Chinese hamster ovary cell adhesion to human platelet thrombospondin is dependent on cell surface heparan sulfate proteoglycan

Paul Kaesberg, W. B. Ershler, J. D. Esko, D. F. Mosher

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Abstract

Thrombospondin is a 420-kD platelet alpha-granule glycoprotein that binds specifically to heparin. We examined adhesion to thrombospondin of CHO K1 cells and three mutant CHO lines with varying deficiencies in glycosaminoglycan (GAG) synthesis. In an experiment in which the parent line (K1) had 78% adherence to thrombospondin adsorbed to tissue culture plastic, CHO S745 cells, with < 6% normal GAG synthesis had 11% adherence. CHO S677 cells, with decreased heparan sulfate proteoglycan but increased chondroitin sulfate proteoglycan, had 42% adherence. CHO S803 cells, with decreased heparan sulfate proteoglycan and normal chondroitin sulfate proteoglycan, had 31% adherence. Heparin inhibited K1 cell adhesion to thrombospondin, but not fibronectin, in a concentration-dependent manner. Dermatan sulfate but not chondroitin sulfate was also inhibitory. There was markedly decreased K1 cell adhesion to a thrombospondin core fragment that lacked the heparin binding NH2-terminal domain. Purified heparin binding domain, although poorly adhesive when adsorbed to substratum, inhibited cell adhesion to intact thrombospondin. Adhesion was better for all cell lines tested, including three human tumor cell lines, when thrombospondin was adsorbed at pH 4.0 compared with pH 7.4. When adsorption of thrombospondin was done at pH 7.4, cell adhesion was better when thrombospondin was adsorbed in the presence of ≥ 0.6 mM calcium, compared to 0.1 mM calcium or EDTA. These finding suggest that thrombospondin can adsorb to plastic with varying degrees of exposure of a cell adhesion domain. We conclude that the thrombospondin cell adhesion receptor on CHO cells is a heparan sulfate proteoglycan, and that cell adhesion to thrombospondin depends on conformation of adsorbed thrombospondin.

Original languageEnglish (US)
Pages (from-to)994-1001
Number of pages8
JournalJournal of Clinical Investigation
Volume83
Issue number3
StatePublished - 1989
Externally publishedYes

Fingerprint

Thrombospondins
Heparan Sulfate Proteoglycans
Cricetulus
Cell Adhesion
Ovary
Blood Platelets
CHO Cells
Heparin
Chondroitin Sulfate Proteoglycans
Glycosaminoglycans
Plastics
Calcium
Tumor Cell Line
Fibronectins
Edetic Acid
Adhesives
Adsorption

ASJC Scopus subject areas

  • Medicine(all)

Cite this

Chinese hamster ovary cell adhesion to human platelet thrombospondin is dependent on cell surface heparan sulfate proteoglycan. / Kaesberg, Paul; Ershler, W. B.; Esko, J. D.; Mosher, D. F.

In: Journal of Clinical Investigation, Vol. 83, No. 3, 1989, p. 994-1001.

Research output: Contribution to journalArticle

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abstract = "Thrombospondin is a 420-kD platelet alpha-granule glycoprotein that binds specifically to heparin. We examined adhesion to thrombospondin of CHO K1 cells and three mutant CHO lines with varying deficiencies in glycosaminoglycan (GAG) synthesis. In an experiment in which the parent line (K1) had 78{\%} adherence to thrombospondin adsorbed to tissue culture plastic, CHO S745 cells, with < 6{\%} normal GAG synthesis had 11{\%} adherence. CHO S677 cells, with decreased heparan sulfate proteoglycan but increased chondroitin sulfate proteoglycan, had 42{\%} adherence. CHO S803 cells, with decreased heparan sulfate proteoglycan and normal chondroitin sulfate proteoglycan, had 31{\%} adherence. Heparin inhibited K1 cell adhesion to thrombospondin, but not fibronectin, in a concentration-dependent manner. Dermatan sulfate but not chondroitin sulfate was also inhibitory. There was markedly decreased K1 cell adhesion to a thrombospondin core fragment that lacked the heparin binding NH2-terminal domain. Purified heparin binding domain, although poorly adhesive when adsorbed to substratum, inhibited cell adhesion to intact thrombospondin. Adhesion was better for all cell lines tested, including three human tumor cell lines, when thrombospondin was adsorbed at pH 4.0 compared with pH 7.4. When adsorption of thrombospondin was done at pH 7.4, cell adhesion was better when thrombospondin was adsorbed in the presence of ≥ 0.6 mM calcium, compared to 0.1 mM calcium or EDTA. These finding suggest that thrombospondin can adsorb to plastic with varying degrees of exposure of a cell adhesion domain. We conclude that the thrombospondin cell adhesion receptor on CHO cells is a heparan sulfate proteoglycan, and that cell adhesion to thrombospondin depends on conformation of adsorbed thrombospondin.",
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