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Query: UMLS:C0011849 (diabetes)
 277,896 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Non-obese diabetic mice display a syndrome with dramatic clinical and pathological features similar to those of Type 1 (insulin-dependent) diabetes in man. Circulating autoantibodies to the surface of islet cells were demonstrated in some of these mice by a protein A radioligand assay. To produce monoclonal antibodies to islet cell surface antigens, therefore, we took the spleens of non-obese diabetic mice, transferred the spleen cells into non-immunized recipient mice, which were made immunologically incompetent by a large dose of X-irradiation, and then fused their lymphocytes with FO mouse myeloma cells. After screening the resultant hybrids, one stable hybridoma (3A4) that produced a monoclonal antibody (IgG1) specifically bound to the surface of islet cells was obtained. The purified monoclonal antibody was bound to the surface of transplantable Syrian golden hamster insulinoma cells sevenfold more than control antibody. Adsorption of the antibody on mouse spleen lymphocytes or thymocytes resulted in only a slight decrease in 125I-protein A binding to insulinoma cells. This antibody also reacted with the surface of mouse and rat islet cells, but not with that of rat spleen cells or hepatocytes. A spectrophotometric assay for peroxidase activity demonstrated that six times more peroxidase bound to insulinoma cells incubated with the antibody than to cells treated with control antibody. Furthermore, this antibody could be visually detected in the immunoenzymatic labelling of the surface of insulinoma cells. In summary, we have developed a novel method of producing monoclonal antibodies to the surface of islet cells for probing into the pathogenesis of Type 1 diabetes.
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PMID:Production of monoclonal antibodies to islet cell surface antigens using hybridization of spleen lymphocytes from non-obese diabetic mice. 637 47

A gas-liquid chromatographic method, using a fused silica capillary column, for the determination of red cell sorbitol is described. The capillary column gives complete resolution of the polyols xylitol, inositol, mannitol, sorbitol and galactitol, even when the glucose peak in the red cell chromatogram is dominating. The identity of sorbitol, and its single elution from the capillary column has been confirmed by mass spectrometry. Recovery of sorbitol from various red cell samples was 101% +/- 3.2 (mean +/- SD, n = 7). Precision, estimated from duplicate diabetic red cell sorbitol analyses was CVdup = 3.5% (n = 18) and from run to run analyses CVinterassay = 4.0% (n = 6). Sorbitol levels determined in erythrocytes of 19 healthy subjects were 5.9 +/- 1.6 nmol/ml red cells and in erythrocytes of 18 insulin-dependent diabetics 17.8 +/- 8.2 nmol/ml red cells (means +/- SD). The method described offers a reliable and specific tool to study in vivo polyol pathway activity in relation to some diabetes-associated complications.
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PMID:Determination of sorbitol in erythrocytes of diabetic and healthy subjects by capillary gas chromatography. 661 65

Capillary isoelectric focusing (CIEF) with electro-osmotic zone displacement of normal and pathological hemoglobins (Hb) is reported. CIEF is performed in untreated, open-tubular, fused-silica capillaries of 75 microns internal diameter using methylcellulose for dynamic column conditioning. After direct injection of hemolysates mixed with carrier ampholytes, high resolution separation of Hb variants, including Hb A1c, A, F, D, S, E and A2, is obtained, this permitting unambiguous characterization of Hb patterns of normal adults, newborns, patients with diabetes, different hemoglobinopathies and thalassemia syndromes. Qualitatively, the CIEF data compare well with those obtained by gel isoelectric focusing and high-performance liquid chromatography. CIEF is demonstrated to be a simple, rapid and fully instrumental approach to Hb analysis. Run times of less than 20 min make CIEF an attractive method for routine Hb investigations and screening programs.
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PMID:Application of dynamic capillary isoelectric focusing to the analysis of human hemoglobin variants. 751 28

The human insulin domains, signal peptide, B-chain, C-peptide, and A-chain, were highly expressed in Escherichia coli as recombinant proteins N-terminally fused to glutathione-S-transferase and a histidine-hexapeptide. The recombinant proteins were purified from insoluble cell fraction by affinity chromatography using metal chelating matrix, which was charged with Ni+2 ions. ELISA screening for autoantibodies directed to preproinsulin were performed with sera from patients with recently diagnosed insulin-dependent diabetes mellitus in order to localize the antigenic epitopes within the human preproinsulin. Of the patients, 14% had developed autoantibodies that recognized either the recombinant C-peptide or the signal peptide. No reaction was observed with the A-chain or B-chain.
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PMID:Mapping of antigenic epitopes within the recombinant human preproinsulin related to insulin-dependent diabetes mellitus. 751 60

We previously reported that 2400 base pairs (bp) of 5'-flanking DNA is sufficient for tissue-specific and hormonal/metabolic regulation of the human GLUT4 gene in transgenic mice (Liu, M.-L., Olson, A. L., Moye-Rowley, W. S., Buse, J. B., Bell, G. I., and Pessin, J. E. (1992) J. Biol. Chem. 267, 11673-11676). To further define the DNA sequences required for GLUT4 expression, we generated transgenic mice carrying 1975, 1639, 1154, 730, and 412 bp of the GLUT4 5'-flank (hG4) fused to the chloramphenicol acetyltransferase (CAT) reporter gene. The 1975-hG4-CAT, 1639-hG4-CAT, and 1154-hG4-CAT constructs were expressed in a tissue-specific manner identical to the endogenous murine GLUT4 mRNA. Regulation of these reporter gene constructs in insulin-deficient diabetes also paralleled the endogenous gene. In contrast, 730-hG4-CAT was expressed at high levels only in skeletal muscle and at low levels in all of the other tissues examined. Additionally, expression of 412-hG4-CAT was completely unrestricted. Neither the 730-hG4-CAT nor the 412-hG4-CAT reporter genes displayed any insulin-dependent regulation. These data demonstrate that a skeletal muscle-specific DNA element is located within 730 bp of the GLUT4 5'-flanking DNA but that 1154 bp is necessary to direct the full extent of tissue-specific and insulin-dependent regulation of the human GLUT4 gene in transgenic mice.
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PMID:Transcriptional regulation of the human GLUT4 gene promoter in diabetic transgenic mice. 755 12

The alpha 4 beta 1-integrin (CD49d, CD29) constitutively expressed on leukocytes regulates cell migration to inflammatory sites, cell activation, and development through its interactions with two alternate ligands, vascular cell adhesion molecule-1 (VCAM-1; CD106) expressed on cytokine-activated endothelium, dendritic and stromal cells, and the extracellular matrix protein fibronectin. Another alpha 4-integrin receptor, alpha 4 beta 7, expressed on leukocytes also binds VCAM-1 and fibronectin (FN), and controls homing to mucosal tissues through its interactions with mucosal vascular addressin MAdCAM-1. In vitro studies have shown that alpha 4-dependent cell adhesion is regulated by the activation state of the cell and by divalent cations. However, the existence and role of cells with different alpha 4 activation states in vivo have not been defined. Herein we show that a soluble ligand with the two N-terminal domains of human VCAM-1 fused to a human IgG1 constant region, VCAM-Ig, binds selectively to activated alpha 4-receptors on murine cells, such as those induced by Mn2+ in vitro. To determine whether the cells identified by VCAM-Ig were required under physiologic conditions, we assessed its anti-inflammatory effect. We show that VCAM-Ig is not bound to the majority of murine alpha 4+ cells after in vivo administration, yet it significantly delays the onset of adoptively transferred autoimmune diabetes. Thus, soluble VCAM-Ig can modify alpha 4-dependent disease progression, apparently by its selective action on cells with activated alpha 4-integrin receptors, thereby providing evidence for distinct alpha 4 activation states in vivo.
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PMID:Vascular cell adhesion molecule-Ig fusion protein selectively targets activated alpha 4-integrin receptors in vivo. Inhibition of autoimmune diabetes in an adoptive transfer model in nonobese diabetic mice. 760 69

Myoblasts from human skeletal muscle were isolated from needle biopsy samples of vastus lateralis and fused to differentiated multinucleated myotubes. Specific high-affinity insulin and insulin-like growth factor I (IGF-I) binding, glucose transporter proteins GLUT1 and GLUT4, glycogen synthase and pyruvate dehydrogenase proteins, and their specific mRNAs were identified in fused myotubes. Insulin and IGF-I stimulated 2-deoxyglucose uptake twofold with half-maximal stimulation by insulin at 0.98 +/- 0.12 nmol/l and maximal stimulation at 17.5 nmol/l. Acute insulin treatment (33 nmol/l) doubled glycogen synthase activity and glucose incorporation into glycogen while increasing pyruvate dehydrogenase approximately 30%. In cells cultured from NIDDM subjects, both basal (6.9 +/- 1.0 vs. 13.0 +/- 1.7 pmol.mg protein-1.min-1) and acute insulin-stimulated transport (13.5 +/- 2.0 vs. 22.4 +/- 1.3 pmol.mg protein-1.min-1) were significantly reduced compared with nondiabetic control subjects (both P < or = 0.005). GLUT1 protein content of total membranes from NIDDM subjects was decreased compared with control subjects, while GLUT4 levels were similar between groups. A significant correlation (r = 0.65, P < or = 0.05) was present when maximal rates of insulin-stimulated glucose transport in cell culture from subjects were compared with their corresponding in vivo glucose disposal determined by hyperinsulinemic glucose clamp. In summary, differentiated human skeletal muscle cultures exhibit biochemical and molecular features of insulin-stimulated glucose transport and intracellular enzyme activity comparable with the in vivo situation. Defective insulin-stimulated glucose transport persists in muscle cultures from NIDDM subjects and resembles the reduced insulin-mediated glucose uptake present in vivo. We conclude that this technique provides a relevant cellular model to study insulin action and glucose metabolism in normal subjects and determine the mechanisms of insulin resistance in NIDDM.
Diabetes 1995 Aug
PMID:Insulin action and glucose metabolism in nondiabetic control and NIDDM subjects. Comparison using human skeletal muscle cell cultures. 762

Evidence from in vivo studies indicates that the bile acid pool and bile acid excretion are increased in humans with diabetes mellitus and in experimental diabetic animals, and that both parameters return to normal levels after administration of insulin. To investigate the biochemical background of these changes, the effects of insulin on bile acid synthesis and cholesterol 7 alpha-hydroxylase and sterol 27-hydroxylase, two key enzymes in routing of cholesterol toward bile acids, were studied in cultured rat hepatocytes. Mass production of bile acids was dose dependently diminished, showing significant reduction (-33% to -53%) at physiological concentrations of the hormone (1.4 to 14 nmol/L) and a maximal decrease at 140 nmol/L (-65%). The decrease of bile acid synthesis correlated well with the suppression of cholesterol 7 alpha-hydroxylase and sterol 27-hydroxylase activity. The enzyme activity for cholesterol 7 alpha-hydroxylase, examined in more detail, was dose dependently diminished on incubation of hepatocytes with various concentrations of insulin, reaching maximal reduction at 14 nmol/L of insulin. Maximal decrease of the enzyme activity was seen after 8 hours of incubation (-70%). Insulin strongly reduced the rise in cholesterol 7 alpha-hydroxylase activity induced by incubation with dexamethasone. Sterol 27-hydroxylase activity was inhibited up to -58% after 24 hours of incubation with 140 nmol/L insulin. To study the mechanism of suppression of cholesterol 7 alpha-hydroxylase and sterol 27-hydroxylase activity, the effects of insulin on their respective levels of messenger RNA (mRNA) and gene transcription were assessed. The decrease in enzyme activities could be explained by a concomitant reduction in the cholesterol 7 alpha-hydroxylase (-76%) and sterol 27-hydroxylase (-62%) mRNA level. Transcriptional activity, as assessed by nuclear runoff assays, was decreased to the same extent, i.e., -60% for cholesterol 7 alpha-hydroxylase and -75% for sterol 27-hydroxylase. Transient expression experiments using a construct containing the proximal 348 basepairs of the cholesterol 7 alpha-hydroxylase promoter fused to the chloramphenicol acetyltransferase (CAT) gene (-348Rcat) showed a significant reduction of transcriptional activity (-64%) with insulin, indicating that a sequence important for an insulin-induced transcriptional response is located within the first 348 basepairs, preceding the transcription start of the cholesterol 7 alpha-hydroxylase promoter.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Insulin suppresses bile acid synthesis in cultured rat hepatocytes by down-regulation of cholesterol 7 alpha-hydroxylase and sterol 27-hydroxylase gene transcription. 784 24

To examine the hormonal/metabolic as well as tissue-specific expression of the GLUT4/muscle-fat facilitative glucose transporter gene, we have generated several transgenic mouse lines expressing a human GLUT4 mini-gene which extends 5.3 kilobases (kb) upstream of transcription start and terminates within exon 10. This construct (hGLUT4-11.5) was expressed in a tissue-specific pattern identical to the endogenous mouse GLUT4 gene. The transcription initiation sites of the transgenic construct were similar to the GLUT4 gene expressed in human tissues. To investigate the hormonal/metabolic-dependent regulation of GLUT4, the transgenic animals were made insulin-deficient by streptozotocin (STZ) treatment. In these animals, STZ-induced diabetes resulted in a parallel decrease in endogenous mouse GLUT4 mRNA and the transgenic human GLUT4 mRNA in white adipose tissue, brown adipose tissue, and cardiac muscle. Similarly, insulin treatment of the STZ-diabetic animals restored both the endogenous mouse and transgenic human GLUT4 mRNA levels. To further define cis-regulatory regions responsible for this hormonal/metabolic regulation, the same analysis was performed on transgenic animals which carry 2.4 kb of the human GLUT4 5'-flanking region fused to a CAT reporter gene (hGLUT4[2.4]-CAT). This reporter construct responded similarly to the human GLUT4 mini-gene demonstrating that the element(s) controlling hormonal/metabolic regulation and tissue specificity all reside exclusively within 2.4 kb of the transcriptional initiation site.
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PMID:Hormonal/metabolic regulation of the human GLUT4/muscle-fat facilitative glucose transporter gene in transgenic mice. 848 63

Nonenzymatic glycation of apolipoprotein B (apo B) is a post-secretory modification of low density lipoprotein (LDL) that affects its atherogenic potential and is implicated in the accelerated atherosclerosis associated with diabetes. To facilitate assessment of apo B glycation, we produced hybridomas secreting monoclonal antibodies specific for glycated apo B. SP 2/0 myeloma cells were fused with spleen cells from BALB/c mice immunized with purified apo B glycated non-reductively in vitro. Specificity of monoclonal antibodies secreted by the cloned cell line designated ES12 was demonstrated by immunoblotting and by direct ELISA, wherein the antibodies reacted with glycated epitopes residing in LDL but not in other plasma proteins, and did not react with nonglycated apo B or nonglycated LDL. Immunoblotting of human plasma with ES12 monoclonal antibody yielded an approx. 180,000 molecular weight component showing co-identity with apo B, indicating site specificity for glycated epitopes residing in apo B of the LDL complex and absence of reactivity with other nonenzymatically glycated plasma proteins. This reactivity of ES12 with the physiologic form of glycated apo B that occurs in vivo differs from properties of other antibodies raised against glycated lipoproteins, which recognized glycated residues only after reductive conversion to glucitol-lysine and which do not discriminate between different glycated proteins. In a competitive ELISA, mean concentration of glycated LDL, measured as apo B equivalents, in eight separate plasma samples was 19.7 +/- 1.9 micrograms/ml, representing 3.5 +/- 0.3% of total apo B. The ES12 monoclonal antibody allows specific determination of plasma glycated LDL concentrations, which may have diagnostic and pathogenetic importance.
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PMID:Immunologic detection and measurement of glycated apolipoprotein B with site specific monoclonal antibodies. 850 55


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