluate whether the impact on vegetation size and complication rate of antibiotic regimens differed in patients with positive and negative blood cultures. METHODS: The effect of different antibiotic regimes on vegetation size monitored by using transesophageal echocardiography was evaluated in 183 patients with echocardiographic evidence of infective endocarditis. A total of 223 vegetations attached to the aortic or mitral valves were detected using the transesophageal approach. The patients were followed for a mean of 76 weeks and underwent a minimum of two consecutive transesophageal echocardiographic examinations. RESULTS: Treatment with different kinds of antibiotics corresponded with significant differences in vegetation size; vancomycin-associated treatment was related to a 45% reduction, ampicillin to a 19% reduction, penicillin to a 5% reduction, penicillase-resistant drugs to a 15% increase, and cephalosporin to a 40% increase in vegetation size. Multivariate analysis showed that penicillin, cephalosporin, and penicillase-resistant drug treatments were associated with an increased embolic risk, vancomycin treatment with abscess formation, and cephalosporin medication with increased mortality. Plotting changes in vegetation size against the incidence of embolism and mortality, linear regression analysis suggested a 40-50% reduction in vegetation size, thereby greatly reducing the risk of embolism and mortality. CONCLUSION: Our study shows that different antibiotics have different effects on vegetation size. The highest complication rate was observed when vegetations significantly increased in size during antibiotic treatment. Especially in culture-negative patients, monitoring vegetation size by means of transesophageal echocardiography may prove to be useful for estimating the efficacy of antibiotic treatment.

Anti-Bacterial Agents↗
PubMed1997

The 5-HT transporter gene-linked polymorphic region (5-HTTLPR) in evolutionary perspective: alternative biallelic variation in rhesus monkeys. Rapid communication.

By conferring allele-specific transcriptional activity on the 5-HT transporter gene promoter in humans, the 5-HT transporter gene-linked polymorphic region (5-HTTLPR) influences a constellation of personality traits related to anxiety and increases the risk for neurodevelopmental, neurodegenerative, and psychiatric disorders. Here we have analyzed the presence and variability of the 5-HTTLPR in several species of primates including humans, and other mammals. PCR, Southern blot, and sequence analyses of the 5-HT transporter gene's 5'-flanking region in different mammalian species confirmed the presence of the 5-HTTLPR in platyrrhini and catarrhini (hominoids, cercopithecoids) but not in prosimian primates and other mammals. Since the 5-HTTLPR is unique to humans and simian primates, a progenitor 5-HTTLPR sequence may have been introduced into the genome some 40 Mio, years ago. In humans the majority of alleles are composed of either 14 or 16 repeat elements, while alleles with 18 or 20 repeat elements are rare. In contrast, great apes including orang-utan, gorilla, and chimpanzee display a high prevalence of alleles with 18 and 20 repeat elements. In hominoids all alleles originate from variation at a single locus (polymorphic locus 1). In the 5-HTTLPR of rhesus monkeys (rh5-HTTLPR) we found an alternative locus for length variation (polymorphic locus 2) generated by a 21 bp insertion/deletion event. The existence of a distinct biallelic variation of the 5-HTTLPR in rhesus monkeys but similar allele and genotype frequencies in this species and humans supports the notion that there may be a relationship between functional 5-HT transporter expression, anxiety-related traits, and the complexity of socialization in human and non-human primate populations.

Alleles↗