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  • 1
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 134 (1988), S. 376-386 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The purpose of this study was to examine the nature of the linkage between cell-surface hyaluronate and the plasma membrane. To accomplish this, rat fibrosarcoma cells were cultured in the presence of [3H]-acetate to isotopically label the hyaluronate, and then fixed with glutaraldehyde, which cross-links proteins but does not react directly with hyaluronate. The glutaraldehyde fixation stabilized the cells so that they could be manipulated in ways which would otherwise destroy cells. The fixed cells were then subjected to various treatments, and the amount of hyaluronate remaining on the cell surface was assayed via exhaustive digestion with Streptomyces hyaluronidase. Using this technique, we found that (1) cell-surface hyaluronate was quite stable for extended periods of time even in the presence of a large excess of non-labeled hyaluronate; (2) 4 M guanidine HCI and detergents did not extract a significant portion of cell-surface hyaluronate; (3) solutions of varying ionic strength (0-1 M NaCI) had no effect on the retention of hyaluronate; (4) the cell coat was stable in the range of pH 4-11, but outside this range a significant amount of hyaluronate was released; and (5) treatment with proteases released cell-surface hyaluronate. These results are consistent with the possibility that hyaluronate is covalently linked to a protein associated with the plasma membrane. Further support for this model came from experiments with the detergent Triton X-114, which can be used to separate soluble proteins from hydrophobic proteins. When nonfixed rat fibrosarcoma cells were extracted with this detergent and then partitioned by centrifugation, approximately 30 times as much hyaluronate was present in the detergent fraction which contained the hydrophobic proteins, as compared to the extracts pretreated with trypsin prior to phase separation. Again, these results suggest that cell-surface hyaluronate is directly linked to a hydrophobic core protein intercalated in the plasma membrane.
    Additional Material: 7 Ill.
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  • 2
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 110 (1982), S. 123-128 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The sizes of hyaluronate-containing coats on the surfaces of parent and virus-transformed cell lines (3T3 vs. SV-3T3; BHK vs. PY-BHK) were compared according to the method of Clarris and Fraser (1968, Exp. Cell Res., 49: 181-193) in which fixed red blood cells were allowed to settle slowly on the surface of culture dishes containing the cells. The coats were seen as areas devoid of red blood cells surrounding each of the cultured cells and could be destroyed by the addition of small amounts of streptomyces hyaluronidase, an enzyme specific for hyaluronate. In the case of the parent cell lines (3T3 and BHK), the coats were clearly visible, whereas for their virus-transformed counterparts (SV-3T3 and PY-BHK), the coats were either greatly reduced or absent. To confirm these observations, the amount of hyaluronate associated with each of the cell lines was measured using a direct chemical assay and shown to be significantly greater for the parent cell lines than for their virus-transformed counterparts. In addition, the parent cell lines secreted greater amounts of hyaluronate into the medium and retained a larger fraction of the total amount of hyaluronate at the cell surface than the virus-transformed cells. Thus the larger amount of hyaluronate on the surfaces of the parent cell types may be the result of both a faster rate of production and a decreased rate of release.
    Additional Material: 1 Ill.
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 90 (1977), S. 53-59 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: Heparan sulfate from the surface of a variety of mouse cells at different cell densities was examined by ion-exchange chromatography. The results of this analysis show that: 1The heparan sulfate from new isolates of Swiss 3T3 cells transformed SV40 virus (a DNA tumor virus) elutes from DEAE-cellulose at a lower ionic strength than that from the parent cell type. This finding confirms our earlier observation with an established SV40-transformed cell line (Underhill and Keller, 1975) and eliminates the possibility that this change is caused by extended passage in culture.2For both parent and transformed 3T3 cells, the heparan sulfates from low and high density cultures were the same as judged by chromatography on DEAE-cellulose. This result demonstrates that the transformation-dependent change which we have observed is independent of cell density.3The heparan sulfate from Balb/c 3T3 cells transformed with Kirsten murine sarcoma virus (an RNA tumor virus) elutes from DEAE-cellulose prior to that from parent Balb/c 3T3 cells. This result extends the transformation dependent change in heparan sulfate to the Balb/c 3T3 cell line and to cells transformed with an RNA virus.
    Additional Material: 8 Ill.
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Journal of Cellular Physiology 126 (1986), S. 352-358 
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: The effects of different carbohydrates on cell-to-cell adhesion were examined in an aggregation assay, which consisted of swirling a suspension of cells and monitoring the loss of single cells with a Coulter Counter. Of the carbohydrates tested, only heparin and dextran sulfate induced cell aggregation. This effect occurred in freshly isolated mouse splenocytes and in cultured cells of lymphoid origin (P388, YAA-CI) but not in cell lines of fibroblastic origins (3T3, SV-3T3, BHK, and PY-BHK). Using the YAA-CI cell line for further study, we found that aggregation could be induced by relatively small amounts of heparin (〈10 μg/ml). Binding experiments with 3H-heparin showed that under normal physiological conditions each YAA-CI cell bound approximately 2 × 106 molecules of heparin at saturation with a Kd of 3.5 × 10-7 M. This binding was blocked by both unlabelled heparin and dextran sulfate but not by other carbohydrates. When the pH of the medium was decreased, the heparin-induced aggregation was inhibited, and the Kd of the 3H-heparin binding was increased. In a similar fashion, when the ionic strength of the medium was increased, heparin-induced aggregation was inhibited and the Kd of the interaction was increased. These results suggest that the aggregation is inversely related to the Kd of the interaction and that the binding of heparin to the cell surface is primarily of an ionic nature.
    Additional Material: 5 Ill.
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  • 5
    ISSN: 0021-9541
    Keywords: Life and Medical Sciences ; Cell & Developmental Biology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Medicine
    Notes: In the present study, we examined a panel of human breast cancer cell lines with regard to their expression of CD44 and ability to bind and degrade hyaluronan. The cell lines expressed varying amounts of different molecular weight forms of CD44 (85-200 kDa) and, in general, those that expressed the greatest amounts of CD44 were the most invasive as judged by in vitro assays. In addition, the ability to bind and degrade hyaluronan was restricted to the cell lines expressing high levels of CD44, and both these functions were blocked by an antibody to CD44 (Hermes-1). Moreover, the rate of [3H]hyaluronan degradation was highly correlated with the amount of CD44 (r = 0.951, P 〈 0.0001), as well as with the invasive potential of the cells. Scatchard analysis of the [3H]hyaluronan binding of these cells revealed the existence of significant differences in both their binding capacity and their dissociation constant. To determine the source of this deviation, the different molecular weight forms of CD44 were partially separated by gel filtration chromatography. In all cell lines, the 85 kDa form was able to bind hyaluronan, although with different affinities. In contrast, not all of the high molecular weight forms of CD44 had this ability. These results illustrate the diversity of CD44 molecules in invasive tumor cells, and suggest that one of their major functions is to degrade hyaluronan. © 1994 Wiley-Liss, Inc.
    Additional Material: 7 Ill.
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  • 6
    Publication Date: 2004-01-15
    Description: For circulating lymphocytes to migrate to inflammatory sites, they must first adhere to the target tissue endothelium with sufficient strength to overcome the shear forces of blood flow. We previously reported that dermal papillary vessels in acute graft-versus-host disease (aGVHD) support shear-resistant lymphocyte adherence. We now identify the relevant adhesion molecule(s) directing this binding, showing that interactions between lymphocyte CD44 and hyaluronic acid (HA) expressed on dermal vessels in aGVHD alone confer this shear-resistant attachment. Native HA deposits on vascular endothelium support lymphocyte adherence, whereas HA immobilized on plastic does not. HA expressed at dermal endothelium in aGVHD is thus specialized to support lymphocyte adherence under flow conditions, and CD44-HA interactions may contribute to lymphocytotropism to skin in aGVHD.
    Print ISSN: 0006-4971
    Electronic ISSN: 1528-0020
    Topics: Biology , Medicine
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  • 7
  • 8
    Publication Date: 1993-02-01
    Print ISSN: 0012-1606
    Electronic ISSN: 1095-564X
    Topics: Biology
    Published by Elsevier
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  • 9
    Publication Date: 1988-10-01
    Print ISSN: 0014-4827
    Electronic ISSN: 1090-2422
    Topics: Biology , Medicine
    Published by Elsevier
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  • 10
    Publication Date: 1981-02-01
    Print ISSN: 0014-4827
    Electronic ISSN: 1090-2422
    Topics: Biology , Medicine
    Published by Elsevier
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