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Biomedical subjects

M Ericsson

Publications and source records attributed to M Ericsson.

At least 55 records · Page 3Linked to original sources
PubMed1994

Increased synthesis of DnaK, GroEL, and GroES homologs by Francisella tularensis LVS in response to heat and hydrogen peroxide.

The response of the facultative intracellular bacterium Francisella tularensis LVS to stress was assayed by pulse-labeling with [35S]methionine followed by two-dimensional gel electrophoresis and autoradiography. A temperature increase from 37 to 42 degrees C or exposure to 5 mM hydrogen peroxide induced increased syntheses of at least 15 proteins. Among these proteins were a 75-, a 60-, and a 10-kDa protein. By N-terminal sequence analysis, these three proteins were found to be extensively homologous to the highly conserved chaperone proteins DnaK, GroEL, and GroES of Escherichia coli. Antibodies specific to the DnaK homolog of E. coli reacted with the 75-kDa protein, and antibodies to the GroEL homolog of Legionella micdadei reacted with the 60-kDa protein. A readiness to respond to hydrogen peroxide with synthesis of the chaperone components may be fundamental to the intracellular survival of pathogens such as F. tularensis, which are exposed to oxidative stress while invading the host macrophages.

Amino Acid Sequence↗
PubMed1994

Assembly of vaccinia virus: the second wrapping cisterna is derived from the trans Golgi network.

During the assembly of vaccinia virus, the intracellular mature virus becomes enwrapped by a cellular cisterna to form the intracellular enveloped virus (IEV), the precursor of the extracellular enveloped virus (EEV). In this study, we have characterized the origin of this wrapping cisterna by electron microscopic immunocytochemistry using lectins, antibodies against endocytic organelles, and recombinant vaccinia viruses expressing proteins which behave as Golgi resident proteins. No labelling for endocytic marker proteins could be detected on the wrapping membrane. However, the wrapping membrane labelled significantly for a trans Golgi network (TGN) marker protein. The recycling pathway from endosomes to the TGN appears to be greatly increased following vaccinia virus infection, since significant amounts of endocytic fluid-phase tracers were found in the lumen of the TGN, Golgi complex, and the wrapping cisternae. Using immunoelectron microscopy, we localized the vaccinia virus membrane proteins VV-p37, VV-p42, VV-p21, and VV-hemagglutinin (VV-HA) in large amounts in the wrapping cisternae, in the outer membranes of the IEV, and in the outermost membrane of the EEV. The bulk of the cellular VV-p37, VV-p21, and VV-p42 were in the TGN, whereas VV-HA was also found in large amounts on the plasma membrane and in endosomes. Collectively, these data argue that the TGN becomes enriched in vaccinia virus membrane proteins that facilitate the wrapping event responsible for the formation of the IEV.

Animals↗
PubMed1994

Assembly of vaccinia virus: effects of rifampin on the intracellular distribution of viral protein p65.

The cytoplasmic assembly of vaccinia virus is reversibly blocked by the antibiotic rifampin, leading to the accumulation of partially membrane-delineated rifampin bodies in infected cells. Rifampin-resistant vaccinia virus mutants have point mutations in the D13L gene, which is controlled by a late promoter and expresses a 65-kDa protein, designated p65. To further characterize the mechanism of rifampin inhibition and the function of p65 in virus assembly, we raised antibodies to this protein. Immunoreactive p65 was expressed at late times of infection, and neither its expression nor its turnover was affected by rifampin. Virus-associated p65 could be extracted only with denaturing detergents from purified virions, suggesting that it is an integral viral component. Immunofluorescence studies showed that p65 is localized to the sites of virus assembly. Also, immunoelectron microscopy showed p65 to be associated with viral crescents as well as spherical, immature virions, in both cases predominantly on the inner or concave surface. In the presence of rifampin, p65 was found in large, cytoplasmic inclusion bodies that were distinct from rifampin bodies. The rifampin bodies themselves were labeled with p65 antibodies only after reversal of the rifampin block, predominantly on the viral crescents which rapidly formed following removal of the drug. We propose that p65 functions as an internal scaffold in the formation of viral crescents and immature virions, analogously to the matrix proteins of other viruses.

Amino Acid Sequence↗
PubMed1993

Assembly of vaccinia virus: role of the intermediate compartment between the endoplasmic reticulum and the Golgi stacks.

Vaccinia virus, the prototype of the Poxviridae, is a large DNA virus which replicates in the cytoplasm of the host cell. The assembly pathway of vaccinia virus displays several unique features, such as the production of two structurally distinct, infectious forms. One of these, termed intracellular naked virus (INV), remains cells associated while the other, termed extracellular enveloped virus (EEV), is released from the cell. In addition, it has long been believed that INVs acquire their lipid envelopes by a unique example of de novo membrane biogenesis. To examine the structure and assembly of vaccinia virus we have used immunoelectron microscopy using antibodies to proteins of different subcellular compartments as well as a phospholipid analysis of purified INV and EEV. Our data are not consistent with the de novo model of viral membrane synthesis but rather argue that the vaccinia virus DNA becomes enwrapped by a membrane cisterna derived from the intermediate compartment between the ER and the Golgi stacks, thus acquiring two membranes in one step. Phospholipid analysis of purified INV supports its derivation from an early biosynthetic compartment. This unique assembly process is repeated once more when the INV becomes enwrapped by an additional membrane cisterna, in agreement with earlier reports. The available data suggest that after fusion between the outer envelope and the plasma membrane, mature EEV is released from the cell.

Endoplasmic Reticulum↗
PubMed1993

Characterization of a novel 63 kDa membrane protein. Implications for the organization of the ER-to-Golgi pathway.

Owing to the lack of appropriate markers the structural organization of the ER-to-Golgi pathway and the dynamics of its membrane elements have been elusive. To elucidate this organization we have taken a monoclonal antibody (mAb) approach. A mAb against a novel 63 kDa membrane protein (p63) was produced that identifies a large tubular network of smooth membranes in the cytoplasm of primate cells. The distribution of p63 overlaps with the ER-Golgi intermediate compartment, defined by a previously described 53 kDa marker protein (here termed ERGIC-53), as visualized by confocal laser scanning immunofluorescence microscopy and immunoelectron microscopy. The p63 compartment mediates protein transport from the ER to Golgi apparatus, as indicated by partial colocalization of p63 and vesicular stomatitis virus G protein in Vero cells cultured at 15 degrees C. Low temperatures and brefeldin A had little effect on the cellular distribution of p63, suggesting that this novel marker is a stably anchored resident protein of these pre-Golgi membranes. p63 and ERGIC-53 were enriched to a similar degree by the same subcellular fractionation procedure. These findings demonstrate an unanticipated complexity of the ER-Golgi interface and suggest that the ER-Golgi intermediate compartment defined by ERGIC-53 may be part of a greater network of smooth membranes.

Animals↗
PubMed1992

Reptilian alcohol dehydrogenase. Heterogeneity relevant to class multiplicity of the mammalian enzyme.

Liver alcohol dehydrogenase of the ethanol-active type ('class I enzyme') from the lizard, Uromastix hardwickii, was purified and screened for relationships with other vertebrate forms of the enzyme. Two different acetylated N-termini (acetyl-Gly and acetyl-Ser) and further positional differences already in the N-terminal segments establish the presence of two types of protein chain. The multiplicity is different from that hitherto detected within vertebrate class I alcohol dehydrogenase isozymes but typical of that which would be expected for subunits of different classes. In particular, relationships to class II or to class II-related forms appear likely. This may indicate yet further vertebrate alcohol dehydrogenase multiplicity or discovery of a class II non-mammalian enzyme. The results give prospects of defining gene duplications corresponding to more than one alcohol dehydrogenase class split to at an early vertebrate stage.

Alcohol Dehydrogenase↗
PubMed1991

Analogues of parathyroid hormone modified at positions 3 and 6. Effects on receptor binding and activation of adenylyl cyclase in kidney and bone.

Predictive and spectroscopic methods were used to develop a model of the structures of the 1-34 peptides of parathyroid hormone (PTH) and the PTH-related protein (PTHrP). Circular dichroism (CD) studies of bovine PTH-(1-34) and human PTHrP-(1-34)amide in the presence of trifluoroethanol suggest the presence of 24-26 alpha-helical residues. For both peptides, interactions between amino- and carboxyl-region alpha-helices are predicted to result in a hydrophobic core with externally facing hydrophilic residues that include probable determinants of receptor binding and activation. Two such residues, Ser3 and Gln6, are conserved in all known members of the PTH/PTHrP family. We have synthesized 13 novel analogues of bovine PTH-(1-34) monosubstituted at positions 3 and 6 and have determined their biological activities in renal and bone cell radioreceptor and adenylyl cyclase assays. Position 3 analogues displayed biological activity that was reduced in direct proportion to the volume of the substituent side-chain. Position 6 analogues also displayed reduced biological activity, but no simple correlation with side-chain volume or hydrophobicity was evident. The analogues fully displaced labeled PTH from binding sites in renal membranes and bone cells, but [Phe3]bPTH-(1-34), [Tyr3]bPTH-(1-34), [Phe6] bPTH-(1-34), and [Ser6]bPTH-(1-34) were only partial agonists in one or both adenylyl cyclase assays. Of these, [Phe3]bPTH-(1-34) and [Phe6]bPTH-(1-34) were tested for antagonist activity and were found to inhibit the activation of adenylyl cyclase in response to bPTH-(1-34) or hPTHrP-(1-34)amide. These results indicate that positions 3 and 6 contribute important determinants of PTH receptor binding and activation. Modification at these positions represents a novel approach to the development of antagonists of PTH action.

Adenylyl Cyclases↗
PubMed1991

Behavioural profile of partial D2 dopamine receptor agonists. 1. Atypical inhibition of d-amphetamine-induced locomotor hyperactivity and stereotypy.

The effects of partial D2 dopamine (DA) receptor agonists on the behavioural activation produced by 1.5 and 8.0 mg/kg d-amphetamine were compared with the changes produced by the classical DA antagonist haloperidol. Alterations in behaviour were assessed in standard activity monitoring cages by direct observation of the rats using a rapid time sampling procedure. Haloperidol blocked d-amphetamine (1.5 mg/kg)-induced increases in photocell counts, ambulation, rearing and sniffing up, and after the highest dose of the DA antagonist the animals were mainly inactive. The partial D2 DA agonist SDZ 208-911 was equipotent to haloperidol in blocking the increase in photocell counts and rearing produced by d-amphetamine. However, even high doses of the drug did not reduce the incidence of sniffing or induce inactivity, but qualitative changes in the form of sniffing did occur. Although considerably less potent, preclamol exerted similar effects to SDZ 208-911. The profiles of SDZ 208-912 and terguride were intermediary to those of SDZ 208-911 and haloperidol. All compounds blocked the repetitive sniffing down produced by 8.0 mg/kg d-amphetamine. After a low dose of haloperidol, these stereotyped behaviours were replaced by a behavioural syndrome similar to that observed with low dose d-amphetamine, but inactivity was observed following a further small increase in antagonist dose. The blockade of stereotypy by SDZ 208-911, preclamol and terguride was accompanied only by the low dose d-amphetamine behavioural syndrome; no inhibition of sniffing or induction of inactivity occurred. SDZ 208-912 exhibited a profile with features very similar to that noted with haloperidol. These findings suggest that partial D2 agonists exert similar, but not identical, behavioural effects to classical DA antagonists when dopaminergic function in increased by d-amphetamine. The differences in behavioural profile are discussed in relation to variations in the intrinsic efficacy of the dopaminergic compounds and to differences in the response capability of D2 receptor populations underlying the different behaviours produced by d-amphetamine.

Animals↗
PubMed1991

Targeting of frog prodermorphin to the regulated secretory pathway by fusion to proenkephalin.

We have investigated the sorting and processing of the amphibian precursor prepro-dermorphin in mammalian cells. Dermorphin, a D-alanine-containing peptide with potent opioid activity, has been isolated from the skin of the frog Phyllomedusa sauvagei. The maturation of this peptide from the precursor involves several posttranslational steps. Recombinant vaccinia viruses were used to infect AtT-20, PC12, and HeLa cells to study the sorting and processing of prepro-dermorphin. While this precursor was not processed in any of the examined cell lines, AtT-20 cells were able to process approximately 40% of a chimeric precursor consisting of the first 241 amino acids of prepro-enkephalin fused to a carboxy-terminal part of pro-dermorphin. By immunogold-EM, we could show that the chimeric protein, but not pro-dermorphin, was sorted to dense-core secretion granules. The processing products could be released upon stimulation by 8-Br-cAMP. We conclude that the pro-enkephalin part of the fusion protein contains the information for targeting to the regulated pathway of secretion, while this sorting information is missing in pro-dermorphin. This indicates that sorting mechanisms may differ between amphibian and mammalian cells.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1990

Detection of doubly stained fluorescent specimens using confocal microscopy.

Studies of doubly stained specimens were performed with a confocal scanning microscope. The instrument used provides the possibility of making separate detections of the fluorescent dyes. The optimal choice of excitation wavelengths and optical filters are discussed. The fluorphores that were used are Lucifer Yellow, Texas Red, fluorescein isothiocyanate and tetramethylrhodamine isothiocyanate.

Animals↗
PubMed1990

Spinal epidural abscesses in adults: review and report of iatrogenic cases.

A retrospective review of the medical records of adults with diagnosed spinal epidural abscess (SEA) admitted to the Departments of Neurosurgery and Infectious Diseases at the University Hospital of Umeå, Sweden, during a 10-year-period (1978-1987) is presented. 10 patients were diagnosed as having SEA during the study period. An iatrogenic origin was suspected in 5. Spondylitis was the most common source of infection. Bacteriological aetiology was confirmed in 8 cases and Staphylococcus aureus was the most common aetiological agent. Trauma and degenerative diseases of the spine, were underlying conditions in 6 cases. Laminectomy was performed in 7 cases. 5/6 patients operated within 48 h after onset of neurological symptoms improved. The remaining case with therapy resistant tuberculous spondylitis died. One patient with surgery after 96 h became paretic. 2/3 conservatively treated patients had a successful outcome while the third patient had a permanent paraparesis due to missed diagnosis. Early diagnosis and early laminectomy are still the most important prognostic factors. Recommended initial antibiotic therapy is the combination of a cephalosporin with extended spectrum and metronidazole.

Abscess↗
PubMed1989

The quadriceps femoris muscle in 20-70-year-old subjects: relationship between knee extension torque, electrophysiological parameters, and muscle fiber characteristics.

Comparisons have been made between torque (isometric and isokinetic), electrophysiological (SFEMG, Macro EMG), and muscle fiber characteristics in the vastus lateralis muscle of both legs in healthy subjects aged between 20 and 70 years. Torque was greater in males and decreased with age in both sexes. Multifactorial analysis showed a positive correlation between torque, body surface area, and mean fiber area. These variables explained only about 30-40% of the torque changes. The electrophysiological parameters (Marco EMG amplitudes and fiber density) revealed evidence of reinnervation, indicating preceeding denervation and therefore loss of motor units. It was concluded that this fall out of motor units also contributes to the reduction in torque, when compensatory reinnervation begins to fall. Other factors, such as reduction in muscle fiber contractility, metabolic factors, and central factors, may also play a role in age-related reduction in torque.

Adult↗
PubMed1988

Three-dimensional reconstruction of transmitter-identified central neurons by "en bloc" immunofluorescence histochemistry and confocal scanning microscopy.

A new method for three-dimensional reconstruction of transmitter-identified neurons is presented which involves "en bloc" immunofluorescence histochemistry and confocal scanning microscopy. The technique was applied to different types of neurons in the rat brain and lamprey spinal cord. Thick sections or tissue "blocs" (50-200 micron thick) were incubated with antisera against neuropeptides or monoaminergic markers, followed by fluorescent secondary antibodies. Three-dimensional reconstructions were obtained by scanning the preparations in sequential focal planes with a thin laser beam, while sampling the emitted light in each focal plane. The method is convenient and can be applied to a wide variety of neuron types. The reconstructions obtained are accurate since the "optical serial sections" of the specimen are perfectly aligned, and optic disturbances such as "halo" phenomena do not occur.

Animals↗
PubMed1988

Myocardial turnover of plasma free fatty acids during angina pectoris induced by atrial pacing.

Myocardial extraction of free fatty acids (FFA), together with glucose, lactate, pyruvate, glycerol and oxygen was determined by simultaneous sampling of blood from an artery (a) and the coronary sinus (cs) at rest and during chest pains induced by atrial pacing in seven fasting male patients with ischaemic heart disease. Results were compared to those, at rest and during pacing at heart rate 140 beats min-1, in ten healthy men of similar age. A continuous i.v. infusion of 14C oleate and 3H palmitate enabled the calculation of simultaneous myocardial uptake and release of FFA as well as of the fraction of extracted FFA which underwent direct oxidation. During chest pain lactate net extraction decreased to become, in some patients, negative. FFA extraction, as estimated from the fractional extraction of labelled fatty acid was likewise decreased, while the a-cs O2 difference was not significantly altered. The fractional oxidation of extracted FFA was increased, whereas the calculated fatty acid release from the heart was unaltered. The increase in fractional oxidation was quantitatively correlated with the decrease in lactate extraction suggesting that it was related to the degree or extent of ischaemia. It was also proportional to the decrease in FFA extraction. Thus, in patients with angina pectoris the ischaemic myocardium may be subjected to a limitation not only with regard to oxygen but also substrate flux into the myocardial cells.

Angina Pectoris↗
PubMed1985

Clearance of triglycerides from the circulation and its relationship to serum lipoproteins: influence of age and sex.

In this study of a randomly selected population from 20 to 70 years of age, the clearance of triglycerides from the blood was studied after i.v. injection of an artificial triglyceride emulsion Intralipid. In women, the triglyceride clearance from the blood decreases with increasing age, but for men there was no change with age. Women had faster fractional removal rates of i.v. injected triglycerides than men in all age-groups. The triglyceride clearance showed a negative correlation to Broca index (obesity), serum triglyceride, serum cholesterol, triglycerides in very low density lipoproteins, and to cholesterol in very low density lipoproteins and in low density lipoproteins, but showed a strong positive correlation to cholesterol in high density lipoproteins.

Adult↗