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Biomedical subjects

G Dawson

Publications and source records attributed to G Dawson.

At least 235 records · Page 13Linked to original sources
PubMed1981

Glycosphingolipid-high density lipoprotein-3 interactions. I. Transfer of glycosphingolipid from phosphatidylcholine vesicles to high density lipoprotein-3.

Single bilayer vesicles (d less than 1.02 g/ml) of 3H-glycosphingolipids and [14C]phosphatidylcholine in the molar ratio of 1:7 were prepared by ethanolic injection of the lipid mixture into buffer, concentrated, and incubated with human serum high density lipoprotein-3 (HDL3; d = .14 g/ml) at 37 degrees C. Equilibrium ultracentrifugation of the incubation mixtures on a 0-22% NaBr gradient revealed the presence of three discrete lipid-protein complexes of density 1.03, 1.06, and 1.12 g/ml (Peaks I, II, and III, respectively). Each peak was homogeneous upon reultracentrifugation and the protein and radioactivity eluted as a single peak upon Sepharose CL-6B chromatography. Compositional analysis showed peak I to contain 2.6% protein (apo-A-I peptide) and 4.3% cholesterol, peak II to contain 17.6% protein (apo-A-I peptide) and 6.3% cholesterol, and peak III to have a composition similar to HDL3. Electron microscopy of negatively stained samples confirmed the homogeneity of the peaks and the similarity between peak III and HDL3. Peak II particles were larger than HDL3; peak I particles resembled fused or aggregated vesicles which could be removed by ultracentrifugation; disc-shaped particles were not seen in any of the fractions. Direct incubation of HDL3 or human serum with 3H-glycosphingolipid dispersions did not yield a glycolipid . HDL3 complex as judged by density gradient ultracentrifugation and Sepharose CL-6B chromatography. However, incubation of 3H-glycolipid/phosphatidylcholine vesicles with serum did result in transfer of 3H-glycolipid to the HDL fraction. It was concluded that glycolipids incorporated into a lipid membrane structure can interact with, and become incorporated into, high density lipoprotein.

Gangliosidoses↗
PubMed1981

Caprine beta-mannosidosis. Inherited deficiency of beta-D-mannosidase.

A rapidly fatal neurovisceral storage disease was discovered in both male and female offspring of clinically normal Nubian goats. Microscopic examination of fixed tissues revealed extensive demyelination and ubiquitous distribution of lysosomal storage vacuoles containing dispersed floccular material and membranous fragments. Urine was found to contain elevated levels of both mannose and N-acetylglucosamine, suggestive of an oligosaccharide storage disease. Brain was found to contain 2.2 mumol/g of a trisaccharide Man(beta 1-4)GlcNAc(beta 1-4)GlcNAc; (Jones, M. Z., and Laine, R. A. (1980) Fed. Proc. 39, 2521 and Jones, M. Z., and Laine, R. A. (1981) J. Biol. Chem. 256, 5181-5184). A profound deficiency of beta-D-mannosidase activity was found in a number of tissues from affected goats; obligate heterozygotes showed a partial enzyme deficiency. Other lysosomal hydrolase activities were normal or elevated over normal, including alpha-D-mannosidase, confirming that this was a hitherto undescribed inborn error of glycoprotein catabolism.

Animals↗
PubMed1981

Stimulation of glycolipid synthesis and exchange by human serum high density lipoprotein-3 in human fibroblasts and leukocytes.

Upon exposure to either human skin fibroblasts or human circulating leukocytes, the composition of human serum high density lipoprotein-3 (HDL3) was modified by the apparent loss of apolipoprotein A-II and a 2- to 4-fold increase in glycosphingolipid content. Exposure of HDL3 to leukocytes produced an increase in the content of lactosylceramide, which is the major glycolipid in leukocytes, whereas exposure of HDL3 to human skin fibroblasts produced predominantly an increase in trihexosylceramide, which is the major glycolipid in fibroblasts. Other protein components of HDL3 (such as apolipoprotein A-I) were unaffected and there were no major changes in either neutral lipid or phospholipid composition. The increase in glycosphingolipid content of both cells and reisolated HDL3 particles was HDL3 concentration-dependent up to a concentration of 1 mg/ml and appeared to be the result of a stimulation of cellular glycolipid synthesis by HDL3 and subsequent transfer to HDL3 in the medium. A similar stimulation could not be produced by either low density lipoprotein or lipoprotein-deficient human serum. The coaddition of HDL3 and lipoprotein-deficient serum reduced both the loss of apolipoprotein A-II and the change in HDL3 glycolipid content, but not the increase in cellular glycolipid content, suggesting that modification of the apolipoprotein A-II peptide may enhance the ability of HDL3 to acquire new glycolipid from cells.

Cells, Cultured↗
PubMed1981

Synthesis of gangliosides by cultured oligodendrocytes.

Gangliosides are enriched in the nervous system compared to other tissues. The synthesis of gangliosides by monolayer cultures of isolated oligodendrocytes has not previously been investigated. Cells were labeled with [3H] galactose at preselected times and gangliosides isolated by phase partition, purified, and identified by chromatography. Cultured oligodendrocytes showed selectivity in their synthesis of gangliosides, which was expressed in the type of ganglioside synthesized as well as in the change of incorporation over time in culture. For the first ten days, there was very little incorporation of [3H] galactose in gangliosides, but this was followed by a stimulation of uptake for GM3, GM1/GD3, and GD1 gangliosides, reaching a maximum after approximately 25-30 days in vitro. There was little incorporation into GM2 or trisialogangliosides throughout the life of the cultures. Since oligodendrocytes synthesize extensive membranes during this period, one may speculate that the de novo-synthesized gangliosides are used for membranes.

Animals↗
PubMed1981

Distinct high-affinity binding sites for benzomorphan drugs and enkephalin in a neuroblastoma--brain hybrid cell line.

The high-affinity binding of benzomorphan drugs (ethylketocyclazocine and N-allylnorcyclazocine) and [DAla2,DLeu5] enkephalin was examined in a mouse neuroblastoma--Chinese hamster brain clonal hybrid cell line (NCB-20). Scatchard analysis of saturation binding isotherms indicated the presence of a single binding site for 3H-labeled [DAla2,DLeu5]enkephalin (Kd = 3 nM) and multiple binding sites for [3H]ethylketocyclazocine (Kd = 4 and 20 nM) and N-[3H]allylnorcyclazocine (Kd = 0.5 and 15 nM). Both ethylketocyclazocine and N-allylnorcyclazocine competed (Ki = 10 and 30 nM, respectively) with [3H][DAla2,DLeu5]enkephalin binding in NCB-20 cells but neither [DAla2,DLeu5]enkephalin nor morphine could completely inhibit the specific binding of [3H]ethylketocyclazocine (7 nM) or N-[3H]allylnorcyclazocine (3 nM). Furthermore, not all benzomorphan drugs (e.g., ethylketocyclazocine) were totally efficacious in displacing 3 nM N-[3H]allylnorcyclazocine binding in the presence or absence of high concentrations of [DAla2,DLeu5]enkephalin. The data presented suggest that benzomorphan drugs interact with three distinct high-affinity binding sites: (i) a site that binds enkephalin and morphine in addition to ethylketocyclazocine and N-allylnorcyclazocine; (ii) a site that binds both ethylketocyclazocine and N-allylnorcyclazocine but not enkephalin and morphine; and (iii) a site that binds N-allylnorcyclazocine but not enkephalin, morphine, or ethylketocyclazocine. The first of these sites was comparable to the delta opiate receptor expressed in NG108-15 and N4TG1 cell lines based on the potency series obtained for various opiates and benzomorphan drugs in competition studies with [3H][DAla2,DLeu5]-enkephalin. However, the specific high-affinity benzomorphan binding sites thus far are unique and may represent biochemical correlates of kappa and sigma opiate receptors which have been proposed to exist on the basis of physiological studies.

Analgesics, Opioid↗
PubMed1981

Possible role of cyclic AMP in the receptor-mediated regulation of glycosyltransferase activities in neurotumor cell lines.

Exposure of mouse neuroblastoma cell line N4TGl to opiates or [D-Ala2,D-Leu5] enkephalin produced a naloxone-reversible inhibition of cyclic AMP synthesis and prevented, in a concentration-dependent manner, the formation of both ganglioside GM2 (GalNAc-[NeuNAc]-Gal-Glc-ceramide) from GM3 (NeuNAc-Gal-Glc-ceramide) and ganglioside GM1 (Gal-GalNAc-[NeuNAc]-Gal-Glc-ceramide) from GM2 in cell-free extracts. In contrast, the receptor-mediated elevation of intracellular cyclic AMP levels by agents such as prostaglandin E1 (in the presence of isobutylmethylxanthine) or the addition of the cyclic AMP derivatives (dibutyryl cyclic AMP) markedly stimulated the activities of UDP-GalNAc:GM3,N-acetylgalactosaminyltransferase and UDP-Gal:GM2,galactosyltransferase. An overall increase in the synthesis of gangliosides more complex than GM3 was also observed in the mouse neuroblastoma x hamster brain explant hybrid cell line NCB-20 following elevation of cyclic AMP levels by treatment with serotonin and pargyline. The data presented support the hypothesis that cyclic AMP may have a role in the regulation of sialoglycosphingolipid biosynthesis.

Animals↗
PubMed1980

Maintenance of isolated oligodendrocytes in long-term culture.

A new procedure for isolating oligodendrocytes from ovine white matter is described. The method separates oligodendrocytes into two bands on a linear sucrose gradient. Five criteria have been employed to classify the separated cells. It is shown by indirect immunofluorescence with specific antisera that 97% of the cells from both bands carry galactocerebroside, a specific surface marker for oligodendrocytes, on their plasma membranes and 95% of the cells retain myelin basic protein as distinct patches on their surfaces. Isolated cells conform ultrastructurally to current concepts of oligodendrocytes. The cells incorporate [3H]galactose into galactocerebroside and carrier free H2(35)SO4 into sulfatide, specific markers for oligodendrocytes. The specific activity of 2',3'-cyclic nucleotide-3'-phosphodiesterase in the two cell fractions is comparable to that reported for isolated oligodendrocytes by others. It is concluded that conservatively, 95% of the cells in both fractions are oligodendrocytes. Cells from both bands survive in culture for months. In vitro the cells extend two or more processes, contain 'gliosomes', and surround themselves with extensive sheet-like membranes; i.e. they exhibit the morphological characteristics ascribed to oligodendrocytes in explant cultures. Conservatively 90% of cultured cells stain with an antimyelin basic protein serum. The staining is localized in the cytoplasm and processes. The cells also stain with antigalactocerebroside and antioligodendrocyte sera. Cells remain differentiated for up to 70 days in vitro as evidenced by their incorporation of [3H]galactose and H2(35)SO4 into galactosyl and sulfogalactosylceramide, respectively.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed1980

A lysosomal storage disorder in mice characterized by a dual deficiency of sphingomyelinase and glucocerebrosidase.

Lipid and lysosomal enzyme levels in the tissues of a strain of mice afflicted with an autosomal rescessive neuroviscereal storage disorder were examined. Sphingomyelinase and glucocerebrosidase activities were consistently diminished in a wide variety of tissues obtained from the affected mice. The activities of these enzymes were clearly attenuated in new-born mice, which at this age, were otherwise indistinguishable from littermates and age-matched controls. The deficiency of sphingomyelinase was more pronounced than glucocerebrosidase. There was progressive accumulation of sphingomyelin, glucocerebroside, lactosylceramide and unesterified cholesterol in the tissues of these mice in the postnatal period. Gangliosides GM2 and GM3 accumulated in the brain of the animals, and GM3 and asialo-GM2 were stored in the liver. Furthermore, there was a large increase in the quantity of hepatic bis(monoacylglycero)phosphate. The accumulation of lipids was parallelled by a progressive elevation in the activity of several lysosomal hydrolases in various tissues. Heterozygous mice were biochemically indistinguishable from normal controls. The phenotypic manifestations in these metabolically mutated animals are compared with those in Niemann-Pick disease and Gaucher's disease in humans.

Animals↗
PubMed1980

Purification and properties of two forms of human alpha-L-fucosidase.

High (100 000) and low (50 000) molecular weight forms of alpha-L-fucosidase (alpha-fucosidase I and II) were purified to apparent homogeneity from human spleen, liver brain and kidney on the basis of differential affinity for epsilon-amino-caproyl fucosamine-agarose bead columns. Alpha-fucosidase I (the "bound" form) consisted of two 50 000 dalton monomers; however, both forms can aggregate to tetramer and hexamer forms. Most previous studies on alpha-fucosidase have been carried out on this form of human alpha-fucosidase although the bound and unbound forms of the enzyme were present in equal amounts in human spleen. The bound (100 000) form is a sialoglycoprotein whereas the unbound (50 000) form, is a neutral mannose-rich glycoprotein. Other differences with respect to amino acid composition, pH optimum, electrophoretic mobility, KM, thermal stability, and natural substrate specificities were observed. All preparations hydrolysed 4-methylumbelliferyl-alpha-L-fucoside, flucosyllactose, and lacto-N-fucopentaose I, but the unbound fraction (alpha-fucosidase II) preferentially hydrolyused lacto-N-fucopentaose II. The unbound (mannose-rich) alpha-fucosidase II was taken up by human skin fibroblasts with higher affinity (5% per 2 h per 2 times 10(5) cells) than the bound (sialo-) alpha-fucosidase I (<1% per 2 times 10(5) cells). Uptake was inhibited by other lysosomal hydrolases, fetal calf serum, mannose 6-phosphate and phosphomannans and to a lesser extent by heparins. Our studies suggest that alpha-fucosidase I is not a simple dimer of alpha-fucosidase II and represents a less-biologically active form of the enzyme.

Carbohydrate Metabolism, Inborn Errors↗
PubMed1980

Comparison between the cellular proteins of hairy cell leukemia and the leukemic phase of other lymphoproliferative diseases.

Cellular proteins from malignant cells of the leukemic phase of hairy cell leukemia and other lymphoproliferative diseases characterized by immunological markers were evaluated by sodium dodecyl sulfate gradient-polyacrylamide gel electrophoresis. The protein patterns from eight patients with hairy cell leukemia were essentially identical. The protein patterns from eight patients with chronic lymphocytic leukemia, seven patients with acute lymphocytic leukemia, and four patients with poorly differentiated lymphocytic lymphoma were examined and did not demonstrate a consistent pattern within each disease. The protein patterns of one patient each with T-cell malignant lymphoma, lymphoblastic lymphoma, or acute monocytic leukemia were also examined. The protein pattern for hairy cell leukemia is distinctly different from that of all the other diseases studied; differences were distinct even within and between immunological subtypes.

Electrophoresis, Polyacrylamide Gel↗
PubMed1979

Purification and characterization of a glycopeptide derived from Phaseolus vulgaris leukoagglutinating phytohemagglutinin.

A glycopeptide obtained from tryptic digestion of citraconylated leukoagglutinating (L-subunit) phytohemagglutinin was isolated and purified. The composition of its ten amino acid and seven hexose residues, when compared to a previous analysis of the NH2-terminal amino acid sequence, indicated that the glycopeptide was derived from residues 11 through 20, with the oligosaccharide unit linked N-glycosidically to asparagine at the twelfth position. Structural studies involving a combination of alpha-mannosidase digestion, periodate oxidation, and permethylation, in conjunction with computerized mass-spectrometric and masschromatographic techniques, suggested a branched sequence of five mannose and two glucosamine residues similar to that found in many mammalian glycoproteins.

Chemical Phenomena↗
PubMed1979

Fatty acid differences amongst leukemia cell types, with special reference to hairy cell leukemia: a preliminary report.

Fatty acid analyses of malignant cells from hairy cell leukemia patients revealed a profile which was rich in arachidonate and relatively low in linoleate and oleate. This profile was distinct from that of mononuclear cells derived from patients with other lymphoproliferative diseases or myeloproliferative disorders, and may reflect the unique membrane morphology of the hairy cell.

Arachidonic Acids↗
PubMed1979

Human acoustic neurinomas: nervous system specific biochemical parameters.

A series of 24 human acoustic neurinomas from 24 patients has been assayed for several biochemical parameters characteristic of the nervous system. S 100 protein, 2', 3'-cyclic nucleotide 3'-phosphohydrolase activity, and the myelin lipids galactosylceramide and sulfogalactosylceramide (sulfatide). Myelin basic protein was not detected. These finding further support the neuroectodermal origin of the human acoustic neurinoma, and provide additional biochemical markers for further study.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗