Comparisons of the ability of follicular fluid glycosaminoglycans and chemically desulfated heparin to compete for heparin-binding sites on granulosa cells.
Glycosaminoglycans (GAGs) were purified from bovine follicular fluid, and their effectiveness to compete for heparin-binding sites in granulosa cells was evaluated. The GAGs dermatan sulfate (DS) and heparan sulfate (HS) were purified by anion-exchange high-performance liquid chromatography. Approximately 5 micrograms of protein from suspensions of bovine granulosa cells were incubated with 101 pmoles of [3H]heparin and 0.01-5.0 mg/ml of HS or DS for 2 h at 37 degrees C in 40 mM tris(hydroxymethyl)aminomethane (Tris), pH 7.35. Heparan sulfate obtained from small and medium follicles displaced [3H]heparin in a dose-dependent manner from 0.1 to 5 mg/ml, but HS from large follicles did not displace [3H]heparin. The DS obtained from small, medium, and large follicles displaced [3H]heparin in a dose-dependent manner, and the potency of the DS to displace [3H]heparin increased as the size of the follicles from which the DS was purified increased. Those results were independent of the maturational state of the granulosa cells. In a separate experiment, heparin (17.1% sulfate) was N-desulfated (11.8%), and the desulfated heparin did not displace [3H]heparin. It was concluded that the effectiveness of follicular HS and DS to compete for heparin-binding sites on granulosa cells was dependent on the maturation of the follicle from which the fluid was obtained rather than on the source of granulosa cells. The binding interaction of the GAGs relies, to some extent, on the presence and positions of sulfate moieties.
Bellin
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Endocrinology-Reproductive Physiology Program, University of Wisconsin, Madison 53706.
Wentworth
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D000818:Animals; D001667:Binding, Competitive; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003871:Dermatan Sulfate / Q000378:metabolism; D005260:Female; D006025:Glycosaminoglycans / Q000378:metabolism*; D006107:Granulosa Cells / Q000378:metabolism*; D006493:Heparin / Q000378:metabolism*; D006497:Heparan Sulfate / Q000378:metabolism; D006080:Ovarian Follicle / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism*
D003871:Dermatan Sulfate; D006025:Glycosaminoglycans; D006493:Heparin; D006497:Heparan Sulfate; D011956:Receptors, Cell Surface; C037680:heparin receptor