Rheumatoid arthritis (RA) is a widely prevalent (1-3%) chronic systemic disease thought to have an autoimmune component; both humoral and cellular mechanisms have been implicated. Primary osteoarthritis (OA) is considered to be distinct from rheumatoid arthritis, and here damage is thought to be secondary to cartilage degeneration. In rheumatoid arthritis, immune complexes are present that consist exclusively of immunoglobulin, implying that this is both the 'antibody' (rheumatoid factor [RF]) and the 'antigen' (most commonly IgG). Autoantigenic reactivity has been localized to the constant-region (C gamma 2) domains of IgG. There is no evidence for a polypeptide determinant but carbohydrate changes have been reported. We have therefore conducted a study, simultaneously in Oxford and Tokyo, to compare in detail the N-glycosylation pattern of serum IgG (Fig. 1) isolated from normal individuals and from patients with either primary osteoarthritis or rheumatoid arthritis. The results, which required an evaluation of the primary sequences of approximately 1,400 oligosaccharides from 46 IgG samples, indicate that: (1) IgG isolated from normal individuals, patients with RA and patients with OA contains different distributions of asparagine-linked bi-antennary complex-type oligosaccharide structures, (2) in neither disease is the IgG associated with novel oligosaccharide structures, but the observed differences are due to changes in the relative extent of galactosylation compared with normal individuals. This change results in a 'shift' in the population of IgG molecules towards those carrying complex oligosaccharides, one or both of whose arms terminate in N-acetylglucosamine. These two arthritides may therefore be glycosylation diseases, reflecting changes in the intracellular processing, or post-secretory degradation of N-linked oligosaccharides.

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(4,5). AP remains in amyloid deposits for prolonged periods without protein catabolism (6) and might thereby contribute to the persistence which underlies their pathogenicity. However, the structure ofAP has not been well characterized. AP from only one individual has been completely sequenced at the protein level (7), and the two reports of the SAP cDNA sequence (8, 9), as well as preliminary reports of the glycostructure (10,11,37), are conflicting. We have therefore compared the complete covalent structures of SAP and AP, with particular emphasis on the oligosaccharide and its role in turnover of the SAP glycoprotein.MATERIALS AND METHODS Proteins. SAP was isolated from the serum (13) of two healthy subjects, from two large pools of normal serum containing 1-ml aliquots from each of >5000 different healthy volunteer blood donors, and from a pool of malignant effusion fluids containing material from >300 donors. AP was isolated from two spleens containing AA amyloid, two spleens and two livers with AL amyloid, one spleen containing [ArgW]apolipoprotein A-I amyloid, and the spleen, liver, heart, kidney, and adrenals of an individual patient with AA amyloidosis. Amyloidotic tissue was homogenized in 0.01 M Tris/0.14 M NaCl/0.01 M EDTA, pH 8.0, and the supernatant after centrifugation at 50,000 x g for 30 min was dialyzed extensively into 0.01 M Tris/0.14 M NaCl/0.002 M CaCl2, pH 8.0, before extraction of AP precisely as described for SAP from serum (13). All SAP/AP preparations were >991% pure in heavily overloaded SDS/polyacrylamide gradient gels (Excelgel, Pharmacia) run under reducing conditions and stained with brilliant blue R-350 or with silver.Glycan Structure Analysis. Oligosaccharides attached to SAP and AP were released by hydrazinolysis and radiolabeled with sodium borotritide at the reducing terminus, which incorporates a single label into each chain independently of its structure, so that the relative molar proportions of all glycans present can be estimated directly (14). These oligosaccharides were analyzed by high-voltage paper electrophoresis in pyridine/acetic acid water (3:1:387 by volume) at pH 5.4, 80 V/cm. They were then converted to neutral glycans by neuraminidase digestion and were analyzed on a highresolution 1.5 cm x 2 m Bio-Gel P-4 (<400 mesh) column by elution at 550C with water (14). The structure was determined by sequential exoglycosidase digestion in combination with permethylation and GS-MS (14). Heterogeneity of sialic acid linkage in the A-1 and A-2 structures was determined by digestion with neuraninidase from Newcastle disease virus, specific for a2-3 bonds, and neuraminidase from Arthrobacter ureafaciens, which cleaves a2-3(6) bonds, followed by selective exoglycosidase digestion and product analysis using permethylation and GC-MS (14). Oligosaccharide Modification. Bovine al acid glycoprotein (AGP) (Sigma) was desialylated by incubation at 800C for 1 hr at 2-5 mg/ml in 12.5 mM H2SO4 (16), resulting in the expected alteration in electrophoretic mobility in 1% agarose gel i...

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The proportion of oligosaccharide chains on the Fc fragment of IgG which terminate with N-acetylglucosamine (GIcNAc) rather than galactose is increased in rheumatoid arthritis and tuberculosis, and in sera from patients with Crohn's disease, probably because of decreased activity of a galactosyltransferase in B lymphocytes. We We confirm here that the percentage of agalactosyl IgG rises in Crohn's disease, show that it also rises in a minority of patients with ulcerative colitis, and investigate its relationship to clinical and biochemical parameters in both diseases.