Glycosaminoglycans provide a binding site for thyroglobulin in orbital tissues of patients with thyroid-associated ophthalmopathy.
The presence of thyroglobulin (Tg) in orbital tissues of patients with thyroid-associated ophthalmopathy (TAO) supports a role of Tg in TAO pathogenesis. To search for Tg-binding sites in orbital tissues, because Tg is a heparin-binding protein, we investigated its binding to glycosaminoglycans (GAGs) that are abundant in orbital tissues: chondroitin sulfate B (CSB) and C (CSC) and hyaluronic acid (HA). Both in solid phase and solution phase assays purified human Tg bound to GAGs. In solid-phase assays, binding was increased by coincubation with heparin or GAGs in solution, or with an antibody against a Tg heparin-binding sequence (Arg2489-Glu2503), possibly suggesting crosslinking of Tg molecules induced by GAGs or by the presumably bivalent antibody. Orbital tissue extracts from TAO patients that contained Tg were subjected to high-salt treatment, which resulted in separation of Tg from GAGs, as observed by column chromatography. After separation from GAGs, the Tg in orbital tissue extracts acquired the ability to bind to immobilized CSB, and heparin enhanced binding, resembling the findings with purified human Tg. Therefore, we conclude that GAGs provide binding sites for Tg in orbital tissues, which may explain the presence of Tg in orbital tissues of patients with TAO.
Marinò
Michele
M
Department of Endocrinology, University of Pisa, Pisa, Italy. m.marino@endoc.med.unipi.it
Lisi
Simonetta
S
Pinchera
Aldo
A
Marcocci
Claudio
C
Menconi
Francesca
F
Morabito
Eugenia
E
Macchia
Marco
M
Sellari-Franceschini
Stefano
S
McCluskey
Robert T
RT
Chiovato
Luca
L
D000273:Adipose Tissue / Q000378:metabolism*; D000368:Aged; D001665:Binding Sites; D002809:Chondroitin Sulfates / Q000378:metabolism; D003871:Dermatan Sulfate / Q000378:metabolism; D005260:Female; D006025:Glycosaminoglycans / Q000737:chemistry / Q000378:metabolism*; D006111:Graves Disease / Q000378:metabolism* / Q000601:surgery; D006493:Heparin / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D006820:Hyaluronic Acid / Q000378:metabolism; D008297:Male; D008875:Middle Aged; D009915:Orbit / Q000378:metabolism*; D010047:Ovalbumin / Q000378:metabolism; D011485:Protein Binding; D013954:Thyroglobulin / Q000737:chemistry / Q000378:metabolism*
D006025:Glycosaminoglycans; D003871:Dermatan Sulfate; D006820:Hyaluronic Acid; D006493:Heparin; D010047:Ovalbumin; D002809:Chondroitin Sulfates; D013954:Thyroglobulin