Binding of antibodies in sera from Type 1 (insulin-dependent) diabetic patients to glutamate decarboxylase from rat tissues. Evidence for antigenic and non-antigenic forms of the enzyme

An islet protein of Mr 64000, identified as the ?-amino butyric acid (GABA)-synthesizing enzyme, glutamate decarboxylase, is a major target for antibodies in Type 1 (insulin-dependent) diabetes mellitus. This enzyme is also expressed in brain and in some other tissues and may exist in multiple forms. The aim of this study was to determine the ability of antibodies from diabetic patients to recognize glutamate decarboxylase from rat islets, brain and other normal rat tissues. Glutamate decarboxylase was detected at high activity levels in brain and at lower levels in islets, kidney, liver, pituitary gland, thyroid gland, adrenal gland, testis and ovary. The ability of antibodies in sera of diabetic patients to immunoprecipitate enzyme activity from detergent extracts of tissues was determined. Antibodies in sera from diabetic patients were found to bind the enzyme from islet and brain extracts, but bound less than 20% of the activity from other tissues. The ability of antibodies to immunoprecipitate the brain enzyme was significantly correlated with the presence of antibodies to the islet 64 kilodalton antigen. These studies show that the glutamate decarboxylase activity expressed in brain shares antigenic determinants with the islet 64 kilodalton antigen. Isoforms of the enzyme expressed in other nonneuronal tissues may be antigenically distinct and may lack determinants recognized by diabetes-associated antibodies.</p

Binding of antibodies in sera from Type 1 (insulin-dependent) diabetic patients to glutamate decarboxylase from rat tissues. Evidence for antigenic and non-antigenic forms of the enzyme

An islet protein of Mr 64000, identified as the ?-amino butyric acid (GABA)-synthesizing enzyme, glutamate decarboxylase, is a major target for antibodies in Type 1 (insulin-dependent) diabetes mellitus. This enzyme is also expressed in brain and in some other tissues and may exist in multiple forms. The aim of this study was to determine the ability of antibodies from diabetic patients to recognize glutamate decarboxylase from rat islets, brain and other normal rat tissues. Glutamate decarboxylase was detected at high activity levels in brain and at lower levels in islets, kidney, liver, pituitary gland, thyroid gland, adrenal gland, testis and ovary. The ability of antibodies in sera of diabetic patients to immunoprecipitate enzyme activity from detergent extracts of tissues was determined. Antibodies in sera from diabetic patients were found to bind the enzyme from islet and brain extracts, but bound less than 20% of the activity from other tissues. The ability of antibodies to immunoprecipitate the brain enzyme was significantly correlated with the presence of antibodies to the islet 64 kilodalton antigen. These studies show that the glutamate decarboxylase activity expressed in brain shares antigenic determinants with the islet 64 kilodalton antigen. Isoforms of the enzyme expressed in other nonneuronal tissues may be antigenically distinct and may lack determinants recognized by diabetes-associated antibodies.</p

Detection of pancreatic islet 64,000 Mr autoantigens in insulin-dependent diabetes distinct from glutamate decarboxylase

Patients with insulin-dependent diabetes (IDDM) possess antibodies to islet proteins of Mr-64,000. Potential autoantigens of this Mr include glutamate decarboxylase (GAD) and 65 kD heat shock protein. We have detected two distinct antibody specificities in IDDM that bind 50,000 Mr or 37,000/40,000 Mr proteolytic fragments of 64,000 Mr proteins. In this study, we investigated relationships of these proteolytic fragments to GAD and heat shock proteins. Polyclonal antibodies to GAD bound 50,000 Mr fragments of islet antigen. Recombinant GAD65, but not GAD67, blocked binding to this antigen, suggesting that 50,000 Mr fragments are derived from islet GAD65. In contrast, GAD antibodies did not recognize 37,000/ 40,000 Mr fragments, and neither GAD isoforms blocked autoantibody binding to precursors of these fragments. The 37,000/40,000 Mr fragments, but not the 50,000 Mr fragments, were detected after trypsin treatment of immunoprecipitates from insulinoma cells that lacked expression of major GAD isoforms. Antibodies in IDDM did not bind native or trypsinized islet heat shock proteins. Thus, IDDM patients possess antibodies to GAD, but also distinct antibodies to a 64,000 Mr protein that is not related to known GAD isoforms or heat shock proteins.</p

Detection of pancreatic islet 64,000 Mr autoantigens in insulin-dependent diabetes distinct from glutamate decarboxylase

Patients with insulin-dependent diabetes (IDDM) possess antibodies to islet proteins of Mr-64,000. Potential autoantigens of this Mr include glutamate decarboxylase (GAD) and 65 kD heat shock protein. We have detected two distinct antibody specificities in IDDM that bind 50,000 Mr or 37,000/40,000 Mr proteolytic fragments of 64,000 Mr proteins. In this study, we investigated relationships of these proteolytic fragments to GAD and heat shock proteins. Polyclonal antibodies to GAD bound 50,000 Mr fragments of islet antigen. Recombinant GAD65, but not GAD67, blocked binding to this antigen, suggesting that 50,000 Mr fragments are derived from islet GAD65. In contrast, GAD antibodies did not recognize 37,000/ 40,000 Mr fragments, and neither GAD isoforms blocked autoantibody binding to precursors of these fragments. The 37,000/40,000 Mr fragments, but not the 50,000 Mr fragments, were detected after trypsin treatment of immunoprecipitates from insulinoma cells that lacked expression of major GAD isoforms. Antibodies in IDDM did not bind native or trypsinized islet heat shock proteins. Thus, IDDM patients possess antibodies to GAD, but also distinct antibodies to a 64,000 Mr protein that is not related to known GAD isoforms or heat shock proteins.</p

Antibodies to islet 37k antigen, but not to glutamate decarboxylase, discriminate rapid progression to IDDM in endocrine autoimmunity

Apart from islet cell antibodies (ICAs), antibodies to glutamate decarboxylase (GAD), insulin autoantibodies (IAAs), and a novel islet antigen (37k antigen) are potential markers for insulin-dependent diabetes mellitus (IDDM). GAD is also an antigen in stiff-man syndrome (SMS), and both SMS and IDDM are associated with ICAs and autoimmunity to other endocrine organs. We investigated possible links between antibody responses to islet antigens with autoimmunity to other endocrine organs and determined which specific antibodies can identify individuals who progress to IDDM. Antibodies to GAD were detected in â?¥90 of both diabetic and nondiabetic patients with ICAs and other endocrine autoimmunity, in 59 of ICA-positive IDDM patients without endocrine autoimmunity, in all patients with SMS, but in only 1-3 of healthy (nondiabetic) and autoimmune disease control subjects. GAD antibody levels were increased in ICA-positive IDDM patients with polyendocrine autoimmunity compared with those without. In contrast, antibodies to 37k antigen were only detected in patients who developed acute-onset IDDM. IAAs were also associated with IDDM. Thus, certain factors enhance antibody responses to GAD in polyendocrine autoimmunity, but this does not necessarily lead to development of IDDM or SMS. Antibodies to 37k antigen are strongly associated with acute-onset IDDM and are useful serological markers for disease.</p

Antibodies to islet 37k antigen, but not to glutamate decarboxylase, discriminate rapid progression to IDDM in endocrine autoimmunity

Apart from islet cell antibodies (ICAs), antibodies to glutamate decarboxylase (GAD), insulin autoantibodies (IAAs), and a novel islet antigen (37k antigen) are potential markers for insulin-dependent diabetes mellitus (IDDM). GAD is also an antigen in stiff-man syndrome (SMS), and both SMS and IDDM are associated with ICAs and autoimmunity to other endocrine organs. We investigated possible links between antibody responses to islet antigens with autoimmunity to other endocrine organs and determined which specific antibodies can identify individuals who progress to IDDM. Antibodies to GAD were detected in â?¥90 of both diabetic and nondiabetic patients with ICAs and other endocrine autoimmunity, in 59 of ICA-positive IDDM patients without endocrine autoimmunity, in all patients with SMS, but in only 1-3 of healthy (nondiabetic) and autoimmune disease control subjects. GAD antibody levels were increased in ICA-positive IDDM patients with polyendocrine autoimmunity compared with those without. In contrast, antibodies to 37k antigen were only detected in patients who developed acute-onset IDDM. IAAs were also associated with IDDM. Thus, certain factors enhance antibody responses to GAD in polyendocrine autoimmunity, but this does not necessarily lead to development of IDDM or SMS. Antibodies to 37k antigen are strongly associated with acute-onset IDDM and are useful serological markers for disease.</p

Antibodies to GAD and tryptic fragments of islet 64K antigen as distinct markers for development of IDDM: studies with identical twins

Insulin-dependent diabetes mellitus (IDDM) is associated with antibodies to a 64,000-M(r) islet cell protein, at least part of which is identified as glutamic acid decarboxylase (GAD). These antibodies are detected as two distinct antibody specificities to 50,000-M(r) and 37,000/40,000-M(r) tryptic fragments of the autoantigen (50K and 37K antibodies, respectively). We determined the frequencies of antibodies to intact GAD, tryptic fragments of islet 64,000-M(r) antigen, islet cell antibodies (ICAs), and insulin autoantibodies (IAAs) in sera from 58 nondiabetic identical twins of patients with IDDM, of whom 12 subsequently developed diabetes. ICA, antibodies to intact GAD, and those to tryptic fragments were detected at similar frequencies in prediabetic twins (67-75), but only 25 had IAA. Of 46 twins who remain nondiabetic, GAD antibodies, 50K antibodies, and ICA were detected in 6 (13), 7 (15), and 5 (11), respectively, whereas only 1 (2) possessed 37K antibodies and 2 (4) had IAA. Eight of 9 twins with 37K antibodies and all 6 twins with ICA > 20 Juvenile Diabetes Foundation U have developed diabetes. Antibodies to GAD are sensitive markers for diabetes development but may also be present in genetically susceptible individuals who are unlikely to develop disease. Antibodies to 37,000/40,000-M(r) fragments of the 64,000-M(r) antigen or high-titer ICA were the best markers for diabetes development in these twins.</p

Antibodies to GAD and tryptic fragments of islet 64K antigen as distinct markers for development of IDDM: studies with identical twins

Insulin-dependent diabetes mellitus (IDDM) is associated with antibodies to a 64,000-M(r) islet cell protein, at least part of which is identified as glutamic acid decarboxylase (GAD). These antibodies are detected as two distinct antibody specificities to 50,000-M(r) and 37,000/40,000-M(r) tryptic fragments of the autoantigen (50K and 37K antibodies, respectively). We determined the frequencies of antibodies to intact GAD, tryptic fragments of islet 64,000-M(r) antigen, islet cell antibodies (ICAs), and insulin autoantibodies (IAAs) in sera from 58 nondiabetic identical twins of patients with IDDM, of whom 12 subsequently developed diabetes. ICA, antibodies to intact GAD, and those to tryptic fragments were detected at similar frequencies in prediabetic twins (67-75), but only 25 had IAA. Of 46 twins who remain nondiabetic, GAD antibodies, 50K antibodies, and ICA were detected in 6 (13), 7 (15), and 5 (11), respectively, whereas only 1 (2) possessed 37K antibodies and 2 (4) had IAA. Eight of 9 twins with 37K antibodies and all 6 twins with ICA > 20 Juvenile Diabetes Foundation U have developed diabetes. Antibodies to GAD are sensitive markers for diabetes development but may also be present in genetically susceptible individuals who are unlikely to develop disease. Antibodies to 37,000/40,000-M(r) fragments of the 64,000-M(r) antigen or high-titer ICA were the best markers for diabetes development in these twins.</p