Isolation and identification of glycosaminoglycans associated with purified nuclei from rat liver

Furukawa, Kiyoshi; Terayama, Hiroshi

doi:10.1016/0304-4165(77)90010-1

Cited by 57 publications

(24 citation statements)

References 30 publications

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“…Indeed, intracellular hyaluronan receptors have been isolated from hepatic ceh (18), suggesting a role for this glycosaminoglycan in maintaining cell shape and integrity under the influence of external pressure (18). Staining of nuclear heterochromatin in both vascular and other cell types using our hyaluronan binding probes is also in agreement with published histochemical (8,43) and biochemical (21,37) data, but the functional implications of these findings are not known.…”

Section: Discussionsupporting

confidence: 80%

Association of hyaluronan with rat vascular endothelial and smooth muscle cells.

Eggli

Graber²

1995

J Histochem Cytochem.

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We investigated the distribution of hyaluronan (hyaluronic acid) in rat vascular tissue fixed by an osmium tetroxide (or glutaraldehyde) microwave technique and embedded in epoxy resin (or Lowiayl K4M), using hyaluronan binding proteins coupled to 15-20-nm gold particles as ultrastructural markets in a one-step post-embedding procedure. The intra-and extracellular aspects of vascular endothelial and smooth muscle cell plasma membranes revealed distinct IntroductionOn the basis principally of biochemical evidence, both normal and especially pathologically altered arterial walls appear to be relatively rich in hyaluronan (25,36,53). A number of attempts have also been made to identlfy this non-sulfated glycosaminoglycan histochemically within vascular tissues. In early studies, the nonspecific glycosaminoglycan-precipitating cationic dye Ruthenium Red was used and electron microscopic examination of arterial walls fixed in its presence revealed a close association between testicular hyaluronidase-digestible Ruthenium Red-positive granules and arterial smooth muscle cell membranes (62). The introduction of biotinylated hyaluronan binding proteins as a more specific means for detecting hyaluronan (44) stimulated the undertaking of several light microscopic investigations for its localization in vessels of both normal and pathologically altered tissues (10,12,32,59,60,63). However, histochemical analyses using these probes have consistently failed to reveal a convincing relationship between hyaluronan and vessel walls, at least in normal tissues (12,32,59,60,63 hyaluronan in the vascular walls of several rat tissues was achieved using a modified histochemical probe (16). Materials and MethodsSpecimen Handling. Adult (3-6 months) pigmented (wild-type) and non-pigmented (Wistar) rats were sacrificed by an overdose of pentobarbital. Pieces of abdominal wall skin 2 x 2 mm and 1 mm thick were prepared and immersed in fixative. Eyes were enucleated and the halfposterior to the ciliary body cut away. The lens was removed and the entire anterior portion fied; it was further dissected into 8-12 radial segments before dehydration and embedding.Microwave Oven Set-up. For all experiments, a conventional microwave oven with a maximal power output of 650 W was used. A water load (200 ml in a beaker) placed in a rear corner of the oven (34) served as a damper to increase irradiation times to 10 sec or more. Glass vials (24 x 40 mm) containing the specimens and 10-ml fixative were placed in the center of the microwave Oven and heated at maximal power output and a frequency of 2450 MHz.Osmium TeaOXidelMimve Fixation and Embedding in Epoxy Resin.Rat abdominal wall skin blocks and anterior eye portions were fixed for 2 min at ambient temperature in 2% (vlv) Os04 solution (in 0.2 M sodium cacodylate buffer; pH 7.2, 450 mOsm). They were then transferred to a microwave oven and heated until temperatures of 43-46'C were attained (-12 sec) (16). Samples were maintained in the fixative solution for 10 min at ambient temperature before washin...

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Section: Discussionsupporting

confidence: 80%

Association of hyaluronan with rat vascular endothelial and smooth muscle cells.

Eggli

Graber²

1995

J Histochem Cytochem.

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“…40,41 Hyaluronan is taken up by cells, 42,43 and cancer cells take up more hyaluronan than do normal cells. 42 Hyaluronan is transported to the nucleus and nucleolus, [43][44][45] where it stimulates cell cycle progression. Some nuclear proteins involved in cell cycle progression 46 and RNA splicing 47 are hyaluronan-binding proteins.…”

Section: Discussionmentioning

confidence: 99%

Hyaluronidase reduces human breast cancer xenografts in SCID mice

Shuster

Frost

Csóka

et al. 2002

Intl Journal of Cancer

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A hyaluronan-rich environment often correlate with tumor progression. and may be one mechanism for the invasive behavior of malignancies. Eradication of hyaluronan by hyaluronidase administration could reduce tumor aggressiveness and would provide, therefore, a new anti-cancer strategy. Hyaluronan interaction with its CD44 receptor and the resulting signal transduction events may be among the mechanisms for hyaluronan-associated cancer progression. We have shown previously that hyaluronidase treatment of breast cancer cells in vitro not only eradicates hyaluronan but also modifies expression of CD44 variant exons of tumor cells. We now determine if such effects occur in vivo and if it is accompanied by tumor regression. SCID mice bearing xenografts of human breast carcinomas were given intravenous hyaluronidase. Tumor volumes decreased 50% in 4 days. Tumor sections showed decreased hyaluronan. Intensity of staining for CD44s was not affected, whereas staining for specific CD44 variant exon isoforms was greatly reduced in residual tumors. Necrosis was not evident. Hyaluronidase, used previously as an adjunct in cancer treatment, presumably to enhance penetration of chemotherapeutic drugs, may itself have intrinsic anti-cancer activity. Removing peritumor hyaluronan appears to cause an irreversible change in tumor metabolism. Continuous hyaluronan binding to CD44 variant exon isoforms may also be required to stabilize inherently unstable isoforms that participate perhaps in tumor progression. Further investigation is required to confirm a cause and effect relationship between loss of hyaluronan, changes in CD44 variant exon expression and tumor reduction. If confirmed, hyaluronidase may provide a new class of anti-cancer therapeutics and one without toxic side effects.

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“…After chondroitinase digestion, the reactions were terminated by heating in a boiling water bath for 5 min and the digested samples were desalted using a Sephadex G-10 microanalysis desalting spin column (Harvard Apparatus Inc. MA, USA) and freeze-dried. Next the freeze-dried sample (0.3-5 mg) was dissolved in 20 mM Tris-HCl buffer (pH 8) containing 2 mM magnesium chloride and digested with endonuclease (2500 m units/mg) for 12 h at 37°C [21] to remove any nucleic acid contaminants. After endonuclease digestion, the reactions were terminated by heating in boiling water bath for 5 min and the samples were desalted using a Sephadex G-10 microanalysis desalting spin columns and the resulting purified HS samples were freeze-dried.…”

Section: Tissues Processing Isolation and Purification Of Heparan Sumentioning

confidence: 99%

Isolation and characterization of heparan sulfate from various murine tissues

et al. 2006

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Heparan sulfate (HS), is a proteoglycan (PG) found both in the extracellular matrix and on cell surface. It may represent one of the most biologically important glycoconjugates, playing an essential role in a variety of different events at molecular level. The publication of the mouse genome, and the intensive investigations aimed at understanding the proteome it encodes, has motivated us to initiate studies in mouse glycomics focused on HS. The current study is aimed at determining the quantitative and qualitative organ distribution of HS in mice. HS from brain, eyes, heart, lung, liver, kidney, spleen, intestine and skin was purified from 6-8 week old male and female mice. The recovered yield of HS from these organs is compared with the recovered whole body yield of HS. Structural characterization of the resulting HS relied on disaccharide analysis and 1 H-NMR spectroscopy. Different organs revealed a characteristic HS structure. These data begin to provide a structural understanding of the role of HS in cell-cell interactions, cell signaling

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Isolation and identification of glycosaminoglycans associated with purified nuclei from rat liver

Cited by 57 publications

References 30 publications

Association of hyaluronan with rat vascular endothelial and smooth muscle cells.

Association of hyaluronan with rat vascular endothelial and smooth muscle cells.

Hyaluronidase reduces human breast cancer xenografts in SCID mice

Isolation and characterization of heparan sulfate from various murine tissues

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