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A Hallberg

Publications and source records attributed to A Hallberg.

At least 73 records · Page 4Linked to original sources
PubMed1992

Modulation of neutrophil superoxide generation by inhibitors of protein kinase C, calmodulin, diacylglycerol and myosin light chain kinases, and peptidyl prolyl cis-trans isomerase.

To assess the role of protein kinase C (PKC) in the respiratory burst of adherent human polymorphonuclear leukocytes (PMNL), reduction of ferricytochrome C by cells triggered with a phorbol ester (PMA), ionophore A23187, serum-treated zymosan (STZ) or three lipid derivatives, 3-decanoyl-sn-glycerol (G-3-OCOC9), (R,R)-1,4-diethyl-2-O-decyl-L-tartrate (Tt-2-OC10) and 3-decyloxy-5-hydroxymethylphenol (DHP) was examined in a microtiter plate procedure in the presence of inhibitors of PKC and, for comparison, inhibitors of calmodulin, diacylglycerol and myosin light chain kinases and the peptidyl-prolyl cis-trans isomerase activity of fujiphilin. 1) Of the protein kinase inhibitors examined, Ro 31-7549 and staurosporine reduced responses to all stimuli except possibly STZ; in contrast, K252a and the myosin light chain kinase inhibitors ML-7 and ML-9 blocked responses to A23187 and STZ better than those triggered by PMA. H-7 reduced responses to A23187, DHP and G-3-OCOC9, and calphostin, palmitoyl carnitine, sphingosine and the multifunctional drugs TMB-8 and W-7 reduced A23187; they also, when examined, reduced decane derivative-induced O2- production more effectively than PMA- and STZ-triggered responses. Polymyxin B, 4 alpha-PMA and retinal displayed no inhibitory capacity. 2) Of the selective calmodulin antagonists, CGS 9343B, Ro 22-4839 and calmidazolium did not inhibit the oxidative response irrespective of the stimulus used, whereas metofenazate reduced those evoked by A23187, DHP, G-3-OCOC9 and STZ.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Isomerases↗
PubMed1991

The correlation of the oxidation potentials of structurally related dibenzo[1,4]dichalcogenines to their antioxidance capacity in biological systems undergoing free radical-induced lipid peroxidation.

A series of structurally related dibenzo[1,4]dichalcogenines possessing similar lipid solubilities but greatly differing oxidation potentials were tested for their ability to inhibit stimulated lipid peroxidation in ADP/Fe2+/ascorbate-treated liver microsomes and in hepatocytes treated with either t-butylhydroperoxide or diquat. In general, there was a close correlation between the half-wave oxidation potential of a particular compound and its antioxidant activity in the microsome and cell systems, with compounds possessing the lowest potential being the most potent antioxidants and vice versa. The Te/O and Te/S and S/O-substituted compounds, with oxidation potentials between 0.65 and 0.87 V, demonstrated most potent activity. Above this potential the antioxidant activity of the structure declined rapidly. The Te/O and Te/S compounds are among the most potent synthetic antioxidants described possessing IC50 values in the microsome system lower than 0.5 microM. This study clarifies the critical role of redox potential of an antioxidant site on a particular molecule without the complication of variable lipid solubility and may allow the definition of an optimal potential for antioxidant activity in biological systems.

Animals↗
PubMed1991

Investigation of the possible dopaminergic toxicity of 1-methyl-3-phenyl-1,2,3,6-tetrahydropyridine, an isomer to the neurotoxin MPTP.

1-Methyl-3-phenyl-1,2,3,6-tetrahydropyridine (M-3-PTP) is an analogue to the Parkinson-producing dopaminergic toxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), M-3-PTP, and simple analogues thereof, are versatile intermediates in organic synthesis. The present study was undertaken to investigate the possible dopaminergic toxicity of M-3-PTP. Male albino mice were injected with 50 mg/kg of either MPTP or M-3-PTP and dopamine (DA) and its metabolites were determined 2 hr and 7 days after the administration. Two hr after MPTP profound acute changes in brain DA metabolism were found, i.e. an approximately 50% reduction in the concentration of DA together with a 10-fold increase in the level of 3-methoxytyramine. Seven days after MPTP, DA and metabolites were markedly reduced which is consistent with a degeneration of the dopaminergic neurones. In contrast M-3-PTP produced no acute or long-term alterations in the concentrations of DA and its metabolites in mouse brain. Furthermore, in vitro experiments show that M-3-PTP does not inhibit monoamine oxidase B. Thus, the present data show that M-3-PTP is devoid of dopaminergic toxicity in mouse brain and is not likely to produce Parkinson's disease in humans. The lack of toxicity is probably explained by the low affinity of M-3-PTP for monoamino oxidase B.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗
PubMed1990

Extracellular ATP increases cytoplasmic free Ca2+ concentration in clonal insulin-producing RINm5F cells. A mechanism involving direct interaction with both release and refilling of the inositol 1,4,5-trisphosphate-sensitive Ca2+ pool.

Effects of extracellularly applied ATP (added as disodium salt) on stimulus-secretion coupling were investigated in clonal insulin-producing RINm5F cells. Cytoplasmic free Ca2+ concentration [( Ca2+]i), electrical activity, membrane potential, formation of InsP3 and insulin release were measured. Addition of ATP in a Ca2(+)-containing medium promoted a rapid rise in [Ca2+]i, which was followed by a slow decline towards the basal level. In a Ca2(+)-free medium, the ATP-induced increase in [Ca2+]i was smaller, but still enough to elicit insulin secretion. Upon normalization of the extracellular Ca2+ concentration, the response to ATP recovered instantaneously. The presence of glucose in the incubation medium was a prerequisite to obtain a pronounced effect of ATP in the absence of extracellular Ca2+. However, glucose did not enhance the response to ATP in a Ca2(+)-containing medium. The effect of ATP was dose-dependent, with a clearly detectable increase in [Ca2+]i at 1 microM and a maximal response being obtained at 200 microM-ATP. The response to ATP was unaffected by activating adenylate cyclase by forskolin, but was abolished by 10 nM of the phorbol ester phorbol 12-myristate 13-acetate. The effects of ATP on [Ca2+]i could not be accounted for by a generalized increase in plasma-membrane permeability, as evident from the failure of the nucleotide to increase the fluorescence of the nuclear stain ethidium bromide. After stimulation with ATP there was an increase in membrane potential, in both the absence and the presence of extracellular Ca2+. Blockage of the voltage-activated Ca2+ channals with D-600, in a Ca2(+)-containing medium, decreased the effect of ATP on [Ca2+]i slightly. Patch-clamp measurements using the cell-attached patch configuration revealed that the RINm5F cells produce spontaneous action potentials, the frequency of which increased markedly on addition of ATP. Whole-cell recordings demonstrated that the increase in spike frequency was not associated with the development of an inward current, but was rather accountable for by a decrease in the activity of the ATP-regulated K+ channels. Addition of 200 microM-ATP stimulated phospholipase C activity, as evident from the formation of InsP3, both in the absence and in the presence of extracellular Ca2+. Thus in the absence of extracellular Ca2+ the stimulatory effect of ATP on insulin release can be explained by InsP3-induced mobilization of intracellularly bound Ca2+. Hence, in the RINm5F cells extracellular ATP acts in a manner similar to other Ca2(+)-mobilizing agents.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenosine Triphosphate↗
PubMed1990

Glutathione in human melanoma cells. Effects of cysteine, cysteine esters and glutathione isopropyl ester.

Thiols are of great importance for the regulation of many cellular functions including metabolism, transport and cell protection. In this study the usefulness of L-cysteine methyl and octyl esters, of N,S-diacetyl-L-cysteine methyl ester and glutathione isopropyl ester as cellular cysteine and GSH delivery systems was investigated in the human IGR 1 melanoma cell line. The L-cysteine methyl and octyl esters proved to be highly toxic to the cells. Treatment of the cultures with 1 mM N,S-diacetyl-L-cysteine methyl ester or 3 mM glutathione isopropyl ester for 24 h resulted in marked elevation of the cellular glutathione level without apparent or with slight cell loss, respectively. Thus the administration of the latter two compounds seems to be suitable for inducing GSH elevation in the cultured melanoma cells.

Acetylcysteine↗
PubMed1990

Coexpression of the platelet-derived growth factor (PDGF) B chain and the PDGF beta receptor in isolated pancreatic islet cells stimulates DNA synthesis.

Suspensions rich in pancreatic beta cells were transfected by means of electroporation or by using the liposome technique with DNA constructs coding for the B chain of platelet-derived growth factor (PDGF) and the PDGF alpha and beta receptors to induce a mitotic response in this slowly replicating cell type. Transfection with the B-chain construct induced synthesis of the PDGF B-chain homodimer (PDGF-BB) as assessed by the presence of 125I-labeled PDGF-BB competing activity in the conditioned medium of the transfected islet cells. Moreover, islet cells transfected with the PDGF beta-receptor construct exhibited increased immunofluorescence staining with a PDGF beta-receptor antibody. These cells also displayed increased 125I-labeled PDGF-BB binding compared with control transfected cells. Cotransfection with the B-chain construct or the addition of 10% fetal bovine serum or purified PDGF all induced DNA synthesis in islet cells transfected with the PDGF beta-receptor construct. Islet cells transfected with the PDGF alpha-receptor construct did not respond with stimulation of [3H]thymidine incorporation to any of the PDGF isoforms (PDGF-AA, -AB, or -BB). Cotransfection of the PDGF alpha- and beta-receptor constructs resulted in a loss of the DNA synthesis response to PDGF. The beta cells exhibited elevated levels of [3H]inositol trisphosphate after transfection with the B-chain and beta-receptor constructs, indicating activation of phospholipase C. Islet cells transfected with the different receptor constructs exhibited different patterns of tyrosine phosphorylation upon ligand activation. The results demonstrate that pancreatic islet cells can be stimulated to increase DNA synthesis by transfection with the PDGF beta-receptor gene, whereas cotransfection with the alpha-receptor gene may attenuate the growth response.

Animals↗
PubMed1990

Effects of long-term chloroquine exposure on the phospholipid metabolism in retina and pigment epithelium of the mouse.

The influence of 6 month's treatment of normal mice with chloroquine on neuroretina and retinal pigment epithelium has been investigated biochemically and morphologically. All classes of neuroretinal phospholipids, except lysophosphatidylcholine, showed increased 14C-glucose incorporation after chloroquine treatment. No metabolic changes were observed in the pigment epithelium after the chloroquine treatment. Morphological signs of phospholipidosis were only evident in the ganglion cells of the neuroretina. It is concluded that long-term treatment with chloroquine does not affect pigment epithelium phospholipid metabolism but leads to morphological and biochemical signs of phospholipidosis in the neuroretina.

Animals↗
PubMed1990

Synthesis of 3-dialkylaminochromans via thallium(III)-induced cyclization of allyl aryl ethers.

The sulfur-containing 3-alkylaminochromans, 5-methoxy-3-[N-(2- methylthioethyl)-propylamino]chroman (15), 5-hydroxy-3-[N-(2-methylthioethyl)propylamino]chroman (5) and 5-methoxy-8-methylthio-3-(dipropylamino)chroman (6), have been prepared from 8-bromo-5-methoxy-3-chromanol (11). This precursor was synthesized from 3-allyloxy-4-bromoanisole (8), by a thallium(III)-mediated ring-closure reaction. Compound 11 also served as starting material for the synthesis of 8-bromo-3-(dipropylamino)-5-methoxychroman (7).

Chromans↗
PubMed1989

Metabolism of N-acetyl-L-cysteine. Some structural requirements for the deacetylation and consequences for the oral bioavailability.

Rat liver, lung and intestine homogenates deacetylated N-acetyl-L-cysteine. Nearly stoichiometric amounts of L-cysteine were recovered. In rat liver, the enzyme activity was associated with the cytosolic fraction. Liver cytosol was much less active. N-Acetyl-D-cysteine or the disulphide of N-acetyl-L-cysteine were not deacetylated or in other ways consumed in vitro. Isolated, perfused rat liver did not retain or metabolize N-acetyl-L-cysteine to any measurable extent during single-pass experiments. N-Acetyl-L-cysteine or N-acetyl-D-cysteine were injected into a ligated segment of rat intestine in situ. After 1 hr 2% of the L-isomer and 35% of the D-isomer remained in the intestinal lumen. Systemic plasma levels were less than 3 microM of the L-form and congruent to 40 microM of the D-form. We conclude that deacetylation in the intestinal mucosa and possibly in the intestinal lumen is the major factor determining the low oral bioavailability of N-acetyl-L-cysteine. The deacetylation is discussed on the basis of the subcellular localization and the structural requirement of the reaction.

Acetylation↗
PubMed1989

Transfection of insulin-producing cells with a transforming c-Ha-ras oncogene stimulates phospholipase C activity.

Pancreatic islet beta-cells and insulin-producing RINm5F cells were electroporated in the presence of the c-Ha-ras oncogene, to assess the possible involvement of the encoded product in coupling extracellular receptors to phospholipase C. After two days the c-Ha-ras-transfected cells increased their expression of c-Ha-ras mRNA. These cells were also found to contain more [3H]InsP3, suggesting an increased basal (non-ligand-activated) phospholipase C activity. In addition, the transfected cells were unable to respond to ligand (bombesin) activation of phospholipase C. The ras-transfected insulin-producing cells showed enhanced phosphorylation of a 200 kDa substrate crossreacting with an antibody to an 80 kDa protein kinase C substrate. The phorbol ester 12-O-tetradecanoyl 13-acetate and bombesin also induced phosphorylation of the 200 kDa substrate. All of these changes occurred without changes in the rates of [3H]thymidine incorporation. The results suggest that the mutated c-Ha-ras oncogene directly or indirectly stimulates the basal phospholipase C activity of these cells.

Animals↗
PubMed1989

Plasma membrane associated ATP as a regulator of the secretory activity of the pancreatic beta-cell.

beta-Cell-rich pancreatic islets from ob/ob-mice were used for evaluating how ATP associated with the plasma membrane participates in the regulation of insulin release. Increase of Ca2+ initiates insulin release from permeabilized beta-cells only in the presence of Mg-ATP. When bound to the inner part of the plasma membrane ATP depolarizes the beta-cells by closing a glucose-regulated K+-channel. It is possible that ATP in a plasma membrane compartment modulates insulin release also by stimulating ion pumps and exchange processes. ATP can regulate the secretory activity by binding also to the exterior of the beta-cells. The addition of ATP resulted in stimulation of insulin release related to polyphosphoinositide breakdown. It is suggested that the granule fusion with the plasma membrane is followed by release of sufficient amounts of ATP and ADP for activating a P2-purinoceptor. This receptor may consequently be part of a system for amplifying the secretory response to glucose and other agents facilitating the entry of Ca2+.

Adenosine Triphosphate↗
PubMed1989

Differential sensitivity to beta-cell secretagogues in cultured rat pancreatic islets exposed to human interleukin-1 beta.

The early stages of insulin-dependent diabetes mellitus are characterized by a selective inability to secrete insulin in response to glucose, coupled to a better response to nonnutrient secretagogues. The deficient glucose response may be a result of the autoimmune process directed toward the beta-cells. Interleukin-1 (IL-1) has been suggested to be one possible mediator of immunological damage of the beta-cells. In the present study we characterized the sensitivity of beta-cells to different secretagogues after human recombinant IL-1 beta (rIL-1 beta) exposure. Furthermore, experiments were performed to clarify the biochemical mechanisms behind the defective insulin response observed in these islets. Rat pancreatic islets were isolated and kept in tissue culture (medium RPMI-1640 plus 10% calf serum) for 5 days. The islets were subsequently exposed to 60 pM human recombinant IL-1 beta during 48 h in the same culture conditions as above and examined immediately after IL-1 exposure. The rIL-1 beta-treated islets showed a marked reduction of glucose-stimulated insulin release. Stimulation with arginine plus different glucose concentrations, and leucine plus glutamine partially counteracted the rIL-1 beta-induced reduction of insulin release. The activities of the glycolytic enzymes hexokinase, glucokinase, and glyceraldehyde 3-phosphate dehydrogenase, were similar in control and IL-1-exposed islets. Treatment with IL-1 also did not impair the activities of NADH+- and NADPH+-dependent glutamate dehydrogenase, glutamate-aspartate transaminase, glutamate-alanine transaminase, citrate synthase, and NAD+-linked isocitrate dehydrogenase. The oxidation of D-[6-14C]glucose and L-[U-14C]leucine were decreased by 50% in IL-1-treated islets. Furthermore, there was a significant decrease in the ratios of [2-14C]pyruvate oxidation/[1-14C]pyruvate decarboxylation and L-[U-14C]leucine oxidation/L-[1-14C]leucine decarboxylation, indicating that IL-1 decreases the proportion of generated acetyl-coenzyme-A residues undergoing oxidation. However, in the presence of IL-1 there was a significant increase in L-[U-14C]glutamate oxidation. These combined observations suggest that exposure to IL-1 induces a preferential decrease in glucose-mediated insulin release and mitochondrial glucose metabolism. This mitochondrial dysfunction seems to reflect an impairment in proximal steps of the Krebs cycle. It is conceivable that the IL-1-induced suppression and shift in islet metabolism can be an explanation for the beta-cell insensitivity to glucose observed in the early phases of human and experimental insulin-dependent diabetes mellitus.

Animals↗
PubMed1989

Human interleukin 1 beta stimulates islet insulin release by a mechanism not dependent on changes in phospholipase C and protein kinase C activities or Ca2+ handling.

Isolated islets from adult rats or obese hyperglycemic (ob/ob) mice were incubated with human recombinant interleukin 1 beta in order to study whether the acute effects of the cytokine on islet insulin release are associated with changes in islet phospholipase C activity, Ca2+ handling or protein phosphorylation. The cytokine stimulated insulin release both at low and high glucose concentrations during one hour incubations. In short-term incubations (less than 1 min) interleukin 1 beta did not affect the production of inositoltrisphosphate. Addition of interleukin 1 beta affected neither the cytoplasmic free Ca2+ concentration at rest nor that observed subsequent to stimulation with a high concentration of glucose. Furthermore, the endogenous protein kinase C activity, as visualized by immunoprecipitation of a 32P-labelled substrate for this enzyme, was not altered by interleukin 1 beta. Separation of 32P-labelled proteins by means of 2-dimensional gel electrophoresis failed to reveal any specific effects of the cytokine on the total protein phosphorylation activity. These results suggest that the stimulatory effects on insulin release exerted by interleukin 1 beta are not caused by acute activation of phospholipase C and protein kinase C or by an alteration of islet Ca2+ handling of the B-cells.

Animals↗
PubMed1989

High-glucose stimulation of 64,000-Mr islet cell autoantigen expression.

This study examined the effect of various extracellular glucose concentrations on the expression of a previously described 64,000-Mr islet cell autoantigen associated with insulin-dependent diabetes mellitus. The protein was precipitated from patient serums incubated with Triton X-100 lysates of [35S]methionine-labeled rat pancreatic islets that had been cultured in 5, 11, or 28 mM glucose for 6 h or 3 days. In both types of experiment, 28 mM glucose was the most efficient stimulator of 64,000-Mr autoantigen production. In contrast, the class I antigens of the major histocompatibility complex, precipitated by a rabbit polyclonal antiserum, were not influenced by differences in glucose concentrations. Our data indicate that expression of islet cell antigens may be increased during the course of hyperglycemia and suggest that the functional activity of islet cells influences their antigenicity.

Animals↗
PubMed1988

Use of liposomes to introduce substances into pancreatic islet cells.

The liposome technique is widely used to transport substances that cannot normally traverse the plasma membrane into the cell. The interactions of liposomes with the plasma membrane of pancreatic islet cells have not previously been studied. We evaluate the suitability of the liposome technique for introducing substances into the pancreatic beta-cell to which the cell membrane is impermeable. Liposomes were synthesized with an ether-injection method, and the cell-liposomal interactions were investigated by means of radioactive labeling and the fluorescent aqueous space marker 6-carboxyfluorescein. Experiments were performed on freshly isolated mouse pancreatic islets and on free islet cell preparations. With fluorescence microscopy, liposomes were observed to fuse spontaneously with islet cells, and the corresponding internalized volumes were quantified with spectrofluorometric measurements. The liposome association with islets and islet cell suspensions, as assessed by radioactive labeling, was found to increase with the liposome concentration. The effects of liposome membrane lipid composition on the fusion rate were found to be decreased in the presence of glucolipid. In addition, polyethylene glycol failed to affect the liposomal uptake. Freshly isolated islets incubated with liposomes containing glucose 6-phosphate were observed to release slightly more insulin than islets incubated with "empty" liposomes. In conclusion, liposomes fuse spontaneously with islet cells in vitro, and the uptake of liposomes is regulated by the lipid composition of the liposomal bilayer and the amount of liposomes present. The function of the beta-cell can be altered with the liposome technique, e.g., by addition of biologically active molecules such as glucose 6-phosphate.

Animals↗
PubMed1987

Characterization of the inositol 1,4,5-trisphosphate-induced Ca2+ release in pancreatic beta-cells.

Pancreatic beta-cells isolated from obese-hyperglycaemic mice released intracellular Ca2+ in response to carbamoylcholine, an effect dependent on the presence of glucose. The effective Ca2+ concentration reached was sufficient to evoke a transient release of insulin. When the cells were deficient in Ca2+, the Ca2+ pool sensitive to carbamoylcholine stimulation was equivalent to that released by ionomycin. Unlike intact cells, cells permeabilized by high-voltage discharges failed to generate either inositol 1,4,5-triphosphate (InsP3) or to release Ca2+ after exposure to carbamoylcholine. However, the permeabilized cells released insulin sigmoidally in response to increasing concentrations of Ca2+. Also in the absence of functional mitochondria these cells exhibited a large ATP-dependent buffering of Ca2+, enabling the maintenance of an ambient Ca2+ concentration corresponding to about 150 nM even after several additional pulses of Ca2+. InsP3, maximally effective at 6 microM, promoted a rapid and pronounced release of Ca2+. The InsP3-sensitive Ca2+ pool was rapidly filled and lost its Ca2+ late after ATP depletion. The transient nature of the Ca2+ signal was not overcome by repetitive additions of InsP3. It was possible to restore the response to InsP3 after a delay of approx. 20 min, an effect which had less latency after the addition of Ca2+. These latter findings argue against degradation and/or desensitization as factors responsible for the transiency in InsP3 response. It is suggested that Ca2+ released by InsP3 is taken up by a part of the endoplasmic reticulum (ER) not sensitive to InsP3. On metabolism of InsP3, Ca2+ recycles to the InsP3-sensitive pool, implying that this pool indeed has a very high affinity for the ion. The presence of functional mitochondria did not interfere with the recycling process. The ER in pancreatic beta-cells is of major importance in buffering Ca2+, but InsP3 only modulates Ca2+ transport for a restricted period of time following immediately upon its formation. Thereafter the non-sensitive part of the ER takes over the continuous regulation of Ca2+ cycling.

Animals↗
PubMed1986

Hydatidiform moles: methods for culture and cytogenetic analyses.

Placental tissue with macroscopic visible vesicles was processed for chromosome analysis. Enzymatic tissue dissociation, culture initiation in Chang's medium, and final culture in a medium with high glucose content were used, which increased the success rate for karyotyping from 55% to 93%. Median culture time necessary for obtaining a karyotype decreased to about 1 week, irrespective of the chromosomal constitution. This was achieved in spite of increasing time for transportation. The metaphases obtained permitted detailed analysis of chromosomal polymorphisms. In 8 of 44 cases, the karyotype was established on short-term culture alone, which proved to be a valuable supplement in this study.

Cells, Cultured↗