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- Bäcker, Annika E., 1965, et al.
(författare)
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Biochemical and enzymatic characterization of blood group ABH and related histo-blood group glycosphingolipids in the epithelial cells of porcine small intestine.
- 1997
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Ingår i: Glycobiology. - : Oxford University Press (OUP). - 0959-6658 .- 1460-2423. ; 7:7, s. 943-53
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Tidskriftsartikel (refereegranskat)abstract
- Non-acid glycosphingolipids were isolated from small intestinal epithelial cells of a single blood group A pig. One very predominant blood group compound was obtained chemically pure upon HPLC fractionation. It was characterized by mass spectrometry and 1H NMR spectroscopy to be the type 1 chain blood group A hexaglycosylceramide. Support for the presence of minute amounts of additional A glycolipids was obtained by mass spectrometry and immunostaining of TLC plates with anti-A antibodies specific for A type 2 chain, A type 3 and 4 chain, and the ALe(b) determinant. Among precursor chains, globoside (type 4) and lactotetraosylceramide (type 1) were immunologically identified, whereas no neolactotetraosylceramide (type 2) and gangliotetraosylceramide reactivities were detected. We addressed the question whether the predominant expression of type 1 chain based A glycolipids reflects a restricted glycolipid precursor chain specificity of the alpha 1-2 fucosyl- and/or the alpha 1-3 N-acetylgalactosaminyltransferases, or if the biosynthesis of the precursor chains themselves is regulated. All precursor core saccharides, lacto- (type 1), neolacto-(type 2), and gangliotetraosylceramide as well as globopentaosylceramide (type 4), could serve as acceptors for fucose in vitro when a crude microsomal fraction obtained from mechanically released, porcine intestinal epithelial cells was used as an enzyme source. Under the same conditions an N-acetylgalactosamine residue could be transferred to the blood group H structures based on these core saccharide chains. Lactotriaosylceramide, but not gangliotriaosylceramide, could serve as an acceptor for UDP-galactose. When the product was digested with beta-galactosidase (EC 3.2.1.23) from S.pneumoniae, under conditions where it specifically cleaves Gal beta 1-4 residues, approximately 40% of the radioactivity was cleaved off, indicating that a substantial amount of neolactotetraosylceramide was made in vitro, as opposed to the predominance of lactotetraosylceramide-based structures found in vivo.
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- Kulseth, Mari Ann, et al.
(författare)
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Serglycin expression during differentiation of U937-1 cells
- 1998
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Ingår i: Glycobiology. - 0959-6658 .- 1460-2423. ; 8:8, s. 747-753
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Tidskriftsartikel (refereegranskat)abstract
- Serglycin is the major proteoglycan in most hematopoietic cells, including monocytes and macrophages. The monoblastic cell line U937-1 was used to study the expression of serglycin during proliferation and differentiation. In unstimulated proliferating U937-1 cells serglycin mRNA is nonconstitutively expressed. The level of serglycin mRNA was found to correlate with the synthesis of chondroitin sulfate proteoglycan (CSPG). The U937-1 cells were induced to differentiate into different types of macrophage-like cells by exposing the cells to PMA, RA, or VitD3. These inducers of differentiation affected the expression of serglycin mRNA in three different ways. The initial upregulation seen in the normally proliferating cells was not observed in PMA treated cells. In contrast, RA increased the initial upregulation, giving a reproducible six times increase in serglycin mRNA level from 4 to 24 h of incubation, compared to a four times increase in the control cells. VitD3 had no effect on the expression of serglycin mRNA. The incorporation of (35S)sulfate into CSPG decreased approximately 50% in all three differentiated cell types. Further, the (35S)CSPGs expressed were of larger size in PMA treated cells than controls, but smaller after RA treatment. This was due to the expression of CSPGs, with CS-chains of 25 and 5 kDa in PMA and RA treated cells, respectively, compared to 11 kDa in the controls. VitD3 had no significant effect on the size of CSPG produced. PMA treated cells secreted 75% of the (35S)PGs expressed, but the major portion was retained in cells treated with VitD3 or RA. The differences seen in serglycin mRNA levels, the macromolecular properties of serglycin and in the PG secretion patterns, suggest that serglycin may have different functions in different types of macrophages.
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- Lind, Thomas, et al.
(författare)
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cDNA cloning and expression of UDP-glucose dehydrogenase from bovine kidney
- 1999
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Ingår i: Glycobiology. - 0959-6658 .- 1460-2423. ; 9:6, s. 595-600
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Tidskriftsartikel (refereegranskat)abstract
- We have isolated a cDNA encoding UDP-glucose dehydrogenase from a bovine kidney cDNA-library, the first mammalian cDNA clone published. [After submission of the manuscript, a study appeared describing the molecular cloning and characterization of the human and mouse UDP-glucose dehydrogenase genes(Spicer et al., 1998).] The enzyme catalyzes the conversion of UDP-glucose to UDP-glucuronicacid, an essential precursor in glycosaminoglycan biosynthesis. The cDNA has an open reading frame of 1482 nucleotides coding for a 55 kDa protein. Expression of the enzyme in COS-7 cells showed a 3-fold increase inUDP-glucose dehydrogenase activity; also, the C-terminal 23 amino acidswas shown not to be necessary for enzyme activity. Northernblots from human and mouse tissues reveal high expression in liver and low in skeletal muscle. Human tissues have a majortranscript size of 3.2 kilobases and a minor of 2.6 whereas mousetissues have a single 2.6 kilobase transcript. We have also developed a sensitive and direct assay using UDP-[14C]Glc as a substrate for detection of small amounts of UDPGDH activity.
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| 9. |
- Malmström, J, et al.
(författare)
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Heparan sulfate upregulates platelet-derived growth factor receptors on human lung fibroblasts
- 1998
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Ingår i: Glycobiology. - : Oxford University Press (OUP). - 0959-6658 .- 1460-2423. ; 8:12, s. 55-1149
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Tidskriftsartikel (refereegranskat)abstract
- Heparan sulfate is a molecule that possesses a large structural variability and which has been shown to inhibit the proliferation of fibroblasts in vitro. The aim of this study was to determine whether the anti-proliferative effects of heparan sulfate were exerted by regulation of the activity of the platelet-derived growth factor and/or of the platelet-derived growth factor receptors. Both l-iduronate-rich, anti-proliferative and the l-iduronate-poor, non-anti-proliferative heparan sulfate species, were incubated with confluent human embryonic lung fibroblasts for 24 h. The mRNA levels for PDGF-AA, PDGF-BB, and their receptors were measured. Binding studies were performed with [125I]-PDGF-BB and [125I]-EGF for 2 h at 4 degreesC in cultures preincubated with both types of heparan sulfate for 24 h. In separate experiments, cultures were incubated together with heparan sulfate and [125I]-PDGF-BB for 2 h at 4 degreesC. Increases of two- to threefold in the mRNA levels for both the alpha- and the beta-receptors of PDGF was obtained after treatment with both types of heparan sulfate, whereas the mRNA levels of both the PDGF-AA and the PDGF-BB were essentially unaffected. A sixfold increase in binding was only noted for [125I]-PDGF-BB in cultures pre-treated with the anti-proliferative heparan sulfate for 24 h, whereas no effect was noted with use of the non-anti-proliferative heparan sulfate. Incubating the [125I]-PDGF-BB and the anti-proliferative heparan sulfate together for 2 h resulted in a smaller, threefold increase in binding. This indicates that the anti-proliferative heparan sulfate both stabilizes and increases expression of the PDGF receptors. To investigate whether the increased number of PDGF receptors could affect cell activity, cells were preincubated with anti-proliferative heparan sulfate and then treated with PDGF-BB. This resulted in an increase in mitogenicity compared to cells treated only with PDGF-BB. Neither an increase in binding for [125I-EGF] nor an increase in the mitogenic response of EGF could be observed in cultures pre-treated with the anti-proliferative heparan sulfate. The results indicate that the extracellular matrix itself may regulate important biological phenomena such as cell proliferation and matrix production through affecting the expression of receptors of PDGF, which initiate both stimulatory and inhibitory signals.
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