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Biomedical subjects

M Sandberg

Publications and source records attributed to M Sandberg.

At least 73 records · Page 4Linked to original sources
PubMed1995

Differential effects of N-methyl-D-aspartate on Ca2+ homeostasis in developing and adult rat striatum: in vivo microdialysis approach.

This in vivo study concerns developmental differences in the sensitivity of striatal neurons to N-methyl-D-aspartate (NMDA). Changes in calcium homeostasis in adult vs immature rats at postnatal days 8-10, evoked by NMDA, were evaluated by measurements of 45Ca efflux and of Ca2+ taurine and phosphoethanolamine concentrations in striatal microdialysates. The efflux of [14C]sucrose was employed in order to measure changes in extracellular space volume. In adult rats the addition of 5 mM NMDA for 20 min to the perfusion medium resulted in a 30-40% increase in 45Ca efflux, and in a 15% increase in [14C]sucrose efflux. Ten minutes after NMDA perfusion, 45Ca and [14C] sucrose efflux returned to the baseline. No significant changes in Ca2+ or amino acid concentrations were observed in the dialysate of the adult rat striatum. NMDA perfusion in the striatum of immature rats initially induced a transient (5 min) increase in the efflux of 45Ca (by 13%) and [14C]sucrose (by 9%). This was followed by a prolonged (lasting 45-50 min) 45% decrease in 45Ca efflux, an 80% reduction of Ca2+ concentration, and increases in taurine and phosphoethanolamine concentrations in the dialysate, whereas [14C]sucrose efflux recovered within 10 min. These data illustrate differences in the NMDA response between developing and adult rat striatum. Only in developing rats did NMDA induce a large and prolonged influx of extracellular calcium to neurons that may explain the enhanced NMDA neurotoxicity in immature rats.

Aging↗
PubMed1995

Phenotypic modification of human osteosarcoma cells with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate.

Treatment of the U-2 OS human osteosarcoma cell line with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) dramatically decreased the rate of DNA synthesis. This decrease in proliferation as well as the change in morphology of the TPA-treated cells can be blocked by the protein kinase C inhibitor GF 109203X. The U-2 OS cells are known to express the c-sis oncogene [platelet-derived growth factor (PDGF) B-chain], PDGF-A, and receptors for PDGF, thus providing a potential autocrine loop of growth stimulation. TPA was found to induce the expression of both the PDGF-A and the PDGF-B chains. However, the levels of the PDGF receptor beta subunits and of the PDGF-BB inducable tyrosine phosphorylation of the PDGF receptor were markedly reduced. The TPA treatment of the U-2 OS cells also induced changes typical for maturing bone cells, such as increased expression levels of alkaline phosphatase and osteopontin. The expression levels of type I collagen and bone sialoprotein were reduced. The results show a TPA-dependent down-regulation of the PDGF receptor beta subunits that correlates with an increased expression of osteoblast phenotypic markers.

Cell Division↗
PubMed1994

Liquid chromatographic determination of acidic beta-aspartyl and gamma-glutamyl peptides in extracts of rat brain.

This work describes further development of our previously presented method for determination of acidic sulfur/phosphor-containing amino acids, gamma-glutamyl di/tripeptides, and beta-aspartyl dipeptides. Automated precolumn fluorogenic derivatization was performed with o-phthaldialdehyde/beta-mercaptoethanol and the derivatives were separated by reversed-phase liquid chromatography. The method was optimized for the analysis of brain tissue extracts. Due to the complex sample matrix, three separation schemes with complementary selectivities were developed. Different extraction protocols were evaluated and sonication of frozen tissue powder in methanol-H2O (9:1, v/v) yielded the highest recoveries and precision. beta-Mercaphtoethanol and EDTA were added to the extraction media to inhibit spontaneous oxidation of thiol-containing amino compounds. Analyte identification was based on retention times and recovery of standards added to extracts. The following compounds were identified in rat cerebral cortex (mean tissue concentration +/- SD, n = 6): gamma-glutamylglutamine (38.5 +/- 12.6 microM), gamma-glutamylglutamate (14.4 +/- 6.0 microM), gamma-glutamyltaurine (4.9 +/- 2.2 microM), beta-aspartylglycine (4.0 +/- 0.4 microM), beta-aspartyltaurine (3.7 +/- 0.6 microM), O-phosphoserine (3.2 +/- 0.8 microM), gamma-glutamylcysteine (1.9 +/- 0.3 microM), gamma-glutamylglycine (1.1 +/- 0.1 microM), and gamma-glutamylcysteateglycine (0.8 +/- 0.1 microM). In addition over 15 unidentified components were found. Cysteate, cysteine sulfinate, homocysteate, homocysteine sulfinate, O-Sulfoserine, gamma-glutamylaspartate, gamma-glutamylcysteate, gamma-glutamylhistidine, and beta-aspartylalanine were not present at concentrations above 1 microM.

Animals↗
PubMed1994

Structure of dodecyl sulfate-protein complexes at subsaturating concentrations of free detergent.

Earlier neutron small-angle scattering experiments had revealed the low resolution structure of the complex between sodium dodecyl sulfate (SDS) and the single polypeptide (452 amino acid residues) of a water-soluble enzyme. The saturated complex consists of three globular micelles which are connected by short flexible polypeptide segments. New experiments, described here, were performed at subsaturating concentrations of free SDS in equilibrium with the complex. The data show a decrease in stoichiometry from one bound dodecyl sulfate (DS) anion per two amino acid residues near the critical micelle concentration (CMC) to one per four residues at half the CMC. At 0.3 CMC, a two-micelle complex is formed by the recombination of the small amino-terminal micelle with the middle one; and the center-to-center distance between the carboxyl-terminal micelle and the middle one decreases from 7.5 to 6.2 nm. These structural data allow us to better understand earlier results obtained with high-performance agarose gel chromatography of the same SDS-protein complexes.

Journal Article↗
PubMed1994

Increased intra- and extracellular concentrations of gamma-glutamylglutamate and related dipeptides in the ischemic rat striatum: involvement of glutamyl transpeptidase.

The present work relates to the possibility that the ATP-independent enzyme gamma-glutamyl transpeptidase (EC 2.3.2.2), which has been postulated to be part of an amino acid uptake system, is active during cerebral ischemia. This was evaluated in the ischemic rat striatum by determination of intra- and extracellular concentrations of gamma-glutamyl dipeptides (the products of the transpeptidation) and glutathione (the physiological gamma-glutamyl donor). An ischemic period (0-30 and 31-60 min) resulted in prominent increases in the respective concentration of extracellular gamma-glutamylglutamate (24- and 67-fold), gamma-glutamyltaurine + gamma-glutamylglycine (5.8- and 19-fold), and gamma-glutamylglutamine (2.6- and 6.8-fold) as revealed using in vivo microdialysis. The changes coincided with increased respective extracellular concentrations of glutamate (83- and 115-fold), taurine (17- and 25-fold), glycine (4.6- and 6.1-fold), and glutamine (1.7- and 2.1-fold). Furthermore, under anoxic conditions in vitro (0-30 and 0-60 min), respective striatal tissue concentrations were increased for gamma-glutamylglutamate (20- and 17-fold), gamma-glutamyltaurine (6.7- and 11-fold), gamma-glutamylglutamine (1.7- and 1.2-fold), and gamma-glutamylglycine (14- and 18-fold), whereas glutathione levels were, on an average, decreased by approximately 350 microM. In summary, gamma-glutamyl transpeptidase is involved in de novo dipeptide synthesis in the mammalian brain during anoxic conditions, indicating transport of amino acids such as glutamate.

Amino Acids↗
PubMed1994

Dietary supplementation with very long-chain n-3 fatty acids in patients with atopic dermatitis. A double-blind, multicentre study.

The purpose of this study was to investigate whether fish oil and/or corn oil had a beneficial effect on the clinical state of atopic dermatitis, and to evaluate the dietary intake of nutrients in this group of patients. In a double-blind, multicentre study lasting 4 months, during wintertime, 145 patients with moderate to severe atopic dermatitis were randomly assigned to receive either 6 g/day of concentrated n-3 fatty acids, or an isoenergetic amount of corn oil. As local treatment, only an emollient cream or hydrocortisone cream was allowed. The fatty acid pattern in serum phospholipids, and the dietary intake of nutrients were monitored in a subgroup of patients, and the results were compared with a group of patients with psoriasis. The overall clinical score, as evaluated by the physicians, improved during the trial by 30% in the fish oil (P < 0.001) and 24% in the corn oil group (P < 0.001). This was also consistent with the results from a selected skin area, and it was further confirmed by the total subjective clinical score reported by the patients. There were no significant differences in the clinical scores between the two groups at baseline, and at the end of the study. In the fish oil group, the amount of n-3 fatty acids in serum phospholipids was significantly increased at the end of the trial, compared with pretreatment values (P < 0.001), whereas the level of n-6 fatty acids was decreased (P < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1994

Expression of ICAM-1 and HLA class II in acute cellular and vascular rejection of human kidney allografts.

We studied the relationship between ICAM-1 and class II expression on graft tubular cells and the relationship with graft inflammation in 50 kidney transplants monitored with serial aspiration biopsies after transplantation. Of the 50 grafts, 26 had an acute rejection 17 +/- 10 days after transplantation, 5 also had acute vascular rejection (AVR) and 24 had no rejection. The initial post-transplant ICAM-1 and class II expression was low in all grafts. All 21 grafts with acute cellular rejection (ACR) displayed ICAM-1 induction, with a peak at the beginning of acute blastogenic rejection and declining over 20 days to prerejection levels. Class II expression reached a peak later and also declined later to prerejection levels. In the grafts with irreversible AVR both ICAM-1 and class II expression remained elevated. The 24 grafts with no rejection displayed no ICAM-1 or class II induction on tubular cells during the follow-up. The differences between ICAM-1 and classs II expression in biopsies with rejection and with no rejection were statistically significant. The results demonstrate that ICAM-1 was induced early during ACR on the graft tubular cells and that it disappeared rapidly in reversible rejections. The induction of class II antigens was slightly slower but quantitatively greater. In the irreversible rejections with a combination of ACR and AVR both ICAM-1 and class II expression remained elevated.

Acute Disease↗
PubMed1994

Human cyclic GMP-dependent protein kinase I beta overexpression increases phosphorylation of an endogenous focal contact-associated vasodilator-stimulated phosphoprotein without altering the thrombin-evoked calcium response.

The role of the cGMP-dependent protein kinase (cGK) and one of its major substrates, the vasodilator-stimulated phosphoprotein (VASP), in the regulation of a receptor-evoked calcium response was investigated. The human type I beta cGK was stably transfected in human embryonic kidney 293 cells and Swiss mouse 3T6 fibroblasts, which contained significant or no detectable levels of the focal adhesion protein VASP, respectively. Western blot analysis and protein kinase activity measurements demonstrated an 8-fold overexpression of cGK-I beta in 293 cells (7-fold in 3T6 cells), representing an intracellular cGK concentration of 0.33 microM. In experiments with intact 293 cells expressing cGK-I beta, beta-phenyl-1,N2-etheno-cGMP and 8-(p-chlorophenylthio)-cGMP were capable of converting up to 30-40% of the 46-kDa VASP to its 50-kDa phospho- form, equivalent to results observed with cGMP analogs that cause a marked inhibition of the stimulated Ca2+ transient in intact human platelets. In contrast to platelets, preincubation of fura-2-loaded 293 and 3T6 cells with 8-(p-chlorophenylthio)-cGMP did not significantly inhibit thrombin-evoked calcium transients, although sufficient cGK-mediated VASP phosphorylation was clearly detectable under these conditions in cGK-I beta-expressing 293 cells. These results demonstrate that cGK inhibition of agonist-evoked calcium mobilization is not a mechanism common to all cell types and that VASP phosphorylation may not be an essential or sufficient component of the cGK effect on calcium levels. In contrast, the observed VASP phosphorylation mediated by recombinant human cGK-I beta in intact 293 cells does support the hypothesis that focal adhesions and their associated proteins are important cellular sites of cGK action.

Blood Proteins↗
PubMed1993

Cysteine sulphinate and cysteate: mediators of cysteine toxicity in the neonatal rat brain?

Excitotoxic amino acids contain two acidic groups, but cysteine represents an exception to this rule. The hypothesis that cysteine toxicity is mediated by the oxidized and diacidic metabolites cysteine sulphinate and/or cysteate was tested in the present study. The issue was approached in three different ways. Firstly, the distribution of brain injury after subcutaneous administration of cysteine (1 mg/g) to 4-day-old rats was compared with that caused by cysteine sulphinate (3 mg/g). Secondly, the effects of excitatory amino acid receptor antagonists on cysteine and cysteine sulphinate toxicity were investigated. Thirdly, the cerebral concentrations of cysteine sulphinate were determined after cysteine administration and compared with those obtained after cysteine sulphinate injection. The cerebral cortex was the region most vulnerable to cysteine toxicity, followed by the hippocampus (especially the medial subicular neurons), amygdala, caudoputamen, cerebellum and septum. Pronounced extravasation of red blood cells was observed in lesioned areas. One day after cysteine administration, the injury was infarction-like and sharply demarcated. Cysteine sulphinate-induced damage resembled cysteine-induced lesions in some respects: the anterior cingulate and retrosplenial cortices, as well as medial subicular cells, were quite vulnerable. However, the differences prevailed. Cysteine sulphinate, but not cysteine, killed neurons of the superficial part of the tectum, the medial habenula, the ventromedial hypothalamus and the arcuate nucleus. Further, while cysteine toxicity was prominent in deep cortical layers, cysteine sulphinate preferentially damaged superficial cortical neurons. Cysteine toxicity was abolished by pretreatment with MK-801, a selective NMDA antagonist, but not by 2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo(F)quinoxaline, a selective AMPA receptor blocker. In contrast, the considerably smaller lesion seen after cysteine sulphinate administration was only partially prevented by MK-801. Large (19-fold) increases in cortical cysteine sulphinate concentration were noted after injection of a toxic dose of cysteine. This corresponds to 90 nmol cysteine sulphinate/g protein. The cysteate concentration was not increased above the detection limit. Injection of a toxic dose of cysteine sulphinate elevated cysteine sulphinate concentration in the frontomedial cortex (a region consistently injured by cysteine sulphinate) almost three orders of magnitude more than that observed after cysteine administration. Taken together, these results strongly suggest that neither cysteine sulphinate nor cysteate alone mediate cysteine toxicity.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acids↗
PubMed1993

Effect of dietary supplementation with very-long-chain n-3 fatty acids in patients with psoriasis.

BACKGROUND: In several studies dietary fish oil has been found to have beneficial effect on psoriasis, but the results are contradictory and based mainly on open studies or studies of small numbers of patients. METHODS: In a four-month double-blind, multicenter trial, we randomly assigned 145 patients with moderate-to-severe psoriasis to receive in their diet either highly purified ethyl esters of n-3 fatty acids ("fish oil"; 6 g of oil per day, containing 5 g of eicosapentaenoic and docosahexaenoic acid) or an isoenergetic amount of corn oil containing mainly n-6 fatty acids. All the patients were advised to reduce their intake of saturated fatty acids. A 48-hour dietary recall was performed, and the fatty-acid pattern in the serum phospholipids was monitored in a subgroup of patients. RESULTS: In the fish-oil group, n-3 fatty acids were increased in serum phospholipids (P < 0.001), the ratio of arachidonic acid to eicosapentaenoic acid decreased (P < 0.001), and the level of n-6 fatty acids decreased (P < 0.001). In the corn-oil group, only docosahexaenoic acid increased significantly (P < 0.05). The ratio of polyunsaturated to saturated fatty acids increased in both groups. Plasma concentrations of triacylglycerol decreased from base line in the fish-oil group (P < 0.05). The score on the Psoriasis Area and Severity Index, as evaluated by the physicians, did not change significantly during the trial in either group. This was also true of a total subjective score reported by the patients, but a selected area of skin in the corn-oil group showed a significant reduction in the clinical signs (P < 0.05). Scaling was reduced from base line in both groups (P < 0.01). The fish-oil group had less cellular infiltration (P < 0.01), and the corn-oil group had improvement in desquamation and redness (P < 0.05). There was no significant difference in clinical manifestations between the groups. Among the patients in the fish-oil group, an increase in the concentration of n-3 fatty acids in serum phospholipids was not accompanied by clinical improvement, whereas in the corn-oil group there was a significant correlation between clinical improvement and an increase in eicosapentaenoic acid and total n-3 fatty acids. CONCLUSIONS: Dietary supplementation with very-long-chain n-3 fatty acids was no better than corn-oil supplementation in treating psoriasis. Clinical improvement was not correlated with an increase in the concentration of n-3 fatty acids in serum phospholipids among the patients in the fish-oil group, whereas there was a significant correlation between clinical improvement and an increase in eicosapentaenoic acid and total n-3 fatty acids in the corn-oil group.

Adult↗
PubMed1993

Gamma-L-glutamyl-L-glutamate is an endogenous dipeptide in the rat olfactory bulb which activates N-methyl-D-aspartate receptors.

The concentration of gamma-L-glutamyl-L-glutamate (gamma-glu-glu), a potent displacer of excitatory amino acid receptor binding, and other structurally related dipeptides were determined in extracts of the rat olfactory bulb by a novel liquid chromatographic method. Whole-cell patch clamp recordings of currents from freshly isolated neurons showed that gamma-glu-glu produced inward currents at negative holding potentials, provided microM concentrations of glycine were added and no Mg(2+)-ions were present, indicating activation of N-methyl-D-aspartate receptors. Consistently, geometry optimization of gamma-glu-glu using molecular mechanics calculations, suggested a bent conformer with structural features supporting this view.

Animals↗
PubMed1993

Electrochemical detection of dipeptides with selectivity against amino acids.

Electrolysis of a basic mobile phase containing biuret reagent [Cu(II) and a tartrate salt] at high (> 1.2 V vs. Ag/AgCl) potentials modifies the glassy carbon electrode. This modified anode oxidizes dipeptides, yielding signals expected for a one-electron transfer, even at low (down to 0.7 V vs. Ag/AgCl) potentials and in the absence of intentionally added copper(II) ion in the reagent or mobile phase. The same modification demonstrates a selectivity to alpha-dipeptides over amino acids that is unprecedented. The product of the anodic reaction is reduced at a downstream cathode at low positive potentials. Sensitivities for several amino acids and dipeptides are reported under several conditions. Neither the anodic nor the cathodic signals for the buiret complex of the tripeptide Ala-Ala-Ala are significantly altered because of the modification.

Amino Acid Sequence↗
PubMed1993

Isolation and characterization of a human pseudogene for the regulatory subunit RI alpha of cAMP-dependent protein kinases and its sublocalization on chromosome 1.

Using a human cDNA (complementary DNA) encoding the regulatory subunit RI alpha of cAMP-dependent protein kinases (PKA) as a probe, a pseudogene for this PKA isoform was isolated from a human genomic library. The human RI alpha pseudogene was 89% similar to the open reading frame of the expressed human RI alpha at the nucleotide level. Several stop codons were found, indicating that the pseudogene does not encode a functional protein. The pseudogene also contained several frameshift mutations, small deletions, and insertions. No introns were found in the region corresponding to the open reading frame of the expressed RI alpha cDNA. Specific oligonucleotides for the RI alpha gene and pseudogene were constructed and used as primers in polymerase chain reaction (PCR) to amplify human DNA from healthy blood donors. Probing of the PCR products using oligonucleotides specific for the RI alpha gene and RI alpha pseudogene, respectively, showed the presence of both genes in the human genome. When DNA extracted from various somatic cell hybrids was amplified, it was shown that the RI alpha gene was located on human chromosome 17, whereas the RI alpha pseudogene was located in the p21-p31 region on chromosome 1. This is the first report describing a pseudogene for a subunit of cAMP-dependent protein kinases from any species.

Amino Acid Sequence↗
PubMed1993

Transient expression of type III collagen by odontoblasts: developmental changes in the distribution of pro-alpha 1(III) and pro-alpha 1(I) collagen mRNAs in dental tissues.

The expression of pro-alpha 1(III) and pro-alpha 1(I) collagen mRNAs in mouse and human dental tissues during tooth development and after its completion was analyzed by in situ hybridization, with use of [35S]-labeled RNA probes. The expression of pro-alpha 1(III) mRNA was also compared to that of the protein product, as localized by immunostaining with polyclonal antibodies to type III collagen and the N-terminal propeptide of type III procollagen. Contrary to many previous reports, our results suggest that odontoblasts express type III collagen. While pro-alpha 1(III) transcripts were less intensely expressed in odontoblasts than pro-alpha 1(I) transcripts, the amounts of both mRNAs increased in odontoblasts with progressing dentin formation, and decreased toward its completion. In contrast to pro-alpha 1(III) mRNA, pro-alpha 1(I) mRNA was still detectable in odontoblasts of fully developed teeth. Type III collagen immunoreactivity was observed in the early predentin, and again in predentin toward the completion of dentinogenesis, when mRNA was no longer detected. Also in the pulp, the protein product, unlike pro-alpha 1(III) mRNA, was relatively strongly expressed. Hence, these immunostaining patterns were inversely related to the expression of pro-alpha 1(III) mRNA, suggesting accumulation of the protein. The mesenchymal cells, when condensed in the region of the future mandibular bone, expressed pro-alpha 1(III) mRNA intensely, whereas osteoblasts expressed pro-alpha 1(I) but not pro-alpha 1(III) transcripts strongly. Cell type- and developmental stage-related differences in the expression of the two mRNAs suggest that type I/type III collagen ratio influences the structure of dental tissues.

Aging↗
PubMed1993

Expression of types I, III and IV collagen genes in fibrotic skin and nerve lesions of toxic oil syndrome patients.

We have studied the skin and nerve fibrosis in toxic oil syndrome by in situ hybridization using specific cDNA probes for types I, III, and IV collagens. Fibroblasts with high levels of type I and III collagen mRNA were observed in biopsies from fibrotic skin areas. Similarly, type IV collagen mRNA was abundant in cells within the fibrotic process of the nerves. These results suggest that the excessive accumulation of collagen in toxic oil syndrome results from transcriptional activation of collagen genes in a subpopulation of fibroblasts.

Autoradiography↗
PubMed1993

Excitatory amino acids in the cerebrospinal fluid of asphyxiated infants: relationship to hypoxic-ischemic encephalopathy.

Asphyxiated (n = 27) and control infants (n = 25) were subjected to spinal taps. Amino acids were measured with liquid chromatography and the degree of hypoxic-ischemic encephalopathy was determined in each case. In asphyxiated infants, the concentrations of aspartate and glutamate were 286% and 387% (p < or = 0.01 and p < or = 0.05) of the control values, respectively. The cerebrospinal fluid aspartate levels were significantly (p < or = 0.05) higher in the group with severe (3.4 mumol/l) compared with the group with mild hypoxic-ischemic encephalopathy (1.0 mumol/l). Glutamate was also higher in the group with severe (12.3 mumol/l) than in the groups with mild (2.7 mumol/l) or moderate (3.2 mumol/l) hypoxic-ischemic encephalopathy (p < or = 0.05). High concentrations of excitatory amino acids were present in the CSF of asphyxiated infants which may exert excitotoxic effects.

Amino Acids↗
PubMed1992

Patterns of expression of the six alternatively spliced exons affecting the structures of the COL1 and NC2 domains of the alpha 1(XIII) collagen chain in human tissues and cell lines.

Reverse transcription-polymerase chain reactions and RNA protection experiments were used to examine alternative splicing of the six exons 3B-5, 12, and 13 affecting the COL1 and NC2 domains of type XIII collagen in seven human tissues and four cell lines. Distinct differences in the proportions of the variant mRNAs were found. With respect to the COL1 domain, all studied samples contained mRNAs corresponding to the shortest COL1 variants of 57 and 66 residues, with the former variant being prominent in most samples. Most of the samples also contained notable amounts of mRNAs that corresponded to the longest COL1 variants, mainly those of 104 and 95 residues. Particularly the extent of inclusion of exon 12 and 13 sequences, encoding most of the NC2 domain, varied according to the type of tissue or cell analyzed. Bone, cartilage, and colon adenocarcinoma samples contained little or none of the mRNAs corresponding to the long NC2 variants, whereas in fibroblast, lung, muscle, and osteosarcoma cells, those mRNAs were the major variants. The relative proportions of the various combinations of exons 3B-5, 12, and 13 were evaluated in four of the RNA samples. Interestingly, each of these samples appeared to contain only one to three major combinations of the six exons, representing about 40% to nearly 100% of all variants.

Alternative Splicing↗