Biomedical subjects

A Hallberg

Publications and source records attributed to A Hallberg.

At least 55 records · Page 3Linked to original sources

PubMed1997

Design, synthesis, and biological activities of four angiotensin II receptor ligands with gamma-turn mimetics replacing amino acid residues 3-5.

Disulfide cyclization is a powerful method for reducing the conformational space of a peptide. This in turn may enable the study of its bioactive conformation. Several analogues of angiotensin II (Ang II) containing a disulfide bridge between amino acids 3 and 5 have been reported. Among these the cyclic octapeptides c[Hcy3,5]-Ang II, c[Cys3,5]-Ang II, and c[Pen 3,5]-Ang II showed significant activity at Ang II receptors. We have performed conformational analysis studies using theoretical calculations and 1H-NMR spectroscopy on tripeptide model compounds of these cyclic octapeptides which show that the cyclic moieties of c[Cys3,5]-Ang II and c[Pen3,5]-Ang II preferentially assume an inverse gamma-turn conformation. On the basis of these results, we substituted amino acid residues 3-5 in Ang II with two different gamma-turn mimetics giving four diastereomeric Ang II analogues. Interestingly, two of these are equipotent to Ang II in binding to AT1 receptors. In the contractile test using rabbit aorta rings, one of the analogues is an agonist with full contractile activity approximately equipotent to c[Pen3,5]-Ang II but 300-fold less potent than Ang II. This low potency may suggest that Ang II does not adopt a gamma-turn in the 3-5 region when interacting with the receptor.

Angiotensin II↗

PubMed1997

Unexpected binding mode of a cyclic sulfamide HIV-1 protease inhibitor.

Two cyclic, C2-symmetric HIV-1 protease inhibitors, one sulfamide and one urea derivative, both comprising phenyl ether groups in the P1/P1' positions, were cocrystallized with HIV-1 protease, and the crystal structures were determined to 2.0 A resolution. The structure of the urea 2 showed a conformation similar to that reported for the related urea 3 by Lam et al., while the sulfamide 1 adopted an unanticipated conformation in which the P1' and P2' side chains were transposed.

Azepines↗

PubMed1997

Cyclic HIV-1 protease inhibitors derived from mannitol: synthesis, inhibitory potencies, and computational predictions of binding affinities.

Ten C2-symmetric cyclic urea and sulfamide derivatives have been synthesized from L-mannonic gamma-lactone and D-mannitol. The results of experimental measurement of their inhibitory potencies against HIV-1 protease were compared to calculated free energies of binding derived from molecular dynamics (MD) simulations. The compounds were selected, firstly, to enable elucidation of the role of stereochemistry for binding affinity (1a-d) and, secondly, to allow evaluation of the effects of variation in the link to the P1 and P1' phenyl groups on affinity (1a and 2-5). Thirdly, compounds with hydrogen bond-accepting or-donating groups attached to the phenyl groups in the P2 and P2' side chains (6 and 7) were selected. Binding free energies were estimated by a linear response method, whose predictive power for estimating binding affinities from MD simulations was demonstrated.

Anti-HIV Agents↗

PubMed1997

Characterization of susceptible chiasma configurations that increase the risk for maternal nondisjunction of chromosome 21.

Recent studies of trisomy 21 have shown that altered levels of recombination are associated with maternal non-disjunction occurring at both meiosis I (MI) and meiosis II (MII). To comprehend better the association of recombination with nondisjunction, an understanding of the pattern of meiotic exchange, i.e. the exchange of genetic material at the four-strand stage during prophase, is required. We examined this underlying exchange pattern to determine if specific meiotic configurations are associated with a higher risk of non-disjunction than others. We examined the crossover frequencies of chromosome 21 for three populations: (i) normal female meiotic events; (ii) meiotic events leading to MI non-disjunction; and (iii) those leading to MII non-disjunction. From these crossover frequencies, we estimated the array of meiotic tetrads that produced the observed crossovers. Using this approach, we found that nearly one-half of MI errors were estimated to be achiasmate. The majority of the remaining MI bivalents had exchanges that clustered at the telomere. In contrast, exchanges occurring among MII cases clustered at the pericentromeric region of the chromosome. Unlike the single exchange distributions, double exchanges from the non-disjoined populations seemed to approximate the distribution in the normal population. These data suggest that the location of certain exchanges makes a tetrad susceptible to non-disjunction. Specifically, this susceptibility is associated with the distance between the centromere and closest exchange. This result challenges the widely held concept that events occurring at MII are largely independent of events occurring at MI, and suggests that all non-disjunction events may be initiated during MI and simply resolved at either of the two meiotic stages.

Chromosomes, Human, Pair 21↗

PubMed1997

Human immunodeficiency virus type 1 proteinase resistance to symmetric cyclic urea inhibitor analogs.

Resistant virus was isolated from virus propagated in cell culture in the presence of the human immunodeficiency virus type 1 (HIV-1) proteinase inhibitor DMP 323, Ro 31-8959, or A-75925. The proteinase gene of resistant virus was sequenced, and key mutations (G48V, V82A, I84V, L90M, and G48V/L90M) were introduced into clones used for the expression, purification, and further characterization of the enzyme. The mutant enzymes were all less active than the wild-type enzyme, as judged by k(cat) and k(cat)/Km values. L90M had a lower Km than the wild type, whereas the G48V/L90M double mutant had an increased Km compared with that of the wild type, contributing to a 10-fold reduction in the k(cat)/Km. Vitality values were used to show that the enzyme of the I84V mutant is the enzyme most resistant to the two cyclic urea inhibitors DMP 323 and AHA 008. Virus with the same mutation is also resistant, although the double mutation L10F/I84V confers even greater resistance. All of these mutants are more resistant to DMP 323 than to AHA 008. The resistance of the I84V mutant may be attributed to a loss of van der Waals interactions with the inhibitor, since the larger amino acid side chain involved in the interaction is replaced by a smaller side chain. This is supported by the lower level of resistance to AHA 008 that was observed. The phenyl groups of AHA 008 should protrude deeper into the S1 and S1' subsites than those of the smaller compound DMP 323, reducing the loss of interaction energy. These results reveal that small structural modifications of inhibitors that do not affect the inhibitory effect on wild-type virus can influence the inhibition of resistant strains. This is of importance for optimizing drugs with respect to their potency and resistance.

Cells, Cultured↗

PubMed1997

A case of apparent trisomy 21 without the Down's syndrome phenotype.

We describe a case of apparent trisomy 21 that does not fulfill the criteria for the clinical diagnosis of Down's syndrome (DS). Our patient was subjected to karyotype analysis and found to have full, non-mosaic trisomy 21 in both blood lymphocytes and skin fibroblasts, while examination of the term placenta, which was performed earlier in the course of a different study, had shown mosaicism (73%) for trisomy 21. FISH analysis showed no obvious rearrangement of the DS chromosomal region in any of the chromosomes 21. Molecular analysis using polymorphic markers on chromosome 21 verified the existence of trisomy for the entire long arm of the chromosome and showed that the origin of the extra chromosome was maternal and was probably the result of a mitotic error. In contrast with the above, the clinical evaluation using the Jackson checklist of 25 signs failed to establish the diagnosis of DS. We believe that our patient might present mosaicism in other tissues that are not available for analysis and can be regarded as an extreme example in the continuous spectrum of karyotype phenotype associations in mosaic cases.

Down Syndrome↗

PubMed1996

Susceptible chiasmate configurations of chromosome 21 predispose to non-disjunction in both maternal meiosis I and meiosis II.

The cause of non-disjunction of chromosome 21 remains largely unknown. Advanced maternal age is associated with both maternal meiosis I (MI) and meiosis II (MII) non-disjunction events. While reduced genetic recombination has been demonstrated in maternal MI errors, the basis for MII errors remains uncertain. We studied 133 trisomy 21 cases with maternal MII errors to test the hypothesis that segregation at MII may also be influenced by genetic recombination. Our data support a highly significant association: MII non-disjunction involves increased recombination that is largely restricted to proximal 21q. Thus, while absence of a proximal recombination appears to predispose to non-disjunction in MI, the presence of a proximal exchange predisposes to non-disjunction in MII. These findings profoundly affect our understanding of trisomy 21 as they suggest that virtually all maternal non-disjunction results from events occurring in meiosis I.

Adult↗

PubMed1995

Effects of N-acetylcysteine stereoisomers on oxygen-induced lung injury in rats.

The effects of the stereoisomers of N-acetylcysteine (L-NAC and D-NAC) on oxygen-induced lung oedema have been studied in rats. The NAC-isomers were given by an osmotic minipump in order to attain continuous administration, either intravenously or intragastrically. In some experiments, plasma concentrations of NAC, cysteine and glutathione (total concentrations, i.e., concentrations obtained after reduction of the samples with dithiothreitol) were recorded. Exposure to oxygen induced an almost two-fold increase of the lung wet weight. When L-NAC or D-NAC were given intravenously, in dose of 1.1 mmol/day/kg body weight, the increase of lung wet weight was prevented by 40-50%. The plasma concentrations were approximately 40 microM (L-NAC) and approximately 90 microM (D-NAC). Following intragastrical administration of the same doses, plasma concentrations of L-NAC and D-NAC reached approximately 3 and approximately 60 microM, respectively. Using this method of administration, only D-NAC significantly diminished the increase of the lung wet weight. The difference in plasma concentrations of the NAC isomers, particularly after intragastric administration, most likely reflects the fact that L-NAC is effectively hydrolysed in most tissues, while D-NAC is resistant to enzymatic hydrolysis, thus penetrating largely intact into the systemic circulation. The data presented shows that NAC, regardless of stereoconfiguration, will protect the lung against oxygen toxicity, provided sufficient systemic levels are obtained. Since D-NAC is not a precursor of L-cysteine, formation of glutathione cannot explain the protective effects of this isomer. L- and D-NAC may therefore act via direct antioxidant/radical scavenging mechanisms and not necessarily as precursors of glutathione in this model.

Acetylcysteine↗

PubMed1995

Islet cell antibodies: variable immunostaining of pancreatic islet cells and carcinoid tissue.

OBJECTIVES: Islet cell antibodies (ICA) in sera of patients with autoimmune diabetes mellitus generally stain not only the insulin-producing beta cells but also the non-beta cells of the islets of Langerhans. The antibodies have been reported to react also with the chromaffin cells of carcinoid tissue. In the present study, we examined in detail the reactivity of 10 ICA-positive sera of patients with new onset insulin-dependent diabetes mellitus (IDDM) and two sera of patients with stiff-man syndrome. DESIGN: The sera were analysed by immunofluorescence and by immunoperoxidase staining of human islets as well as by immunoprecipitations using 35S-methionine labelled rat islet lysates. In addition, immunofluorescence analyses of carcinoid tissues were carried out. RESULTS: Eight of the 10 IDDM-positive sera reacted with all islet endocrine cells, whereas two sera showed staining restricted to the beta cells, as did the two sera of the patients with stiff-man syndrome. All beta cell 'selective' sera, but only 6 of 8 'whole' islet positive sera, immunoprecipitated the 64.kDa glutamic acid decarboxylase (GAD) antigen. The staining of carcinoid tissue was variable and did not correlate with the 'whole' or 'selective' staining pattern of islets. CONCLUSION: The data underline a heterogeneity of ICA and indicate the presence of a separate, non-GAD antigen in islet cells. It is possible that in future studies, a resolution of ICA titres with respect to different types of islet cellular reactivity might provide insights into the pathogenesis of IDDM and improve the prognostic implications of antibody determinations.

Adolescent↗

PubMed1994

Catalytic effects of glutathione peroxidase mimetics on the thiol reduction of cytochrome c.

The reduction of ferric cytochrome c by various thiols was studied. It was found that L-cysteine, L-cysteine methyl ester and D-penicillamine were very efficient reductants for cytochrome c, whereas N-acetylated amino acids (N-acetyl-L-cysteine and N-acetyl-D-cysteine) reacted considerably slower. A series of glutathione peroxidase mimetics and related compounds were studied as catalysts for the N-acetyl-L-cysteine reduction of ferric cytochrome c. Diphenyl diselenide, t-butylthio phenyl selenide, S-(phenylseleno)-glutathione, N-(phenylseleno)-phthalimide and alpha-(phenylselenenyl)-acetophenone were all efficient reduction catalysts. Diphenyl disulfide, Ebselen and several derivatives thereof were less potent catalysts whereas diaryl selenides and diphenyltelluride did not affect the rate of reduction when present in catalytic amounts. The catalysis of diphenyl diselenide, selenosulfides, alpha-(phenylselenenyl)acetophenone, N-(phenylseleno)-phthalimide and Ebselen and derived compounds was suggested to involve the formation of areneselenolate ions as redox-active species capable of transferring one electron to the ferric cytochrome c. The resulting selenium centered arylseleno radicals would then dimerize to regenerate the catalyst in the diselenide form. In the presence of diaryl ditellurides and N-acetyl-L-cysteine, ferric cytochrome c was also rapidly reduced. This reaction was stoichiometric with respect to the ditelluride reagent.

Cytochrome c Group↗

PubMed1994

Antioxidant activity of some diarylselenides in biological systems.

The selenoorganic compounds di(4-aminophenyl)selenide (10) and 4-nitro-4'-amino-diphenylselenide (36) were shown to inhibit lipid peroxidation in ADP/Fe2+/ascorbate-treated microsomes and tert-butylhydroperoxide-treated hepatocytes with IC50s of 3 and 10 microM, and 14 and 10 microM, respectively. In the former system, these inhibition constants compare favourably with those of Ebselen and classical antioxidants such as butylated hydroxytoluene (BHT) and butylated hydroxyanisole (BHA). In the cell system, these selenium compounds were equipotent with BHA but more potent than Ebselen and its analogues. The diamino compound (10) was also an effective inhibitor of lipid peroxidation initiated by diquat redox cycling in hepatocytes, again being equipotent with BHA but more potent than Ebselen and its analogues, which actually stimulated lipid peroxidation in this test system. Manipulation of the amino functions of (10) and (36) by alkylation or acylation altered the antioxidant capacity. Optimal activity in this series was achieved by N-ethylation or N-isobutylation of (10). This produced antioxidants having IC50s below 1 microM in the microsome system, 3-13 microM in the tert-butylhydroperoxide system, and being 100% effective in the diquat model at 50 microM. On the other hand, acylation or alkylation of the amino groups with long chain acyl or alkyl groups reduced the efficacy of the structures below that of the parent diamine. As with other antioxidant compounds, several of the chalcogenides were relatively selective inhibitors of monocyte 5'-lipoxygenase-dependent secretion of LTB4 as compared to their effect on cyclooxygenase-dependent secretion of PGE2 (for example compound 42 had IC50s of 0.6 microM and 10 microM, respectively). No correlation was observed between the redox-properties of the chalcogenides and their respective abilities to inhibit these enzymes.

Adenosine Diphosphate↗

PubMed1994

Diaryl tellurides as inhibitors of lipid peroxidation in biological and chemical systems.

Diaryl tellurides carrying electron-donating substituents in the para positions were found to efficiently inhibit peroxidation of rat hepatocytes, rat liver microsomes and a chlorobenzene solution of phosphatidylcholine. The most active compound in the microsomal assay, bis(4-dimethylaminophenyl) telluride, showed an IC50-value of 30 nM. This compound also caused a dose-dependent delay of the onset of the linear phase of microsomal peroxidation stimulated by iron/ADP/ascorbate. The peak oxidation potentials of the diaryl tellurides (0.50-1.14 V in MeCN) correlated linearly with the IC50-values in this assay, with a point of inflection around 0.85 V. In the hepatocyte system, all compounds showed similar protective activity. It is proposed that diaryl tellurides exert an antioxidative effect by deactivating both peroxides and peroxyl radicals under the formation of telluroxides. These oxides may regenerate the active divalent organotellurides upon exposure to a suitable reducing agent.

Animals↗

PubMed1993

Increased activity of L-type Ca2+ channels exposed to serum from patients with type I diabetes.

Type I diabetes [insulin-dependent diabetes mellitus (IDDM)] is an autoimmune disease associated with the destruction of pancreatic beta cells. Serum from patients with IDDM increased L-type calcium channel activity of insulin-producing cells and of GH3 cells derived from a pituitary tumor. The subsequent increase in the concentration of free cytoplasmic Ca2+ ([Ca2+]i) was associated with DNA fragmentation typical of programmed cell death or apoptosis. These effects of the serum were prevented by adding a blocker of voltage-activated L-type Ca2+ channels. When the serum was depleted of immunoglobulin M (IgM), it no longer affected [Ca2+]i. An IgM-mediated increase in Ca2+ influx may thus be part of the autoimmune reaction associated with IDDM and contribute to the destruction of beta cells in vivo.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗

PubMed1993

Human autoantibodies react with glutamic acid decarboxylase antigen in human and rat but not in mouse pancreatic islets.

The presence of one of the major targets for autoantibodies in Type 1 (insulin-dependent) diabetes mellitus, the enzyme glutamic acid decarboxylase, was studied in human, rat and mouse pancreatic tissue using immunoprecipitation and immunohistochemical techniques. Immunoprecipitation of glutamic acid decarboxylase was attempted with lysates of [35S]-methionine-labelled rat or mouse pancreatic islets using two different glutamic acid decarboxylase antisera, one mouse monoclonal antibody raised against the 65 kDa isoform of the enzyme, sera from six patients with Type 1 diabetes, one patient with stiff-man syndrome and sera from 19 non-obese diabetic mice. The same sera were used for immunoperoxidase staining of cryosections of human, rat or mouse pancreas. Using patient sera glutamic acid decarboxylase was detected by immunoprecipitations from isolated rat islets but not from islets of five different mouse strains tested, including the non-obese diabetic mouse. When using the non-obese diabetic mouse sera, glutamic acid decarboxylase could not be detected in either rat or mouse tissue. Immunoperoxidase staining demonstrated high levels of glutamic acid decarboxylase in human and rat pancreatic islets but low levels in mouse islets. Direct measurements of enzyme activity showed glutamic acid decarboxylase to be present in mouse islets at a level of about 40% of that in rat islets, and subsequent Western blot analyses indicated that mouse islets express the 67 kDa isoform, whereas as in rat islets both the 67 and 65 kDa isoforms are present.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1993

Demonstration of GAD-65 as the main immunogenic isoform of glutamate decarboxylase in type 1 diabetes and determination of autoantibodies using a radioligand produced by eukaryotic expression.

Plasmids containing cDNA for the rat 67- and 65-kD isoforms of glutamate decarboxylase (GAD-67 and GAD-65) were expressed in COS-cells, and lysates of [35S]methionine-labeled cells were used for immunoprecipitations. Sera from 38 patients with type 1 (insulin-dependent) diabetes mellitus, which precipitated a 64-kD antigen from rat islets, reacted with recombinant GAD-65 in relation to their anti-64-kD titers. The eight strongest sera also precipitated recombinant GAD-67, suggesting that certain epitopes are common to both isoforms. Subsequently, [35S]methionine-labeled GAD-65 was purified from COS cell lysates and employed in a binding assay with 50 sera of patients with recent onset of type 1 diabetes mellitus. 38 sera (76%) precipitated labeled GAD-65 with titers that correlated with islet cell antibodies (ICA), determined in a standard immunofluorescence assay. 2 sera were GAD positive but ICA negative, 4 were positive only for ICA, and 6 were negative for both GAD and ICA, as were the sera of 20 controls. The data illustrate that antibodies against GAD-65 are present in a majority of patients with type 1 diabetes mellitus and that autoantibodies against other islet cell antigens also exist. The radioligand-binding assay, which is convenient and sensitive for detecting GAD antibodies, will facilitate the screening of individuals with autoimmune islet cell disease.

Adolescent↗

PubMed1993

Enhanced stimulus-secretion coupling in polyamine-depleted rat insulinoma cells. An effect involving increased cytoplasmic Ca2+, inositol phosphate generation, and phorbol ester sensitivity.

To extend previous observations on the role of polyamines in insulin production, metabolism, and replication of insulin-secreting pancreatic beta cells, we have studied the role of polyamines in the regulation of the stimulus-secretion coupling of clonal rat insulinoma cells (RINm5F). For this purpose, RINm5F cells were partially depleted in their polyamine contents by use of the specific ornithine decarboxylase inhibitor difluoromethylornithine (DFMO), which led to an increase in cellular insulin and ATP contents. Analysis of different parts of the signal transduction pathway revealed that insulin secretion and the increase in cytoplasmic free Ca2+ concentration ([Ca2+]i) after K(+)-induced depolarization were markedly enhanced in DFMO-treated cells. These effects were paralleled by increased voltage-activated Ca2+ currents, as judged by whole-cell patch-clamp analysis, probably reflecting increased channel activity rather than elevated number of channels per cell. DFMO treatment also rendered phospholipase C in these cells more sensitive to the muscarinic receptor agonist carbamylcholine, as evidenced by enhanced generation of inositol phosphates, increase in [Ca2+]i and insulin secretion, despite an unaltered ligand binding to muscarinic receptors and lack of effect on protein kinase C activity. In addition, the tumor promoter 12-O-tetradecanoylphorbol 13-acetate, at concentrations suggested to be specific for protein kinase C activation, evoked an increased insulin output in polyamine-deprived cells compared to control cells. The stimulatory effects of glucose or the cyclic AMP raising agent theophylline on insulin release were not increased by DFMO treatment. In spite of increased binding of sulfonylurea in DFMO-treated cells, there was no secretory response or altered increase in [Ca2+]i in response to the drug in these cells. It is concluded that partial polyamine depletion sensitizes the stimulus-secretion coupling at multiple levels in the insulinoma cells, including increased voltage-dependent Ca2+ influx and enhanced responsiveness to activators of phospholipase C and protein kinase C. In their entirety, our present results indicate that the behavior of the stimulus-secretion coupling of polyamine-depleted RINm5F insulinoma cells changes towards that of native beta cells, thus improving the usefulness of this cell line for studies of beta cell insulin secretion.

Adenosine Triphosphate↗

PubMed1993

Modulation of beta-cell activity and its influence on islet cell antibody (ICA) and islet cell surface antibody (ICSA) reactivity.

Insulin-dependent diabetes mellitus (IDDM) is associated with the formation of autoantibodies against different antigens in the islets of Langerhans, so-called islet cell antibodies (ICA). The expression of a major autoantigen, the beta-cell specific enzyme glutamic acid decarboxylase (GAD), is glucose-dependent in vitro and correlated to insulin release in vitro. In this study the expression of islet autoantigens was examined in vivo and the relationship between beta-cell function and islet cell surface antibody (ICSA) reactivity was tested. Rats were fed for 10 days with glipizide or diazoxide, in order to stimulate or inhibit insulin release, respectively. Frozen sections of pancreata were incubated with ten ICA-positive IDDM sera and analyzed by indirect immunofluorescence. Two sera with a "beta-cell restricted" staining, five with an "all-islet cell" staining and three with a "mixed" pattern were employed. In all three groups, the highest end-point titres were obtained when pancreata of rats treated with glipizide were used. Intermediate titres were seen in control animals and the lowest titres were observed on pancreata from diazoxide-treated rats, regardless of the serum used. In contrast to these observations, no correlation between ICSA reactivity and islet cell activity could be demonstrated. Conflicting results concerning ICSA in previous reports and our failure to show a glucose regulation of ICSA reactivity, indicate that ICSA is a phenomenon with a low degree of specificity.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1992

Alpha-(phenylselenenyl)acetophenone derivatives with glutathione peroxidase-like activity. A comparison with ebselen.

Here we describe a new class of organoselenium compounds possessing glutathione peroxidase-like activity. The parent compound, alpha-(phenylselenenyl)acetophenone (PSAP), increased the rate of reaction of glutathione with H2O2, tert-butylhydroperoxide, cumene hydroperoxide, linoleic acid hydroperoxide and dilinoleyl lecithin hydroperoxide by 7.0, 25.1, 34.1, 19.1 and 8.4-fold, respectively, as assessed by the oxidized glutathione (GSSG) reductase enzyme assay. Direct assay of the removal of hydrogen peroxide and glutathione from reaction mixtures confirmed the peroxidase-like activities of these selenoorganic compounds, but indicate that the conventional coupled GSSG reductase assay may be unsuitable for the assessment of the catalytic capacity of PSAP and Ebselen. One possible mechanism of catalysis by PSAP involves an initial oxidation at selenium. Thiol may then react with the selenoxide to yield a selenium (II) compound, H2O and a disulfide. Compounds derived from PSAP may provide potential selenium-based anti-inflammatory agents.

Acetophenones↗