Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0011849 (diabetes)
 277,896 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The vitreous is a neural extracellular space separated from the blood-vascular compartment by the blood-retinal barrier. Study of the appearance of serum proteins in this space have been carried out in rats with streptozotocin-induced diabetes mellitus, a condition associated with barrier dysfunction. A vitreous sampling technique that avoids contamination with surrounding tissue was employed. In rats 1 month after administration of streptozotocin (fasting serum glucose > or = 375 mg/dl), significant increases in vitreous protein were observed in the absence of discernible eye pathology. Two-dimensional isoelectric focusing and SDS-polyacrylamide gel analysis of the soluble fraction demonstrated 85 polypeptides, 28 of whose electrophoretic positions coincided with positions of serum polypeptides. The remainder were unrelated to serum polypeptide loci. Overall patterns of soluble protein from the vitreous of streptozotocin-injected and normoglycemic-uninjected control animals were virtually identical. Results support a system for selective transfer for certain proteins into the extraneural vitreous space as suggested by Chen and Chen (6).
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PMID:Protein levels in the vitreous of rats with streptozotocin-induced diabetes mellitus. 842 Jun 39

We have examined, by western immunoblot analysis, the sera of 16 insulin-dependent diabetes mellitus patients (IDDM) for the presence of autoantibodies against proteins extracted from islet-cell enriched preparations of normal human pancreata. A novel putative autoantigen recognized by late stage IDDM patients sera was identified, and its amino acid sequence was partially determined. Islets of Langerhans were partially purified by a modified collagenase digestion procedure, and subsequent protein extracts were fractionated by one-dimensional or two-dimensional polyacrylamide gel electrophoresis (1-D or 2-D SDS-PAGE). Immunoblot analysis revealed a 30-kD species which was recognized by 4 of 16 IDDM patients sera, but none of 16 normal sera. The 30-kD protein, appeared as a single band on 1-D SDS-PAGE, but was resolved on 2-D gel electrophoresis as several distinct protein species with different isoelectric points (pI's), ranging from 7 to 9. The amino terminal sequence of one such species was partially determined by microsequencing, and the second through the fourteenth amino acids were found to be identical to the corresponding sequence in human chymotrypsinogen. The fifteenth through the eighteenth amino acids were different from the known chymotrypsinogen sequence. This region corresponds with the site that is cleaved to activate chymotrypsinogen. Based on the size and sequence homology, this antigen appears to be related to chymotrypsinogen. We conclude that this 30-kD species may be an autoantigen in some late stage IDDM patients.
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PMID:IDDM patients' sera recognize a novel 30-kD pancreatic autoantigen related to chymotrypsinogen. 850 58

The class II molecules of the diabetes-prone NOD mice, I-Ag7, showed very limited amounts of stable form when analyzed by SDS-PAGE. We included the analysis of spleen B cells and B lymphoma cells transfected with I-Ag7 genes. Early during bio-synthesis there was invariant chain binding to the alpha beta-chains. Examination of APCs from F1 mice (NOD x C57BL/6) indicated that the same APC expressed high levels of unstable I-Ag7 and normal amounts of stable class II molecules compared with the other haplotype (I-Ab). The half-life of I-Ag7-peptide complexes on the cell surface of APC was significantly shorter than that of other class II haplotypes. Direct biochemical demonstration of peptide interactions with I-Ag7 was difficult to demonstrate. In T cell assays, the immunogenic peptides, including the diabetogenic Ag, were rapidly lost when peptide-pulsed APCs were washed free of peptide. We hypothesize that the weak and unstable peptide-binding property of I-Ag7 molecules does not favor the elimination or inactivation of autoreactive T cells.
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PMID:The class II MHC I-Ag7 molecules from non-obese diabetic mice are poor peptide binders. 854 93

Plasmin-alpha2-antiplasmin complexes (PAP) are considered good markers of fibrinolytic activation in vivo. The presence of neoantigens in these complexes offers the possibility to develop specific immunoassays to determine PAP levels. We have developed a sensitive PAP purification method in vitro by adding urokinase to fresh plasma followed by affinity chromatography to lysine-sepharose and elution with epsilon-aminocaproic acid. This material, characterized by SDS-PAGE and Western blotting, was used to raise monoclonal antibodies (MoAbs). We describe a new enzyme linked immunosorbent assay (ELISA) to quantify PAP complexes in plasma. The assay follows the sandwich principle and is based on two MoAbs, CPL12 and CPL15, that bind to the modified alpha2-antiplasmin moiety and the plasmin moiety of the complex respectively. The calibration curve was constructed with definite concentrations of purified PAP. The lower limit of the assay is 75 ng/ml and the variation coefficients are 3.5% (intra-assay) and 10-6% (interassay). A mean value of 573.5+/-131.4 ng/ml was obtained from PAP concentration in a healthy population (n = 30). Significantly higher PAP levels were observed under diverse clinical conditions in which fibrinolysis is activated: clinical sepsis, acute myocardial infarction (AMI), malignancy, diabetes, pregnancy, elderly people and thrombolytic therapy. From our results we conclude that this ELISA is suitable to measure in vivo plasma PAP levels.
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PMID:Development and clinical application of a new ELISA assay to determine plasmin-alpha2-antiplasmin complexes in plasma. 861 97

Glutamate decarboxylase (GAD65) is a major autoantigen in insulin-dependent diabetes (IDDM) and the neurological disorder Stiff-Man-Syndrome (SMS). We derived a human monoclonal autoantibody (MICA 2) from peripheral blood of a patient newly diagnosed with IDDM, which reacted with GAD65 in Western blots. This indicated that a linear epitope is recognized by MICA 2. Using an epitope cDNA library we mapped the MICA 2 epitope to a contiguous stretch of 26 amino acids (506-531) in the C-terminus of GAD65. Neither blocking experiments with synthetic peptides nor analysis of overlapping decapeptides expressed as fusion proteins allowed us to further narrow down the epitope to the typical size of linear epitopes of 6-8 amino acids. We suggest that a miniconformational epitope provided by amino acids 506-531 is recognized by MICA 2, which withstands SDS gel electrophoresis without destruction or partially refolds during the Western blot procedure. A sequence homology with human heat shock protein 60 (HSP60) maps to this region of GAD65 but no cross-reactivity of MICA 2 with HSP60 occurred. Our data demonstrate that reactivity of an antibody in Western blots does not necessarily define a classic linear epitope of 6-8 amino acids and describe a new autoreactive epitope in GAD65 different from those reported for sera from patients with SMS.
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PMID:Mapping of an autoreactive epitope within glutamate decarboxylase using a diabetes-associated human monoclonal autoantibody and an epitope cDNA library. 874 89

We studied the quantitative and qualitative characteristics of lipoprotein(a) [Lp(a)] as a function of apolipoprotein(a) [apo(a)] phenotype in 87 members (42 males, 45 females) of 20 diabetic families, 26 of whom were diagnosed with non-insulin-dependent diabetes mellitus (NIDDM) with moderate glycaemic control (HbA1c 7.1 +/- 1.2%). Apo(a) phenotyping was performed by a sensitive, high-resolution technique using SDS-agarose/gradient PAGE (3-6%). To date, 26 different apo(a) phenotypes, including a null type, have been identified. Serum Lp(a) levels of NIDDM patients and non-diabetic members of the same family who had the same apo(a) phenotypes were compared, while case control subjects were chosen from high-Lp(a) non-diabetic and low-Lp(a) nondiabetic groups with the same apo(a) phenotypes in the same family. Serum Lp(a) levels were significantly higher in NIDDM patients than in non-diabetic subjects (39.8 +/- 33.3 vs 22.3 +/- 19.5 mg/dl, p < 0.05). The difference in the mean Lp(a) level between the diabetic and non-diabetic groups was significantly (p < 0.05) greater than that between the high-Lp(a) non-diabetic and low-Lp(a) non-diabetic groups. An analysis of covariance and a least square means comparison indicated that the regression line between serum Lp(a) levels [log Lp(a)] and apo(a) phenotypes in the diabetic patient group was significantly (p < 0.01) elevated for each apo(a) phenotype, compared to the regression line of the control group. These data together with our previous findings that serum Lp(a) levels are genetically controlled by apo(a) phenotypes, suggest that Lp(a) levels in diabetic patients are not regulated by smaller apo(a) isoforms, and that serum Lp(a) levels are greater in diabetic patients than in non-diabetic family members, even when they share the same apo(a) phenotypes.
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PMID:Serum lipoprotein(a) concentrations and apolipoprotein(a) phenotypes in the families of NIDDM patients. 878 17

Glutamic acid decarboxylase (GAD) is a major autoantigen in insulin-dependent diabetes mellitus (IDDM). This was initially identified as a 64-65 kD molecule according to migration in gels after immunoprecipitation from pancreatic islets. We studied the antigenicity of two different radiolabelled preparations of GAD, derived either by affinity purification from porcine brain and known to contain GAD 65 and GAD 67, or by expression from a cDNA for human GAD 65 by rabbit reticulocyte lysate (RRL). Radiolabelled immunoprecipitated pellets from the reaction of potent antisera to GAD from patients with IDDM were examined by autoradiography after SDS-PAGE under reducing or non-reducing conditions. Also, preparations of porcine brain GAD were "depleted' of GAD by exposure to antisera, and then similarly re-examined. Autoradiography of radiolabelled GAD either affinity purified from porcine brain, or expressed by RRL, showed that the immunoprecipitated protein migrated under non-reducing conditions according to a M(r) of approximately 110-130 kD, corresponding to dimeric forms of monomeric GAD of approximately 55-65 kD. Depletion by immunoprecipitation of this minor higher M(r) component from preparations of GAD left, in the supernatant, an abundance of GAD of M(r) 64-65 kD corresponding to monomer that was completely non-reactive with potent IDDM sera. We conclude that IDDM sera react with the GAD molecule in a dimeric (or oligomeric) form. Our findings have general connotations for self-tolerance and autoimmunity.
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PMID:Diabetic sera react with the glutamic acid decarboxylase molecule in a dimeric-oligomeric form. 891 80

A role for glycation in diabetic pathology appears beyond doubt and one of the present trends is to focus the poorly explored field of intracellular glycation. In this work we studied the pattern of early glycation in hepatocyte cytosolic proteins from streptozotocin-induced diabetic rats (n=14) compared to control animals (n=8). Glycated proteins were present in the cytosol of control rats and increased three-fold after one month of diabetes, while glycated Hb and glycated plasma proteins rose two- and three-fold, respectively. A good correlation (r=0.82, p<0.001) was found between glycated cytosolic proteins and glycated plasma proteins, suggesting the latter could provide an indirect indication of intracellular glycation. Using PBA affinity chromatography followed by SDS-PAGE we detected 7 major glycated bands in cytosols from control animals which increased dramatically in diabetic rats. Moreover, other glycated proteins, which were undetectable in control animals, became prominent, and more than 15 major bands can thus be resolved. No major differences in the patterns can be seen after 1, 5, or 12 months of diabetes, suggesting that early glycation in cytosolic proteins reaches an equilibrium in a short period of one to two weeks (further supported by the tight correlation with glycated plasma proteins). Through comparison of the patterns obtained with an antiglucytollysine antibody on Western blots with those of silver stained gels from the PBA eluates we present evidence that intracellular glycation is mediated by glucose but mainly by other sugars.
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PMID:Glycation of hepatocyte cytosolic proteins in streptozotocin-induced diabetic rats. 895 99

Our group previously reported an assay for the study of lymphocyte adhesion to insulin-producing cells in which xenogeneic rat insulinoma (RIN) cells were used as targets. The present study found an increased number of RIN-cytoadherent lymphocytes in 63 patients with Type 1 diabetes compared with 150 control subjects and in 211 NOD mice compared with 104 BALB/c mice (p < 0.001). Proteins concentrated from spontaneous RIN cell culture supernatants inhibited increased RIN-adhesion of NOD splenocytes or lymphocytes from diabetic patients (p < 0.001). In addition, increased RIN binding was dose-dependently abolished by RIN membrane extracts. The fact that RIN binding was inhibited by proteins from both membrane and the culture supernatant from RIN cells suggests that soluble inhibitory proteins were spontaneously released into the supernatant from a hydrophobic membrane-bound form. This tended to be confirmed since inhibition obtained with both preparations involved a 55-75 kDa HPLC protein fraction. The possibility that the membrane form of the inhibitory protein was anchored by a glycosylphosphatidylinositol (GPI) tail was evaluated. When RIN cells were treated with PI-PLC, their ability to bind lymphocytes from diabetic patients or NOD splenocytes decreased (p < 0.001) to control levels. Co-incubation with the 55-75 kDa fraction of proteins cleaved from RIN cells by PI-PLC also lowered the number of RIN-adherent NOD splenocytes to control levels. SDS-PAGE and IEF analyses of the 55-75 kDa inhibitory fraction from RIN cell supernatant revealed a major band with Mr 66 kDa and PI5.4, which may correspond to a protein with similar characteristics noted on 2-D electrophoresis of proteins cleaved from RIN cells by PI-PLC. Specific labelling of GPI moieties with 3H-ethanolamine, 3H-glucosamine, or 14C-glucosamine, as well as conversion of the hydrophobic Triton-X114 solubilised form into a hydrophilic form after PI-PLC treatment, confirmed the presence of a GPI anchor in this approximately 66 kDa RIN protein, which could thus be the molecule inhibiting adhesion in the system. Our data suggest that GPI-proteins from insulin-producing cells may influence the immune system both in their membrane-anchored and soluble forms. When considering the binding model, in which beta cells were tumoral and xenogeneic to diabetic lymphocytes, this potential influence of GPI-proteins suggests possible implications in situations of lymphocyte-beta cell interaction, i.e. anti-beta cell autoimmunity, immune reaction against insulinomas, and reaction against islet xenografts.
Diabetes Metab 1996 Dec
PMID:Proteins spontaneously released by rat insulinoma (RIN) cells are anchored on cell membrane by a glycosyl-phosphatidyl-inositol link and inhibit increased RIN cell adhesion of lymphocytes from type 1 diabetic patients and non-obese diabetic mice in vitro. 898 53

New cellular-based reagents are needed to diagnose type I diabetes as well as to monitor the outcomes of clinical trials at early time points. Four new monoclonal antibodies (mAbs) have been shown to demonstrate reduced binding to lymphocytes from identical twins with long-term type I diabetes relative to that observed with lymphocytes from their twin partners without diabetes or from control subjects. Biochemical analysis revealed mAb 3G12EG recognized an unidentified 45-kDa protein, whereas mAb 2E8F1 and 5B6E11 did not appear to precipitate specific proteins as detected by SDS-PAGE. Electrophoresis under reducing and nonreducing conditions and peptide mapping revealed that mAb 8F410 recognizes a novel dimeric form of HLA class I molecule. Predictions from crystallography studies suggested previously this class I dimer as the optimal activation of a single CD8 T-cell. In B-cells from both normal and diabetic individuals, the class I dimer was minimally associated with beta2-microglobulin rapidly formed in the endoplasmic reticulum. These new reagents appear to be able to identify new lymphocyte surface phenotypes associated with diabetes expression in both fresh blood samples and Epstein-Barr virus-established cell lines.
Diabetes 1997 Mar
PMID:New monoclonal antibody diagnostic reagents for type I diabetes: differential lymphocyte surface antigen expression related to disease. 903 90


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