Treatment of neonatal hyperammonaemia.
Explore the source record for details and available documents.
Glucagon↗
Biomedical subjects
C Bachmann
Publications and source records attributed to C Bachmann.
Differential expression of the cationic amino acid transporter 2(B) in the adult rat brain.
L-Arginine is a substrate for the synthesis of proteins, nitric oxide (NO), creatine, urea, proline, glutamate, polyamines and agmatine. In the central nervous system (CNS), arginine is extracted from the blood and exchanged by cells through carriers called cationic amino acid transporters (CAT) and belonging to the so-called system y+. In order to better understand the arginine transport in the CNS, we studied in detail the regional distribution of the cells expressing the CAT2(B) transcript in the adult rat brain by non-radioisotopic in situ hybridization. We show that CAT2(B) is expressed in neurons and oligodendrocytes throughout the brain, but is not detected in astrocytes. The pattern of localization of CAT2(B) in the normal adult rat brain fits closely that of CRT1, a specific creatine transporter. Our study demonstrates that the in vivo expression of CAT2(B) differs from that reported in vitro, implying that local cellular interactions should be taken into account in studies of gene regulation of the CAT2(B) gene. Our work suggests that CAT2(B) may play a role in case of increased NO production as well as arginine or creatine deficiency in the brain.
Age Factors↗
Endogenous synthesis and transport of creatine in the rat brain: an in situ hybridization study.
Creatine is synthesized from arginine by L-arginine:glycine amidinotransferase (AGAT) and S-adenosyl-L-methionine:N-guanidinoacetate methyltransferase (GAMT) and can be taken up by cells by creatine transporters (CRT). While creatine is mainly synthesized by the liver and the kidney, most of other tissues, including the brain, also express AGAT and GAMT. There is evidence that the permeability of the blood-brain barrier (BBB) for creatine is limited, suggesting that the brain is dependent on its own creatine synthesis. In order to better understand creatine synthesis and transport in the central nervous system (CNS), we studied the regional distribution of cells expressing AGAT, GAMT and the creatine transporter CRT1 in the adult rat brain by non-radioisotopic in situ hybridization. AGAT and GAMT presented an ubiquitous neuronal and glial expression, whereas CRT1 was present in neurons and oligodendrocytes throughout the brain, but not in astrocytes. This indicates that all cells in the CNS can synthesize creatine from arginine. The absence of expression of CRT1 in astrocytes and particularly in those contacting capillary endothelial cells (BBB) reinforces the idea that under normal conditions the creatine used by the brain is synthesized mainly in the CNS. Furthermore, the expression of CRT1 by neurons and oligodendrocytes indicates that creatine trafficking is possible in those brain areas of main creatine consumption.
Amidinotransferases↗
Integrating particle image velocimetry and laser Doppler velocimetry measurements of the regurgitant flow field past mechanical heart valves.
This study investigates the transient regurgitant flow downstream of a prosthetic heart valve using both laser Doppler velocimetry (LDV) and particle image velocimetry (PIV). Until now, LDV has been the more commonly used tool in investigating the flow characteristics associated with mechanical heart valves. The LDV technique allows point-by-point velocity measurements and provides enough information about the temporal variations in the flow. The main drawback of this technique is the time consuming nature of the data acquisition process in order to assess an entire flow field area. The PIV technique, on the other hand, allows measurement of the entire flow field in space in a plane at a given instant. In this study, PIV with spatial resolution of 0 (1 mm) and LDV with a temporal resolution of 0 (1 ms) were used to measure the regurgitant flow proximal to the Björk-Shiley monostrut (BSM) valve in the mitral position. With PIV, the ability to measure 2 velocity components over an entire plane simultaneously provides a very different insight into the flow field compared to a more traditional point-to-point technique like LDV. In this study, a picture of the effects of occluder motion on the fluid flow in the atrial chamber is interpreted using an integration of PIV and LDV measurements. Specifically, fluid velocities in excess of 3.0 m/s were recorded in the pressure-driven jet during valve closure, and a 1.5 m/s sustained regurgitant jet was observed on the minor orifice side. Additionally, the effects of the impact and subsequent rebound of the occluder on the flow also were clearly recorded in spatial and temporal detail by the PIV and LDV measurements, respectively. The PIV results provide a visually intuitive way of interpreting the flow while the LDV data explore the temporal variations and trends in detail. This analysis is an integrated flow description of the effects of valve closure and leakage on the pulsatile regurgitation flow field past a tilting-disc mechanical heart valve (MHV). It further reinforces the hypothesis that the planar flow visualization techniques, when integrated with traditional point-to point techniques, provide significantly more insight into the complex pulsatile flow past MHVs.
Blood Flow Velocity↗
Density functional study of pi facial selectivity in diels-alder reactions
Explore the source record for details and available documents.
Journal Article↗
The brain mineralocorticoid receptor: greedy for ligand, mysterious in function.
Glucocorticoids exert their regulatory effects on the hypothalamic-pituitary-adrenocortical axis via two types of corticosteroid receptors: the glucocorticoid receptor and the mineralocorticoid receptor. Whereas the glucocorticoid receptor has a broad distribution in the brain, highest levels of mineralocorticoid receptor are found in the hippocampus. Based on the differential occupancy profile by endogenous glucocorticoids, glucocorticoid receptors are thought to mediate negative feedback signals of elevated glucocorticoid levels, whereas mineralocorticoid receptors control the inhibitory tone of the hippocampus on hypothalamic-pituitary-adrenocortical axis activity. Dysfunction of mineralocorticoid receptors and glucocorticoid receptors are thought to be implicated in stress-related psychiatric diseases such as major depression. Because of its intriguing features, we focus in this review on the mineralocorticoid receptor and provide data which reveal novel aspects of the pharmacology and physiology of mineralocorticoid receptors. Newly obtained results are presented, which help to solve the paradox of why dexamethasone binds with high affinity to mineralocorticoid receptors in vitro, yet binds poorly in vivo. Until recently, mineralocorticoid receptor protein and mRNA levels could only be routinely studied with in vitro cytosol binding assays, in vitro and in vivo receptor autoradiography, Northern blot analysis, and in situ hybridization. These methods are unfortunately hampered by several flaws, such as the necessity of adrenalectomy, no or poor neuroanatomical resolution, the fact that mRNA does not provide the same information as protein, or combinations of these factors. We present immunohistochemical data on mineralocorticoid receptors in the brain obtained by using commercially available antibodies, which alleviate many of these shortcomings. Furthermore, an in vivo microdialysis method is presented which allows the assessment of free corticosterone levels in the brain, which is critical for the study of the pharmacological basis of mineralocorticoid receptor (and glucocorticoid receptor) function. Finally, a novel aspect of the regulation of mineralocorticoid receptors is described which provides evidence that this receptor system is dynamically regulated. In conjunction with previously reported effects of antidepressants, these results have initiated a new concept on the cause of the hypothalamic-pituitary-adrenocortical axis disturbances often seen in stress-related psychiatric disorders such as major depression.
Animals↗
Amyloid and non-amyloid forms of 5q31-linked corneal dystrophy resulting from kerato-epithelin mutations at Arg-124 are associated with abnormal turnover of the protein.
Mutations in kerato-epithelin are responsible for a group of hereditary cornea-specific deposition diseases, 5q31-linked corneal dystrophies. These conditions are characterized by progressive accumulation of protein deposits of different ultrastructure. Herein, we studied the corneas with mutations at kerato-epithelin residue Arg-124 resulting in amyloid (R124C), non-amyloid (R124L), and a mixed pattern of deposition (R124H). We found that aggregated kerato-epithelin comprised all types of pathological deposits. Each mutation was associated with characteristic changes of protein turnover in corneal tissue. Amyloidogenesis in R124C corneas was accompanied by the accumulation of N-terminal kerato-epithelin fragments, whereby species of 44 kDa were the major constituents of amyloid fibrils. R124H corneas with prevailing non-amyloid inclusions showed accumulation of a new 66-kDa species altogether with the full-size 68-kDa form. Finally, in R124L cornea with non amyloid deposits, we found only the accumulation of the 68-kDa form. Two-dimensional gels revealed mutation-specific changes in the processing of the full-size protein in all affected corneas. It appears that substitutions at the same residue (Arg-124) result in cornea-specific deposition of kerato-epithelin via distinct aggregation pathways each involving altered turnover of the protein in corneal tissue.
Amyloid↗
Assay of D-lactate in urine of infants and children with reference values taking into account data below detection limit.
Accumulation of D-lactic acid produced by intestinal bacteria such as streptococci and lactobacilli has been extensively studied in ruminants [1-4]. In humans an increased production of D-lactate by intestinal bacteria under pathological conditions such as the short bowel syndrome can cause metabolic acidosis [5-8]. Since the lactate assays routinely used only measure L-lactate we developed a sensitive method of D-lactate quantification and established reference values in spot urines of infants and children (0 to 4 years of age). The enzymatic method with fluorimetric quantification of NADH is linear up to 2 mmol/l. It has a detection limit of 3.4 micromol/l. Among structurally related organic acids an interference was found only for L-lactate and DL-2-hydroxybutyrate at concentrations which are way beyond their physiological excretion. One hundred and sixty five spot urines of healthy Swiss (S), Austrian (A), German (G) and Chilean (CHI) infants aged from 0 to 4 years were analyzed. The distribution of the data is close to a lognormal one. Values below the detection limit were simulated and age groups were formed. In all populations D-lactate excretion was found highest during the first year of life; it declines with age during infancy and remains stable from 2.5 to 4 years of age. We show that D-lactate is excreted physiologically by healthy infants and children below 4 years of age and present reference values for D-lactate excretion which show some differences between the populations tested.
Aging↗
Fluid dynamics of a pediatric ventricular assist device.
The number of pediatric patients requiring some form of mechanical circulatory assistance is growing throughout the world because of new surgical procedures and the success of pediatric cardiac transplantation. However, the salvage rate for those patients requiring circulatory support may be as low as 25%. Despite the fact that Penn State's 70 cc pneumatic ventricular assist device has been used with a success rate of over 90% in more than 250 patients worldwide, efforts to scale down the pump have encountered difficulties. Animal experiments with a 15 cc version were unsuccessful, with explanted pumps showing extensive thrombus deposition within the pumping chamber. The materials used to fabricate the smaller pump as well as the basic operating principles are identical to the successful adult-sized version. It is therefore believed that reducing the size of the pump altered the internal flow field, and that fluid dynamic factors were responsible for the high degree of thrombus observed with the implanted devices. A dimensional analysis was conducted that revealed significant differences in both Reynolds (Re) and Strouhal (St) numbers between the successful and unsuccessful pumps. Two component laser Doppler velocimetry was then used to characterize the internal flow field quantitatively. Comparison with data from the 70 cc pump showed a reduction in wall shear stress and turbulence levels in the 15 cc pump that would yield an environment conducive to clot formation.
Adult↗
Flow visualization in mechanical heart valves: occluder rebound and cavitation potential.
High density particle image velocimetry, with spatial resolution of O(1 mm), was used to measure the effect of occluder rebound on the flow field near a Bjork-Shiley Monostrut tilting-disk mitral valve. The ability to measure two velocity components over an entire plane simultaneously provides a very different insight into the flow compared to the more traditional point to point techniques (like Laser Doppler Velocimetry) that were utilized in previous investigations of the regurgitant flow. A picture of the effects of occluder rebound on the fluid flow in the atrial chamber is presented. Specifically, fluid velocities in excess of 1.5 m/s traveling away from the atrial side were detected 3 mm away from the valve seat in the local low pressure region created by the occluder rebound on the major orifice side where cavitation has been observed. This analysis is the first spatially detailed flow description of the effects of occluder rebound on the flow field past a tilting-disk mechanical heart valve and further reinforces the hypothesis that the rebound effect plays a significant role in the formation of cavitation, which has been implicated in the hemolysis and wear associated with tilting-disk valves in vivo.
Biomedical Engineering↗
The osmotic swelling characteristics of cardiac valve prostheses.
Several types of mechanical cardiac prostheses have been constructed with Delrin occluders, a material that is subject to osmotic swelling. The leaftets are designed to expand to specific tolerances when immersed in blood. The synthetic blood analogs commonly used in vitro contain hydrophilic compounds that can alter the osmotic expansion of the Delrin occluders. A static leak test chamber was employed to illustrate the effects of various test fluids on the sustained regurgitation phase of Delrin valves.
Equipment Failure Analysis↗
Expression of the human telomerase reverse transcriptase is not related to telomerase activity in normal and malignant renal tissue.
In this study, the association between telomerase activity and the expression of the human telomerase subunits human telomerase RNA (hTR) and human telomerase reverse transcriptase (hTERT) in paired neoplastic and normal renal tissue samples was investigated. Reverse transcription (RT)-PCR on 20 tumor nephrectomy samples revealed that hTR was constitutively expressed both in cancer and normal tissue samples, independent of the telomerase activity status. Remarkably, using in situ hybridization, the expression levels of hTR were found to be markedly higher in the normal tissue than those in the tumors. Expression of hTERT mRNA by RT-PCR was observed in 90% of the cancer samples and, notably, also in 75% of the corresponding normal renal tissue samples. Because all of the normal tissue samples and some of the tumor samples were shown to be telomerase negative, our findings suggest that hTERT mRNA expression is not sufficient for telomerase enzyme activation. Furthermore, semiquantitative RT-PCR revealed equal or even higher hTERT mRNA expression levels in the telomerase-negative normal samples than in the corresponding cancer samples with telomerase activity, contradicting the assumption that a certain threshold level of hTERT mRNA is required for telomerase activation at least in renal tissue. It seems more likely, that other mechanisms, such as posttranscriptional modification of hTERT or inactivation of telomerase inhibitors, are involved in the acquisition of enzyme activity.
Carcinoma, Renal Cell↗
The EVH2 domain of the vasodilator-stimulated phosphoprotein mediates tetramerization, F-actin binding, and actin bundle formation.
Vasodilator-stimulated phosphoprotein (VASP) is a member of the Ena/VASP family of proteins that are implicated in regulation of the actin cytoskeleton. All family members share a tripartite structural organization, comprising an N-terminal Ena/VASP homology (EVH) 1 domain, a more divergent proline-rich central part, and a common C-terminal EVH2 region of about 160-190 amino acids. Using chemical cross-linking, sucrose gradient sedimentation, and gel filtration analyses of different truncated VASP constructs, we demonstrate that the VASP EVH2 region is both necessary and sufficient for tetramerization. Moreover, co-sedimentation and fluorescent phalloidin staining showed that the EVH2 region binds and bundles F-actin in vitro and localizes to stress fibers in transfected cells. Analysis of the functional contribution of highly conserved blocks within this region indicated that residues 259-276 of human VASP are essential for the interaction with F-actin, whereas residues 343-380 are required for tetramerization, probably via coiled-coil formation. Interactions with F-actin are enhanced by VASP tetramerization. The results demonstrate that the C-terminal EVH2 segment is not only conserved in sequence but also forms a distinct functional entity. The data suggest that the EVH2 segment represents a novel oligomerization and F-actin binding domain.
Actins↗
L-arginine uptake, the citrulline-NO cycle and arginase II in the rat brain: an in situ hybridization study.
Nitric oxide (NO) is synthesized from a unique precursor, arginine, by nitric oxide synthase (NOS). In brain cells, arginine is supplied by protein breakdown or extracted from the blood through cationic amino acid transporters (CATs). Arginine can also be recycled from the citrulline produced by NOS activity, through argininosuccinate synthetase (AS) and argininosuccinate lyase (AL) activities, and metabolized by arginase. NOS, AS and AL constitute the so-called citrulline-NO cycle. In order to better understand arginine transport, recycling and degradation, we studied the regional distribution of cells expressing CAT1, CAT3, AS, AL, neuronal NOS (nNOS) and arginase II (AII) in the adult rat brain by non-radioisotopic in situ hybridization (ISH). CAT1, AL and AII presented an ubiquitous neuronal and glial expression, whereas CAT3 and AS were confined to neurons. nNOS was restricted to scattered neurons and a few brain nuclei and layers. We demonstrate by this study that cells expressing nNOS all appear to express the entire citrulline-NO cycle, whereas numerous cells expressing AL do not express AS. The differential expression of these genes within the same anatomical structure could indicate that intercellular exchanges of citrulline-NO cycle metabolites are relevant. Thus vicinal interactions should be taken into account to study their regulatory mechanisms.
Amino Acid Transport Systems, Basic↗
Hyperammonemia: regulation of argininosuccinate synthetase and argininosuccinate lyase genes in aggregating cell cultures of fetal rat brain.
Hyperammonemia in the brain leads to poorly understood alterations of nitric oxide (NO) synthesis. Arginine, the substrate of nitric oxide synthases, might be recycled from the citrulline produced with NO by argininosuccinate synthetase (AS) and argininosuccinate lyase (AL). The regulation of AS and AL genes during hyperammonemia is unknown in the brain. We used brain cell aggregates cultured from dissociated telencephalic cortex of rat embryos to analyze the regulation of AS and AL genes in hyperammonemia. Using RNase protection assay and non-radioactive in situ hybridization on aggregate cryosections, we show that both AS and AL genes are induced in astrocytes but not in neurons of aggregates exposed to 5 mM NH4Cl. Our work suggests that the hyperammonemic brain might increase its recycling of citrulline to arginine.
Ammonia↗
Identification of profilin and src homology 3 domains as binding partners for Drosophila enabled.
Drosophila Enabled (Ena) was first identified as a genetic suppressor of mutations in the Abelson tyrosine kinase and subsequently was shown to be a member of the Ena/vasodilator-stimulated phosphoprotein family of proteins. All members of this family have a conserved domain organization, bind the focal adhesion protein zyxin, and localize to focal adhesions and stress fibers. Members of this family are thought to be involved in the regulation of cytoskeleton dynamics. The Ena protein sequence has multiple poly-(L-proline) residues with similarity to both profilin and src homology 3 binding sites. Here, we show that Ena can bind directly to the Drosophila homolog of profilin, chickadee. Furthermore, Ena and profilin were colocalized in spreading cultured cells. We report that the proline-rich region of Ena is responsible for this interaction as well as for mediating binding to the src homology 3 domain of the Abelson tyrosine kinase. These data support the hypothesis that Ena provides a regulated link between signal transduction and cytoskeleton assembly in the developing Drosophila embryo.
Animals↗
Microheterogeneity of serum glycoproteins in patients with chronic alcohol abuse compared with carbohydrate-deficient glycoprotein syndrome type I.
BACKGROUND: Chronic alcohol abuse alters the normal N-glycosylation of transferrin, producing the carbohydrate-deficient transferrin isoforms. This alteration could be similar to that present in patients with carbohydrate-deficient glycoprotein syndrome type 1 (CDG1). We thus compared the alterations of N-glycans present in patients with alcoholism and patients with CDG1. METHODS: The N-glycans of serum glycoproteins were compared in sera of patients with alcoholism, patients with CDG1, and controls by two-dimensional electrophoresis, neuraminidase, peptide:N-glycosidase F, and endoglycosidase F2 treatments. A specific antibody directed against the amino acid sequence surrounding the N-432 N-glycosylation site of transferrin was prepared (SZ-350 antibody). RESULTS: In patients with alcoholism, the abnormal transferrin and alpha(1)-antitrypsin isoforms were devoid of a variable number of entire N-glycan moieties and were identical with those present in CDG1. In the serum of patients with alcoholism, this finding was less pronounced than in CDG1. In contrast to CDG1, there was no decrease in clusterin or serum amyloid P in patients with alcoholism. The SZ-350 antibody recognized only transferrin isoforms with one or no N-glycan moieties. CONCLUSION: Antibodies directed against specific N-glycosylation sites of glycoproteins could be useful for developing more specific immunochemical tests for the diagnosis of chronic alcohol abuse.
Alcoholism↗
Analysis and regulation of vasodilator-stimulated phosphoprotein serine 239 phosphorylation in vitro and in intact cells using a phosphospecific monoclonal antibody.
The development and functional analysis of a monoclonal antibody (16C2) are reported; the antibody recognizes vasodilator-stimulated phosphoprotein (VASP; an established substrate of both cAMP- and cGMP-dependent protein kinase) only when serine 239 is phosphorylated. VASP serine 239 represents one of the best characterized cGMP-dependent protein kinase phosphorylation sites in vitro and in intact cells. Experiments with purified, recombinant human VASP and various VASP constructs with mutated phosphorylation sites (S157A, S239A, T278A) and experiments with intact cells (human/rat platelets and other cells) treated with cyclic nucleotide-elevating agents demonstrated the specificity of the monoclonal antibody 16C2. Quantitative analysis of the VASP shift from 46 to 50 kDa (indicating VASP serine 157 phosphorylation) and the appearance of VASP detected by the 16C2 monoclonal antibody (VASP serine 239 phosphorylation) in human platelets stimulated by selective protein kinase activators confirmed that serine 239 is the VASP phosphorylation site preferred by cGMP-dependent protein kinase in intact cells. Immunofluorescence experiments with human platelets treated with cGMP analogs showed that the 16C2 monoclonal antibody also detects VASP serine 239 phosphorylation in situ at established intracellular localization sites. Analysis of VASP serine 239 phosphorylation by the 16C2 antibody appears to be the best method presently available to measure cGMP-dependent protein kinase activation in intact cells. Also, the 16C2 antibody promises to be an excellent tool for the evaluation of VASP function in intact cells.
Animals↗