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Altered oligosaccharides as the initiating autoantigen in rheumatoid arthritis
Fedical Hypotheses 10: 347-352, 1985
ALTERED OLIGOSACCHARIDES RHEUMATOID ARTHRITIS
AS THE INITIATING AUTOANTIGEN
IN
Edward E. Gordon, M.D., Chairman, Department of Rehabilitation Medicine, Alexian Brothers Medical Center, Elk Grove Village, Illinois 60007, USA ABSTRACT The linkage of rheumatoid arthritis (RA) to infection though vague has remained a persistent notion for the past fifty years. The hypotheses that streptococcus was responsible; then M. Pneumoniae; the implication of a viral pathogenesis and, more especially, the Epstein-Barr virus, have all not withstood the test of Koch's postulates. Today, autoimmunity holds the field in the pathogenesis of RA. We know in fair detail what happens after failure of selfrecognition develops. Why does synovial tissue become antigenic? Altered receptors of cell surface may be the loci of change, namely, glycoproteins and glycolipids. A hypothesis is constructed based on alteration by viral DNA of these cell surface markers that are Evidence is drawn from insulin dependent then recognized as alien. juvenile diabetes, glomerulonephritis, and Landsteiner blood groups. The initial lesion may be initiated by an infectious agent leading to a chain of events, the first link constituting change of the psecific glycoprotein or glycolipid of a cell to one of antigenic challenge, thus stimulating an autoimmune reaction. The ultimate destructive-proliferative sequences of RA are then set in motion. INTRODUCTION Rheumatoid arthritis (RA) is regarded as an immune disease in which joint and tendon synovia are the primary targets. We are still ignorant of the nature of the initiating autoimmune event that finally leads to synovial and joint destruction. Therefore, the mechanism makes a justifiable subject for speculation. I shall not discuss in any detail the lymphoimmune system, which plays a major part in perpetuating the tissue injury. These subsequent developments, broadly speaking, are fairly clear: lymphcytic and monocytic offshoots evolve which manufacture various immunoglobulins and enzymes leading to destruction of synovial tissue (1,Z). Crucial to injury are antibody-antigen reactions, as well as liberation of proteolytic enzymes, prostaglandins and other cytotoxic substances. But the exciting auto-antigen remains unknown.
347
HYPOTHESIS A hypothesis of the initiation of auto-antigenicity in RA is based on two other autoimmune diseases. An infectious agent, virus or otherwise, invades the cell, tampers with the indigenous DNA, and produces enzyme dyssynthesis, altering glycosyl (oligosaccharides) groups of glycoproteins and glycolipids on the cell membranes. The altered oligosaccharides are sensed as non-self substances, leading to antigenic challenge, Susceptibility to induced autoantigenicity of tissue is genetically determined, linked to the HLA-D locus of chromosome 6 (2,3). Rheumatoid Arthritis The ever recurring ghost of the infectious origin of RA has appeared in the past one hundred years. In the 193Os, Cecil presented evidence for a streptococcal etiology of RA, but his hypothesis has been discarded. More recently mycoplasma has drawn attention, but currently the opinion is strong for the implication of the EpsteinBarr virus (4). Incidence shows a seasonal trend (fall and winter) suggesting an infectious onset. Immunologically, RA is associated with the HLA-DR4 locus (2,3). These characteristics are reminiscent of juvenile diabetes. Juvenile Diabetes A disease that appears to represent strongly rheumatoid arthritis in pathogenesis is juvenile diabetes (JD), which links prior infection to an autoimmune response to the beta calls of the pancreas (5-7). A mild infection for weeks or months often precedes the onset of JD, the end result of which is an autoimmune destruction of the islets of Langerhans with the following features (6): 1. 2. 3. 4. 5. 6.
1::
necrosis of islet cells with lymphocytic infiltration, presence of an IgG antibody against Coxsackie B virus, IgG antibodies directed against islet cells isolated by immunofluorescence, complement dependent beta cell antibody, a number of antibodies with or without complement directed against various beta cells components, cytotoxicity of serum from patients with JD for animal islet cells in vivo and in vitro, cytotoxicity of JD lymphocytes for islet cells in animals, seasonal variation in incidence of JD--highest in fall and winter, never in summer, family history in 10% of patients, HLA association of types DR3 and DR4 antigens.
Comparing these findings with RA (Table 1), a strong parallelism exists for epidemiological and immunological characteristics.
348
Similarities
Table I.
between JD and RA (1,5)
-RA
-JD Remissions
yes
yes
HLA association
BR3 UR4
UR4
Seasonal trend
fall/winter
winter
Symptoms
acute/ subacute
subacute/ insidious
Family history
5-10%
8%
Incidence/year
lo-15/105
?
Suspected virus
myxovirus, Coxsackie
Epstein-Barr
Uses complement
yes
yes
While these do not prove the absolute existence of an infectious agent, one may entertain the hypothesis that such an agent, possibly the Epstein-Barr virus, initiates the organ inflammation and degeneration of RA by auto-immune activity. How is the elaboration of the autoantigen effected? Glycoproteins
and Glycolipids
The character of the cell surface determines the ability of the immune system to recognize the cell as self or foreign (8). Important constituents of the cell surface are glycoproteins and glycolipids. Practically all proteins (except albumin) are linked to short polymers of hexose or substituted hexose molecules through one of five specific amino acids (9,lO). At the maximum these polymers contain no more than fifteen glycosyl radicals; thus they are designated oligosaccharides, in contradistinction to the polysaccharides of proteoglycans. Oligosaccharides are also joined to lipids such as ceramide, in the (9). These sugar residues mark the cells as part of cell membrane self. This function has been particularly well defined in Landsteiner blood groups (9-11). If through some aberration these markers are sensed as foreign, an immune reaction may be directed against the cells initiating the well-known destructive results of auto-immune disease.
In anticipation of the autoimmune model of glomerulonephritis to be presented below, I wish to draw attention to siliac acid, or n-acetyl neuraminic acid, structurally an N-acetyl hexose and a common constituent of oligosaccharides. Other frequently occurring glycosyl radicals are g!actose, glucose, N-acetyl galactose and glucose amines. The free and N-acetylated galactose stand out as differentiating 349
cell markers of the ABO blood group (9-11). Cellular synthesis of the oligosaccharides requires a large number of transferase enzymes to join two glycosyl units at specific carbon atoms (9). Dyssynthesis of a transferase may change the configuration of a glycoprotein, possibly endowing it with antigenic properties and initiating an auto-immune reaction, as glomerulonephritis possibly implies. The Landsteiner Blood Group ABO To illustrate the transformation of an altered glycoprotein on the cell surface into an antigen, we can perform an imaginary experiment on the Landsteiner A-B blood types. The only difference between surface glycosyl groups of A and B cells is that A has n-acetyl galactosamine as the terminal radical in the oligosaccharide chain while the latter has a free galactose (9-11). The rest of the A and B chain is identical. Type B serum has the antibody for A cells; type A serum, for B cells. Now, if by some manipulation of type A blood we can convert the terminal glycosyl, N-acetyl galactosamine, to galactose, what happens? Blood A will now contain cells with B type markers -- terminal galactose radicals -- which are antigenic for A serum, resulting in an antigen-antibody reaction. We have artificially produced an auto-immune disease in type A blood. While this manipulation is artificial, a terminal tetrasaccharide of the group B polymer has been synthesized (12) and antigenic properties demonstrated in the synthetic glycosyl polymer (13). Streptococcal Glomerulonephritis We can now consider this disease, characterized by a precipitating infection and a perpetuating auto-immune process. A recent study of thirty-nine patients early inthecourse ofglomerulonephritis reported eight to have serum n-acetyle neuraminidase activity (14). Normal individuals and patients with other types of renal disease have no such activity at all. Of these patients, all ASLO positive, twenty-eight above 2 SD of the mean of thirty-two controls (0.57 + 1.26), yielded a mean serum level of 10.8 pmol/dl of sialic acid. The elevation occurred within the first eight weeks from onset, then gradually disappeared. Increased concentration of n-acetyl neuraminic acid was not a feature of other types of renal disease such as SLE, pyelitis, membrano-proliferative glomerulonephritis; nor did elevation occur in rheumatic fever, tonsilitis and other Group A and B streptococcal diseases. It is known that streptococcus elaborates a neuraminidase. The question arises whether the elevation of sialic acid in the patients derives from the splitting of this particular hexose residue from the native oligosaccharide on glomerular endothelium. Such action may convert the cell markers into an antigen, which then is challenged by the autoimmune system. The high levels of serum neuraminidase activity and neuraminic acid are no conclusive proof of this explanation of antigenicity in glomerulonephritis. But these findings taken in conjunction with glycosyl determination of blood group 350
specificity are presented to suggest a possible relationship between infection and antigenicity by alteration of cell surface glycoproteins and glycolipids. On the basis of the three paradigms discussed above, one hypothesis of the origin of RA is presented in Table II.
Table II.
Hypothesis
of origin of antigenicity
in RA
Viral Genome -- host DNA -- enzyme dyssynthesis Alteraton
of glycosyl residue of surface oligosaccharides
Antigenic
challenge of altered glycoprotein
(-lipid)
Immune response to altered cell marker Synovial cytotoxicity Rheumatoid
arthritis
The infective viral DNA (Epsten-Barr?) interferes with the indigenous DNA which then transmits faulty instructions for oligosaccharide This results in antigenic glycoproteins and glycolipids synthesis. on the cell membrane. Henceforth, the lymphocytic immune system is aroused against a "foreign" cell with the initiation of synovial inflammation characteristic of RA. CONCLUSION We know nothing of synovial cell glycoproteins in normal joints and in rheumatoid joints shortly after the onset of the disease. The biochemistry of glycoproteins and glycolipids has been enormously enriched in the past ten years both in their isolation and synthesis (8,9,11,12), and immunochemistry of their oligosaccharides is already being pursued (13). These tools offer a means of testing the hypothesis that oligopolymers play a role in the genesis of antibody formation in synovial tissue. While glycoproteins have been measured in serum of pregnant women with rheumatoid arthritis (15), no studies are available in synovium. This tissue can be obtained from surgical specimens with intact synovium and from biopsy and other surgical procedures in rheumatoid patients. The first step in testing this hypothesis would be comparison of the glycosyl residues from the two sources of synovia. By analogy to the findings in glomerulonephritis showing glycoprotein changes early in the disease, synovial biopsies would require similar testing. Isolation of terminal pure polysaccharides, then synthesis would provide probes for testing antigenicity in animals, both of normal and altered glycosyl residues,
351
REFERENCES 1.
Zvailfler NJ. Etiology and pathogenesis of rheumatoid arthritis. p. 417 in Arthritis and Allied Conditions. ed, McCarthy DJ. 9th ed, Lea and Ferbiger, Philadelphia, 1979.
2.
Arthritis
3.
McDevelt HO. compatibility ed. McCarthy
4.
idem. p. 143.
5.
Rabin BS. Immunopathology J, p. 39, Jan 1981.
6.
Cahill GF, Jr, McDevitt HO. Insulin dependent diabetes mellitus: the initial lesion. N Eng J Med 304:1454, 1981.
and rheumatism.
24th Rheumatism
Review.
24(2):133,
1981.
Genetic structure and function of the major histocomplex. p. 229 in Arthritis and Allied Conditions. DJ, 9th ed, Lea and Ferbiger, Philadelphia, 1979.
of insulin dependent
diabetes.
Ill Med
7.
Notkins AL.
8.
Nicolson GL. Transmembrane control of the receptors on normal and tumour cells. I cytoplasmic influence over cell surface components. Biochem Biophys Acta 457:57, 1976.
9.
Sweeley CC, Fung Y-K, Macher BA, Moskal JR and Nunez HA. Structure and metabolism of glycolipids. p. 46 in Glycoproteins and Glycolipids, ed. Walborg. Symposium 80, Am Chem Sot, 1980.
The cause of diabetes.
Sci Am 241:62, 1979.
10.
idem. Walborg EF, Jr. glycolipids, p. 8.
Current concepts of glycoproteins
11.
Watkins WM.
12.
Milat MS, Sinaij P. Chemical synthesis of human blood group B antigenic determinant: type 2 tetrasaccharide. Angew Chem Ed Engl 18:464, 1979.
13.
Feizi T, Wood E, Auge C, David S, Veyrieres A. Blood group I activities of synthetic oligosaccharides assessed by immunoImmunochemistry 15: 733, 1978. assay.
14.
Rodriguez-Iturbe B, Katizar VN, Coello J. Neuraminidase activity and free sialic acid levels in the serum of patients with acute posstreptococcal glomerulonephritis. N Eng J M 304:1506, 1981.
15.
Persellin RH, Weginton DAF, Rutstein JE, Cohen MJ, Goehrs HR, Steele D, Philips VK, Kradel RP. Pregnancy alpha-glycoprotein and rheumatoid arthritis; and Unger A, Kay A, Griffin AJ, Panayi GS. Disease activity and pregnancy-associated A2-glycoprotein in rheumatoid arthritis during pregnancy. Arthritis VIII Pan American Congress of Rheumatology. and Rheumatism. 24 (Suppl): 19, 1982 (Abstracts).
Blood group substances.
352
and
Science 152:172, 1966.
ALTERED OLIGOSACCHARIDES RHEUMATOID ARTHRITIS
AS THE INITIATING AUTOANTIGEN
IN
Edward E. Gordon, M.D., Chairman, Department of Rehabilitation Medicine, Alexian Brothers Medical Center, Elk Grove Village, Illinois 60007, USA ABSTRACT The linkage of rheumatoid arthritis (RA) to infection though vague has remained a persistent notion for the past fifty years. The hypotheses that streptococcus was responsible; then M. Pneumoniae; the implication of a viral pathogenesis and, more especially, the Epstein-Barr virus, have all not withstood the test of Koch's postulates. Today, autoimmunity holds the field in the pathogenesis of RA. We know in fair detail what happens after failure of selfrecognition develops. Why does synovial tissue become antigenic? Altered receptors of cell surface may be the loci of change, namely, glycoproteins and glycolipids. A hypothesis is constructed based on alteration by viral DNA of these cell surface markers that are Evidence is drawn from insulin dependent then recognized as alien. juvenile diabetes, glomerulonephritis, and Landsteiner blood groups. The initial lesion may be initiated by an infectious agent leading to a chain of events, the first link constituting change of the psecific glycoprotein or glycolipid of a cell to one of antigenic challenge, thus stimulating an autoimmune reaction. The ultimate destructive-proliferative sequences of RA are then set in motion. INTRODUCTION Rheumatoid arthritis (RA) is regarded as an immune disease in which joint and tendon synovia are the primary targets. We are still ignorant of the nature of the initiating autoimmune event that finally leads to synovial and joint destruction. Therefore, the mechanism makes a justifiable subject for speculation. I shall not discuss in any detail the lymphoimmune system, which plays a major part in perpetuating the tissue injury. These subsequent developments, broadly speaking, are fairly clear: lymphcytic and monocytic offshoots evolve which manufacture various immunoglobulins and enzymes leading to destruction of synovial tissue (1,Z). Crucial to injury are antibody-antigen reactions, as well as liberation of proteolytic enzymes, prostaglandins and other cytotoxic substances. But the exciting auto-antigen remains unknown.
347
HYPOTHESIS A hypothesis of the initiation of auto-antigenicity in RA is based on two other autoimmune diseases. An infectious agent, virus or otherwise, invades the cell, tampers with the indigenous DNA, and produces enzyme dyssynthesis, altering glycosyl (oligosaccharides) groups of glycoproteins and glycolipids on the cell membranes. The altered oligosaccharides are sensed as non-self substances, leading to antigenic challenge, Susceptibility to induced autoantigenicity of tissue is genetically determined, linked to the HLA-D locus of chromosome 6 (2,3). Rheumatoid Arthritis The ever recurring ghost of the infectious origin of RA has appeared in the past one hundred years. In the 193Os, Cecil presented evidence for a streptococcal etiology of RA, but his hypothesis has been discarded. More recently mycoplasma has drawn attention, but currently the opinion is strong for the implication of the EpsteinBarr virus (4). Incidence shows a seasonal trend (fall and winter) suggesting an infectious onset. Immunologically, RA is associated with the HLA-DR4 locus (2,3). These characteristics are reminiscent of juvenile diabetes. Juvenile Diabetes A disease that appears to represent strongly rheumatoid arthritis in pathogenesis is juvenile diabetes (JD), which links prior infection to an autoimmune response to the beta calls of the pancreas (5-7). A mild infection for weeks or months often precedes the onset of JD, the end result of which is an autoimmune destruction of the islets of Langerhans with the following features (6): 1. 2. 3. 4. 5. 6.
1::
necrosis of islet cells with lymphocytic infiltration, presence of an IgG antibody against Coxsackie B virus, IgG antibodies directed against islet cells isolated by immunofluorescence, complement dependent beta cell antibody, a number of antibodies with or without complement directed against various beta cells components, cytotoxicity of serum from patients with JD for animal islet cells in vivo and in vitro, cytotoxicity of JD lymphocytes for islet cells in animals, seasonal variation in incidence of JD--highest in fall and winter, never in summer, family history in 10% of patients, HLA association of types DR3 and DR4 antigens.
Comparing these findings with RA (Table 1), a strong parallelism exists for epidemiological and immunological characteristics.
348
Similarities
Table I.
between JD and RA (1,5)
-RA
-JD Remissions
yes
yes
HLA association
BR3 UR4
UR4
Seasonal trend
fall/winter
winter
Symptoms
acute/ subacute
subacute/ insidious
Family history
5-10%
8%
Incidence/year
lo-15/105
?
Suspected virus
myxovirus, Coxsackie
Epstein-Barr
Uses complement
yes
yes
While these do not prove the absolute existence of an infectious agent, one may entertain the hypothesis that such an agent, possibly the Epstein-Barr virus, initiates the organ inflammation and degeneration of RA by auto-immune activity. How is the elaboration of the autoantigen effected? Glycoproteins
and Glycolipids
The character of the cell surface determines the ability of the immune system to recognize the cell as self or foreign (8). Important constituents of the cell surface are glycoproteins and glycolipids. Practically all proteins (except albumin) are linked to short polymers of hexose or substituted hexose molecules through one of five specific amino acids (9,lO). At the maximum these polymers contain no more than fifteen glycosyl radicals; thus they are designated oligosaccharides, in contradistinction to the polysaccharides of proteoglycans. Oligosaccharides are also joined to lipids such as ceramide, in the (9). These sugar residues mark the cells as part of cell membrane self. This function has been particularly well defined in Landsteiner blood groups (9-11). If through some aberration these markers are sensed as foreign, an immune reaction may be directed against the cells initiating the well-known destructive results of auto-immune disease.
In anticipation of the autoimmune model of glomerulonephritis to be presented below, I wish to draw attention to siliac acid, or n-acetyl neuraminic acid, structurally an N-acetyl hexose and a common constituent of oligosaccharides. Other frequently occurring glycosyl radicals are g!actose, glucose, N-acetyl galactose and glucose amines. The free and N-acetylated galactose stand out as differentiating 349
cell markers of the ABO blood group (9-11). Cellular synthesis of the oligosaccharides requires a large number of transferase enzymes to join two glycosyl units at specific carbon atoms (9). Dyssynthesis of a transferase may change the configuration of a glycoprotein, possibly endowing it with antigenic properties and initiating an auto-immune reaction, as glomerulonephritis possibly implies. The Landsteiner Blood Group ABO To illustrate the transformation of an altered glycoprotein on the cell surface into an antigen, we can perform an imaginary experiment on the Landsteiner A-B blood types. The only difference between surface glycosyl groups of A and B cells is that A has n-acetyl galactosamine as the terminal radical in the oligosaccharide chain while the latter has a free galactose (9-11). The rest of the A and B chain is identical. Type B serum has the antibody for A cells; type A serum, for B cells. Now, if by some manipulation of type A blood we can convert the terminal glycosyl, N-acetyl galactosamine, to galactose, what happens? Blood A will now contain cells with B type markers -- terminal galactose radicals -- which are antigenic for A serum, resulting in an antigen-antibody reaction. We have artificially produced an auto-immune disease in type A blood. While this manipulation is artificial, a terminal tetrasaccharide of the group B polymer has been synthesized (12) and antigenic properties demonstrated in the synthetic glycosyl polymer (13). Streptococcal Glomerulonephritis We can now consider this disease, characterized by a precipitating infection and a perpetuating auto-immune process. A recent study of thirty-nine patients early inthecourse ofglomerulonephritis reported eight to have serum n-acetyle neuraminidase activity (14). Normal individuals and patients with other types of renal disease have no such activity at all. Of these patients, all ASLO positive, twenty-eight above 2 SD of the mean of thirty-two controls (0.57 + 1.26), yielded a mean serum level of 10.8 pmol/dl of sialic acid. The elevation occurred within the first eight weeks from onset, then gradually disappeared. Increased concentration of n-acetyl neuraminic acid was not a feature of other types of renal disease such as SLE, pyelitis, membrano-proliferative glomerulonephritis; nor did elevation occur in rheumatic fever, tonsilitis and other Group A and B streptococcal diseases. It is known that streptococcus elaborates a neuraminidase. The question arises whether the elevation of sialic acid in the patients derives from the splitting of this particular hexose residue from the native oligosaccharide on glomerular endothelium. Such action may convert the cell markers into an antigen, which then is challenged by the autoimmune system. The high levels of serum neuraminidase activity and neuraminic acid are no conclusive proof of this explanation of antigenicity in glomerulonephritis. But these findings taken in conjunction with glycosyl determination of blood group 350
specificity are presented to suggest a possible relationship between infection and antigenicity by alteration of cell surface glycoproteins and glycolipids. On the basis of the three paradigms discussed above, one hypothesis of the origin of RA is presented in Table II.
Table II.
Hypothesis
of origin of antigenicity
in RA
Viral Genome -- host DNA -- enzyme dyssynthesis Alteraton
of glycosyl residue of surface oligosaccharides
Antigenic
challenge of altered glycoprotein
(-lipid)
Immune response to altered cell marker Synovial cytotoxicity Rheumatoid
arthritis
The infective viral DNA (Epsten-Barr?) interferes with the indigenous DNA which then transmits faulty instructions for oligosaccharide This results in antigenic glycoproteins and glycolipids synthesis. on the cell membrane. Henceforth, the lymphocytic immune system is aroused against a "foreign" cell with the initiation of synovial inflammation characteristic of RA. CONCLUSION We know nothing of synovial cell glycoproteins in normal joints and in rheumatoid joints shortly after the onset of the disease. The biochemistry of glycoproteins and glycolipids has been enormously enriched in the past ten years both in their isolation and synthesis (8,9,11,12), and immunochemistry of their oligosaccharides is already being pursued (13). These tools offer a means of testing the hypothesis that oligopolymers play a role in the genesis of antibody formation in synovial tissue. While glycoproteins have been measured in serum of pregnant women with rheumatoid arthritis (15), no studies are available in synovium. This tissue can be obtained from surgical specimens with intact synovium and from biopsy and other surgical procedures in rheumatoid patients. The first step in testing this hypothesis would be comparison of the glycosyl residues from the two sources of synovia. By analogy to the findings in glomerulonephritis showing glycoprotein changes early in the disease, synovial biopsies would require similar testing. Isolation of terminal pure polysaccharides, then synthesis would provide probes for testing antigenicity in animals, both of normal and altered glycosyl residues,
351
REFERENCES 1.
Zvailfler NJ. Etiology and pathogenesis of rheumatoid arthritis. p. 417 in Arthritis and Allied Conditions. ed, McCarthy DJ. 9th ed, Lea and Ferbiger, Philadelphia, 1979.
2.
Arthritis
3.
McDevelt HO. compatibility ed. McCarthy
4.
idem. p. 143.
5.
Rabin BS. Immunopathology J, p. 39, Jan 1981.
6.
Cahill GF, Jr, McDevitt HO. Insulin dependent diabetes mellitus: the initial lesion. N Eng J Med 304:1454, 1981.
and rheumatism.
24th Rheumatism
Review.
24(2):133,
1981.
Genetic structure and function of the major histocomplex. p. 229 in Arthritis and Allied Conditions. DJ, 9th ed, Lea and Ferbiger, Philadelphia, 1979.
of insulin dependent
diabetes.
Ill Med
7.
Notkins AL.
8.
Nicolson GL. Transmembrane control of the receptors on normal and tumour cells. I cytoplasmic influence over cell surface components. Biochem Biophys Acta 457:57, 1976.
9.
Sweeley CC, Fung Y-K, Macher BA, Moskal JR and Nunez HA. Structure and metabolism of glycolipids. p. 46 in Glycoproteins and Glycolipids, ed. Walborg. Symposium 80, Am Chem Sot, 1980.
The cause of diabetes.
Sci Am 241:62, 1979.
10.
idem. Walborg EF, Jr. glycolipids, p. 8.
Current concepts of glycoproteins
11.
Watkins WM.
12.
Milat MS, Sinaij P. Chemical synthesis of human blood group B antigenic determinant: type 2 tetrasaccharide. Angew Chem Ed Engl 18:464, 1979.
13.
Feizi T, Wood E, Auge C, David S, Veyrieres A. Blood group I activities of synthetic oligosaccharides assessed by immunoImmunochemistry 15: 733, 1978. assay.
14.
Rodriguez-Iturbe B, Katizar VN, Coello J. Neuraminidase activity and free sialic acid levels in the serum of patients with acute posstreptococcal glomerulonephritis. N Eng J M 304:1506, 1981.
15.
Persellin RH, Weginton DAF, Rutstein JE, Cohen MJ, Goehrs HR, Steele D, Philips VK, Kradel RP. Pregnancy alpha-glycoprotein and rheumatoid arthritis; and Unger A, Kay A, Griffin AJ, Panayi GS. Disease activity and pregnancy-associated A2-glycoprotein in rheumatoid arthritis during pregnancy. Arthritis VIII Pan American Congress of Rheumatology. and Rheumatism. 24 (Suppl): 19, 1982 (Abstracts).
Blood group substances.
352
and
Science 152:172, 1966.