Biomedical subjects

A Acheson

Publications and source records attributed to A Acheson.

At least 37 records · Page 2Linked to original sources

PubMed1994

Characterization and crystallization of recombinant human neurotrophin-4.

Neurotrophin-4 (NT-4) is the most recently discovered member of the neurotrophin family. We have expressed, refolded, and purified recombinant human NT-4 from Escherichia coli and compared it with recombinant human NT-4 secreted into the culture medium of baculovirus-infected insect cells. Both preparations were characterized and determined to be indistinguishable according to several biochemical criteria. Recombinant NT-4 from E. coli was crystallized in a form suitable for x-ray analysis, and characterization of these crystals indicated that NT-4 was present as a dimer within the asymmetric unit. NT-4 was active in promoting the survival of rat TrkB receptor-expressing fibroblasts, but was inactive on embryonic chicken sensory neurons, unlike the other members of the neurotrophin family and in contrast to the reported activities of partially purified NT-4.

Animals↗

PubMed1993

Exchange of a single amino acid interconverts the specific activity and gel mobility of human and rat ciliary neurotrophic factors.

Human and rat ciliary neurotrophic factors (CNTF), which share 85% sequence identity, promote the survival of chicken embryo ciliary ganglia neurons in vitro, but display a 4-5-fold difference in specific activity. To explore the origin of this difference and gain insight into the structural organization of CNTF, we created chimeric proteins of these two species. Surprisingly, we found that the differences in two apparently unrelated properties, gel mobility and specific activity, resided in a single amino acid. Substituting arginine residue 63 of rat CNTF into the human sequence created a protein with the properties of rat CNTF. Conversely, substituting the human CNTF glutamine residue 63 into rat CNTF generated a protein with the properties of human CNTF. Binding experiments confirmed that the distinct specific activities of human and rat CNTF and their chimeras reside in structural differences among these ligands rather than species differences in their receptors. Alanine substitution (Q63A) had no effect on the properties of human CNTF, whereas the R63A substitution reduced both the gel mobility and the specific activity of rat CNTF. Finally, a Q95R substitution at a different position of human CNTF had no effect on its properties. These results demonstrate that Arg-63 is both specific and critical in determining the structural differences of human and rat CNTF.

Amino Acid Sequence↗

PubMed1993

Immunological relationships of NGF, BDNF, and NT-3: recognition and functional inhibition by antibodies to NGF.

Polyclonal antibodies raised against mouse 2.5S NGF (mNGF) and against synthetic peptides made from hydrophilic portions of mNGF have been used to compare the immunological properties of mNGF, human recombinant brain-derived neurotrophic factor (hrBDNF), and human recombinant neurotrophin-3 (hrNT-3). Affinity-isolated antibodies raised against intact mNGF reacted with all three neurotrophins when tested by ELISA and totally or partially blocked the bioactivities of the proteins in survival assays of embryonic chicken sensory and sympathetic neurons. On Western blots, mNGF antibodies reacted with all three neurotrophins but less well with hrBDNF and hrNT-3 than with mNGF. Antibodies to hydrophilic peptides within NGF (amino acids 23-35, 59-67, 69-79, and 91-100) showed partial reactivity with some but not all of the neurotrophins when tested by ELISA and on Western blots. The peptide antibodies were also selectively effective in reducing the survival-promoting activity of the neurotrophins on sensory neurons. Results show that mNGF, hrBDNF, and hrNT-3 are immunologically related proteins and that mNGF antibodies react also with other members of the neurotrophin family.

Amino Acid Sequence↗

PubMed1992

Cholinergic differentiation of rat sympathetic neurons in culture: effects of factors applied to distal neurites.

Cholinergic properties are induced in sympathetic neurons by several factors applied to entire neurons in culture. Evidence from work with the rat sweat gland model indicates that factors located in target tissues can induce cholinergic differentiation in vivo. We now report that when leukemia inhibitory factor (LIF), heart cell-conditioned medium (HCCM), or dermal fibroblast-conditioned medium (DFCM) is applied to only distal neurites in compartmented cultures of rat sympathetic neurons, the neurons exhibit an increase in specific choline acetyltransferase activity and a concomitant decrease in levels of tyrosine hydroxylase. LIF, HCCM, and DFCM also induce neurite fasciculation, thus suggesting an additional role of cholinergic switching factors in regulating axon-axon and/or axon-substrate adhesion. These results demonstrate that rat sympathetic neurons have the cellular machinery to respond to cholinergic differentiation cues located in peripheral targets, analogous to the response to nerve growth factor.

Animals↗

PubMed1992

Spatiotemporal increases in epidermal growth factor receptors following peripheral nerve injury.

Non-neuronal cells of peripheral nerve respond to axonal injury with a series of cellular changes that facilitate neuronal regeneration. To characterize the potential role of the epidermal growth factor (EGF) family of proteins in this response, we monitored the expression of EGF receptor mRNA and protein in the injured rat sciatic nerve. EGF receptor mRNA is synthesized in both primary cultured fibroblasts and Schwann cells, and Schwann cells express EGF receptor-like immunoreactivity. In situ hybridization and immunocytochemistry revealed that EGF receptor mRNA and protein are expressed in Schwann cells and fibroblasts of the sciatic nerve in vivo, and that receptor levels increase following nerve injury. Thirty-six hours postlesion, EGF receptors were expressed in gradients along the nerve both proximal and distal to the lesion, with the highest levels localized adjacent to the transection site. By 72 hr, receptor levels were maintained in a gradient in the proximal segment, but were uniformly increased throughout the portions of the distal segment that were analyzed. These changes were similar to those observed for low-affinity NGF receptor mRNA and protein, with transection causing increased expression in both Schwann cells and fibroblasts. Northern blots confirmed that primary cultured fibroblasts express low-affinity NGF receptor mRNA. To determine whether spatiotemporal gradients were a general characteristic of the nerve injury response, we monitored expression of the mRNA encoding the major myelin protein P0. Levels of P0 mRNA decreased initially in cells immediately adjacent to the transection site and, by 72 hr, were uniformly decreased throughout the distal segment. These data suggest that members of the EGF family of proteins may play a role in the peripheral nerve response to injury, and demonstrate a generalized gradient of cellular responses that commence at the transection site and progress distally in the nerve in the absence of intact axons.

Animals↗

PubMed1991

Detection of brain-derived neurotrophic factor-like activity in fibroblasts and Schwann cells: inhibition by antibodies to NGF.

mRNA coding for brain-derived neurotrophic factor (BDNF) has been detected in cultured L929 fibroblasts, rat dermal fibroblasts, and sciatic nerve Schwann cells, as well as in rat skin. Medium conditioned by cultured fibroblasts and Schwann cells also stimulates neurite growth from retinal explants and promotes the survival in culture of BDNF-responsive sensory neurons; biological activity is abolished by antibodies raised against NGF. These results suggest that molecules with BDNF-like activity may be produced by cells in the peripheral nervous system and that the BDNF-like activity in fibroblasts and Schwann cells is derived from molecules immunologically related to NGF. In support of this concept, antibodies against NGF have been found to reduce the biological activity of recombinant BDNF in culture and to cross-react with BDNF on Western blots.

Animals↗

PubMed1991

NCAM polysialic acid can regulate both cell-cell and cell-substrate interactions.

We have proposed previously that the polysialic acid (PSA) moiety of NCAM can influence membrane-membrane apposition, and thereby serve as a selective regulator of a variety of contact-dependent cell interactions. In this study, cell and tissue culture models are used to obtain direct evidence that the presence of PSA on the surface membrane can affect both cell-cell and cell-substrate interactions. Using a neuroblastoma/sensory neuron cell hybrid, it was found that removal of PSA with a specific neuraminidase (endo-N) augments cell-cell aggregation mediated by the L1 cell adhesion molecule as well as cell attachment to a variety of tissue culture substrates. In studies of embryonic spinal cord axon bundling, which involves both cell-cell and cell-substrate interactions, the pronounced defasciculation produced by removal of PSA is most easily explained by an increase in cell-substrate interaction. The fact that in both studies NCAM's intrinsic adhesion function was found not to be an important variable further illustrates that regulation of the cell surface by PSA can extend beyond binding mediated by the NCAM polypeptide.

Animals↗

PubMed1990

Effect of empiric antiarrhythmic therapy in resuscitated out-of-hospital cardiac arrest victims with coronary artery disease.

The effect of empiric antiarrhythmic therapy with quinidine and procainamide on long-term mortality was examined in 209 patients with coronary artery disease resuscitated after out-of-hospital cardiac arrest. The antiarrhythmic agent used was determined by the patient's private physician without knowledge of the study ambulatory electrocardiogram. Of the 209 patients, procainamide was prescribed in 45 (22%), quinidine in 48 (23%) and no antiarrhythmic therapy in 116 (55%). Digoxin therapy was initiated in 101 patients. The 2-year total survival rate for the quinidine, procainamide and nontreated patients was 61, 57 and 71% (p less than 0.05), and for sudden death was 69, 69 and 89% (p less than 0.01), respectively. These observations suggest that empiric antiarrhythmic therapy in survivors of out-of-hospital cardiac arrest did not affect total mortality and was associated with an increased frequency of sudden death.

Aged↗

PubMed1988

The neural cell adhesion molecule (NCAM) as a regulator of cell-cell interactions.

The neural cell adhesion molecule (NCAM) can influence a number of diverse intercellular events, including junctional communication, the association of axons with pathways and targets, and signals that alter levels of neurotransmitter enzymes. These pleiotropic effects appear to reflect the ability of NCAM to regulate membrane-membrane contact required to initiate specific interactions between other molecules. Such regulation can occur through changes in either NCAM expression or the molecule's content of polysialic acid (PSA). When NCAM with a low PSA content is expressed, adhesion is increased and contact-dependent events are triggered. In contrast, the large excluded volume of NCAM PSA can inhibit cell-cell interactions through hindrance of overall membrane apposition.

Animals↗

PubMed1988

Neural cell adhesion molecule regulates cell contact-mediated changes in choline acetyltransferase activity of embryonic chick sympathetic neurons.

E10 chick sympathetic ganglion cells display a cell contact-dependent rise in choline acetyltransferase (ChAT) specific activity over the first several days in culture. This effect can be mimicked by addition of crude membrane fractions prepared from E10 retina and adult chicken brain, but not by those from E10 brain. The effects of both cell-cell and membrane-cell contact are inhibited by the addition of anti-NCAM Fab fragments. The membranes capable of increasing ChAT and those which are ineffective all contain NCAM, however their relative levels of NCAM polysialic acid differ. Whereas membranes with high polysialic acid NCAM are ineffective, selective enzymatic removal of polysialic acid renders them capable of producing an increase in ChAT. The inhibition of NCAM-mediated adhesion produced by Fab fragments can be compensated for by addition of wheat germ agglutinin, but only with membranes whose NCAM has low levels of polysialic acid. Taken together, these data suggest that NCAM can regulate cell contact-mediated increases in ChAT activity. We propose that NCAM-mediated adhesion promotes contact between cell membranes to allow the transmission of an otherwise NCAM-independent signal. In addition, NCAM's polysialic acid moiety appears to influence the ability of cells to transmit this signal, even in the presence of an alternative adhesion mechanism.

Animals↗

PubMed1988

Predictive value of ventricular arrhythmias in resuscitated out-of-hospital cardiac arrest victims.

Twenty-four hour ambulatory electrocardiograms recorded in 103 survivors of out-of-hospital cardiac arrest were analyzed to find those characteristics of the ventricular premature complex (VPC) which provide the best combination of sensitivity, specificity, and predictive accuracy for subsequent mortality. VPC characteristics were grouped as: (1) frequent (greater than or equal to 25 h-1), (2)bigeminal, (3) multiform, (4) early coupled, (5) pairing, (6) repetitive greater than or equal to 2, (7) repetitive greater than or equal to 3, (8) repetitive greater than or equal to 6, (9) the combination of frequent and repetitive, or (10) complex defined as any multiform, early, bigeminal or repetitive VPC. In an average follow-up period of 43 months, 42 deaths occurred, 17 of which were classified as sudden. Each characteristic was a significant predictor for all causes of subsequent death except early coupled VPCs and repetitive VPCs greater than 6. None of the characteristics reached significance as predictors for sudden death. The number of repetitive VPCs when stratified to none, greater than or equal to 2 and greater than or equal to 3 successive VPCs correlated with mortality in an incremental fashion. The combination of frequent VPCs and repetitive VPCs provided the best combination of sensitivity, specificity and predictive accuracy for death from all causes within five years.

Arrhythmias, Cardiac↗

PubMed1988

Clinical characteristics and survival experience of out-of-hospital cardiac arrest victims without coronary heart disease.

Of 270 patients successfully resuscitated from out-of-hospital cardiac arrest, 16% had no evidence of coronary heart disease. In these 43 patients, other forms of heart disease were found in 81% (35/43): cardiomyopathy in 18 patients, valvular disease in six, congenital heart disease in two, and primary arrhythmia in nine. Seven patients had evidence of only pulmonary disease and one had pancreatitis as his precipitating event. Nineteen of the 43 patients (44%) had serum potassium less than 3.6 mEq l-1 in the initial blood sample after cardiac arrest. One- and two-year mortalities were 30% and 43%, respectively, for the group, which is similar to one-year (20%) and two-year (35%) mortalities of the 227 resuscitated patients with coronary heart disease. Patients who survive a sudden death experience and who have no evidence of coronary artery disease are a unique but heterogeneous group who usually have identifiable cardiac or pulmonary disease.

Cardiac Complexes, Premature↗

PubMed1987

High K+-mediated survival of spinal sensory neurons depends on developmental stage.

Elevated concentrations of K+ (35 mM) have previously been shown to support the survival of most embryonic chick sympathetic neurons in vitro (Wakade et al., Exp cell res 144 (1983) 377, [23]) and to be interchangeable with nerve growth factor (NGF) as a survival-promoting agent for these cells (Wakade & Thoenen, Neurosci lett 45 (1984) 71 [21]). In the present study, we show that dorsal root ganglion (DRG) neurons from embryonic day 6 do not survive in the presence of high K+, although both NGF and brain-derived neurotrophic factor (BDNF) each support the survival of more than 50% of the cells at this developmental stage. At E6, high K+ appears to have a cytotoxic effect on BDNF-dependent neurons, and there is also considerable inhibition of neurite outgrowth. At a later developmental stage (E12), high K+ supports the survival of about 40% of DRG cells. This subpopulation of neurons is distinct from that supported by NGF (as evidenced by the additivity of these two agents), but partially overlaps with that supported by BDNF (i.e., the two agents are less than additive). At E12, only approx. 20% of the cells can be supported by either NGF or BDNF, with the rest depending exclusively on one or the other of these factors. This is in contrast to the situation at E6, where there is considerable overlap between NGF- and BDNF-dependent populations.

Animals↗

PubMed1987

Both short- and long-term effects of nerve growth factor on tyrosine hydroxylase in calf adrenal chromaffin cells are blocked by S-adenosylhomocysteine hydrolase inhibitors.

We have previously shown that primary cultures of calf chromaffin cells respond to nerve growth factor (NGF) treatment with a selective induction of tyrosine hydroxylase (TH), which takes 48 h to be manifested. In the present study, we report that short exposure of calf chromaffin cells to NGF (5-60 min) results in TH activation, which involves a change in the Vmax of the enzyme with no change in the number of enzyme molecules, similar to an effect that has been previously reported in PC12 cells. This activation is markedly potentiated when the chromaffin cells are plated on a laminin substrate, such that after 5 min of NGF exposure, there is an approximately fourfold increase in the TH activity. Both short-term activation and long-term TH induction brought about by NGF treatment are blocked by 5'-deoxy-5'-methylthioadenosine and other drugs that act as S-adenosylhomocysteine (SAH) hydrolase inhibitors to block methylation by end-product inhibition. These drugs did not inhibit cyclic AMP-mediated TH activation or increases in the levels of TH. However, measurements of the degree of blockade of methylation in cells treated with these drugs, taken together with conceptual information regarding the nonregulatory nature of methylation in eukaryotic cells, were not consistent with inhibition of methylation as the crucial effect of the drugs to block the effects of NGF. Nonetheless, since SAH hydrolase inhibitors selectively inhibited NGF-mediated effects, and not comparable effects triggered by other stimuli, these compounds provide useful tools in future studies of the biochemical signalling mechanism of NGF.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1986

Analysis of cardiac symptoms preceding cardiac arrest.

Prodromal symptoms and cardiac history were examined in 227 patients with coronary artery disease who were successfully resuscitated after out-of-hospital cardiac arrest. Cardiac arrest was sudden--with either no symptoms or symptoms for less than 1 hour--in 71% of the patients. Nonsudden death--death occurring after more than 1 hour of symptoms--occurred in 29% of the patients. A history of cardiovascular disease was present in 85% of patients with sudden cardiac arrest and in 83% with nonsudden arrest. Cardiac arrest occurred without symptoms in 38% of the patients with sudden cardiac arrest and was the first expression of coronary artery disease in 4% of the entire study group. This study indicates that cardiac arrest usually occurs with symptoms and almost always in the setting of a history of cardiovascular disease.

Angina Pectoris↗

PubMed1986

Methyltransferase inhibitors block NGF-regulated survival and protein phosphorylation in sympathetic neurons.

Nerve growth factor (NGF) and elevated K+ concentrations (35 mM) support the survival of the same population of chick embryonic sympathetic neurons. We have used methyltransferase inhibitors, which block protein methylation in intact cells, to investigate the mechanism(s) by which NGF and high K+ exert their effects. Methyltransferase inhibitors selectively blocked NGF-but not high K+-mediated survival of neurons. The ability of neurons, plated on laminin, to respond rapidly to NGF with neurite outgrowth was used to demonstrate that the blockade of the effects of NGF by methyltransferase inhibitors was reversible. At the molecular level, we studied the rapid decrease in phosphorylation of p70, a 70-kd phosphoprotein of sympathetic neurons regulated by both NGF and high K+. Methyltransferase inhibitors blocked the decrease in p70 phosphorylation induced by NGF but not that by high K+. We conclude that the early molecular events of NGF-mediated neuronal survival differ from those of high K+-mediated neuronal survival in that they involve protein methylation, whereas at a later step, possibly at the level of protein phosphorylation, the two pathways leading to survival of sympathetic neurons converge.

Adenosine↗

PubMed1986

Laminin increases both levels and activity of tyrosine hydroxylase in calf adrenal chromaffin cells.

We have investigated the effects of substrate-bound laminin on levels of enzymes of the catecholamine biosynthetic pathway in primary cultures of calf adrenal chromaffin cells. Laminin increases the levels of the enzymes tyrosine hydroxylase, dopamine-beta-hydroxylase, and phenylethanolamine-N-methyl-transferase. This effect is selective, in that levels of other enzymes (lactate dehydrogenase, aromatic amino acid decarboxylase, and acetylcholinesterase) are not increased. The effect of laminin can be blocked by antibodies directed against a fragment of the heparin-binding domain of the molecule, whereas antibodies directed against other fragments do not block the increase in tyrosine hydroxylase. Thus the laminin domain involved in enzyme regulation in chromaffin cells is apparently the same as that previously implicated in laminin's interactions with neurons to potentiate survival and stimulate neurite outgrowth (Edgar, D., R. Timpl, and H. Thoenen, 1984, EMBO (Eur. Mol. Biol. Organ.) J., 3:1463-1468). The increase in chromaffin cell tyrosine hydroxylase levels is preceded by an activation of the enzyme in which the Vmax (but not the Km) is altered. The effects of laminin appear to be developmentally regulated, since neither activation nor increased levels of tyrosine hydroxylase occur in adult adrenal chromaffin cells exposed to laminin.

Animals↗

PubMed1985

Nerve growth factor.

In contrast to all other molecules which are labelled 'growth factor', NGF is not a mitogen. It is a neurotrophic molecule essential for the development and maintenance of function of specific populations of peripheral and possibly also central neurons. The availability of NGF in large quantities from exocrine glands (e.g. male mouse submandibular gland), where NGF does not play a neurotrophic role, has allowed the purification of NGF, the production of specific antibodies, the determination of its amino acid sequence and finally the molecular cloning of NGF leading to the elucidation of its precursor structure and its genomic organization. Comparison of the biological activities and the immunological properties of NGF isolated from different sources demonstrated that the active centre of the molecule has been highly conserved during evolution, whereas other parts of the molecule determining immunological properties have undergone considerable changes. After a survey of the essential biological actions of NGF, this paper concentrates on two actual questions of NGF research, namely the regulation of NGF synthesis in the target tissues of NGF-responsive neurons, and the molecular mechanism(s) of action of NGF on these neurons.

Animals↗