Purification of amyloid-associated heparan sulphate proteoglycans and galactosaminoglycan free chains from human tissues.
Basement membrane-associated heparan sulphate proteoglycans have been demonstrated immunohistochemically in organs from patients afflicted with various types of amyloidosis. In a recent report, we were able to isolate and partly characterize a basement membrane-associated heparin sulphate proteoglycan from human hepatic amyloid. In the present study proteoglycans were extracted with guanidine from human amyloid-laden kidney, spleen and lymph nodes. All tissues extracted with guanidine contained both heparan sulphate proteoglycan (HSPG) and galactosaminoglycan (CS/DS) free chains. Tissue staining using a monoclonal antibody against basement membrane HSPG revealed the presence of HSPG in amyloid deposits in kidney and spleen. Furthermore, following SDS-PAGE of HSPG from kidney after deaminative cleavage of the HS chains, a 15-kDa and 80-kDa protein appeared, probably representing the core protein(s). In lymph node HSPG, three core proteins of 65, 30 and 25 kDa could be demonstrated on SDS-PAGE, the first reacting with the anti-basement membrane HSPG antibody when subjected to Western blotting subsequent to SDS-PAGE. By immunohistochemistry, we failed to demonstrate any staining of the renal and splenic tissue sections employing an antibody against the decorin core protein.
Stenstad
T
T
Department of Rheumatology, University of Tromsø, Norway.
Magnus
J H
JH
Husby
G
G
Kolset
S O
SO
D000328:Adult; D000682:Amyloid / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D006497:Heparan Sulfate / Q000302:isolation & purification*; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D007668:Kidney / Q000737:chemistry*; D008198:Lymph Nodes / Q000737:chemistry*; D008297:Male; D011134:Polysaccharides / Q000302:isolation & purification*; D011509:Proteoglycans / Q000302:isolation & purification*; D013154:Spleen / Q000737:chemistry*
D000682:Amyloid; D006497:Heparan Sulfate; D011134:Polysaccharides; D011509:Proteoglycans; C007815:galactosaminoglycan