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)

GM1-ganglioside receptor binding by the B subunit of cholera toxin (CtxB) is widely accepted to initiate toxin action by triggering uptake and delivery of the toxin A subunit into cells. More recently, GM1 binding by isolated CtxB, or the related B subunit of Escherichia coli heat-labile enterotoxin (EtxB), has been found to modulate leukocyte function, resulting in the down-regulation of proinflammatory immune responses that cause autoimmune disorders such as rheumatoid arthritis and diabetes. Here, we demonstrate that GM1 binding, contrary to expectation, is not sufficient to initiate toxin action. We report the engineering and crystallographic structure of a mutant cholera toxin, with a His to Ala substitution in the B subunit at position 57. Whereas the mutant retained pentameric stability and high affinity binding to GM1-ganglioside, it had lost its immunomodulatory activity and, when part of the holotoxin complex, exhibited ablated toxicity. The implications of these findings on the mode of action of cholera toxin are discussed.
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PMID:A mutant cholera toxin B subunit that binds GM1- ganglioside but lacks immunomodulatory or toxic activity. 1144 91

The present studies were undertaken to determine the levels of stimulatory and inhibitory guanine nucleotide regulatory proteins (Gs and Gi respectively) and their relationship with adenylyl cyclase activity in aorta from 5-day streptozotocin-induced diabetic (STZ) rats. The levels of Gi alpha-2 as determined by immunoblotting techniques using AS/7 antibody were significantly decreased by about 60% in STZ as compared to control rats, whereas the levels of Gs alpha were not altered. In addition, the stimulatory effect of cholera toxin (CT) on GTP-sensitive adenylyl cyclase was not different in STZ as compared to control rats. On the other hand, the stimulatory effects of GTPgammaS, isoproterenol, glucagon, forskolin (FSK) and sodium fluoride on adenylyl cyclase were enhanced in STZ-rats. Furthermore, GTPgammaS inhibited FSK-stimulated adenylyl cyclase activity in a concentration-dependent manner (receptor independent functions of Gi) in control rats which was almost completely abolished in STZ rats. In addition, receptor-mediated inhibition of adenylyl cyclase by angiotensin II (AII), oxotremorine and atrial natriuretic peptide (ANP) was attenuated in STZ rats. These results suggest that the decreased expression of Gi alpha, but not of Gs alpha, may be responsible for the observed altered responsiveness of adenylyl cyclase to hormonal stimulation and inhibition in STZ-rats. It may thus be suggested that the decreased Gi activity may be one of the possible mechanisms responsible for the impaired vascular functions in diabetes.
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PMID:Alterations in g-protein-linked signal transduction in vascular smooth muscle in diabetes. 1190 Mar 77

The pentameric B-subunit of cholera toxin (CTB) can be used as an efficient mucosal carrier of either immunogenic or tolerogenic T-cell epitopes. In this study a series of fusions was constructed between the genes encoding CTB and the B-chain of human insulin (InsB). The resulting fusion proteins were expressed in Escherichia coli and isolated as cytoplasmic inclusion bodies that were then dissolved and assembled in vitro. GM1 enzyme-linked immunosorbent assay (ELISA), sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analyses showed that the protein construct in which InsB was fused to the C-terminus of a CTB monomer (CI) assembled into structures that both bound to the receptor GM1 ganglioside and reacted with monoclonal antibodies to CTB and insulin. Fusion of InsB to the N-terminus of CTB resulted in protein that could not assemble into pentameric CTB. In vitro assays showed that the CI fusion protein was 300-fold more potent than native insulin at inducing interleukin-2 (IL-2) production by an insulin-specific T-cell hybridoma. When administered orally, the CI fusion protein induced efficient immunological suppression of ovalbumin-specific T-cell responses in mice co-immunized parenterally with insulin and ovalbumin. These results demonstrate the stability, GM1 receptor-binding activity and antigenic authenticity of the CI fusion protein as well as its ability to elicit insulin-specific T-cell responses in vitro. In addition, we demonstrate that the CI fusion protein induces efficient immunosuppression after oral administration, raising the possibility of using such constructs in the treatment of type-1 diabetes.
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PMID:Genetic fusion of human insulin B-chain to the B-subunit of cholera toxin enhances in vitro antigen presentation and induction of bystander suppression in vivo. 1204 53

Nasal administration of beta cell-derived auto-antigens has been reported to suppress the development of autoimmune diabetes. We investigated the tolerogenic effects of insulin conjugated to the B subunit of cholera toxin (CTB). Nasal administration of 1 micro g of CTB-insulin significantly delayed the incidence of diabetes in comparison to CTB treated mice. However, administration of 4 or 8 micro g of the conjugate had no protective effect. Protection induced by CTB-insulin was transferred to naive recipients by splenic CD4+ T cells. This result favours an active cellular mechanism of regulation, which was lost using higher (4-8 micro g) or lower (0.1-0.5 micro g) amounts of the conjugate. When co-administered with diabetogenic T cells, splenic T cells from CTB-insulin-treated mice reduced the lymphocytic infiltration of the islets. Reverse transcription-polymerase chain reaction analysis of recipients' pancreatic glands revealed an increase of TGF-beta and IL-10 transcripts after donor mice tolerization, while levels of IFN-gamma and IL-4 RNAs were unchanged. We observed a significant increase of T cell proliferation after unspecific stimulation in the spleen and pancreatic lymph nodes 24 h after CTB-insulin administration in -comparison to control treatment. Higher amounts of IL-4 and IFN-gamma were noticed in pancreatic lymph nodes of tolerized mice upon in vitro stimulation. Antigen-specific unresponsiveness after immunization and upon subsequent in vitro exposure to homologous antigen was obtained in nasally treated animals. Our results underlined the importance of nasal mucosa as an inducing site of tolerance and provided evidence for similar mechanisms of action to what has been described for the oral route, which favoured a CTB-insulin specific effect.
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PMID:Nasal administration of CTB-insulin induces active tolerance against autoimmune diabetes in non-obese diabetic (NOD) mice. 1239 Mar 7

Oral administration of insulin conjugated to the B chain of cholera toxin (CTB-insulin) in non-obese diabetic (NOD) mice results in diabetes prevention. We investigated the respective contributions of L-selectin (CD62L) and alpha4-integrin pathways during CTB-driven tolerance. Purified CD62L+CD4+ cells from CTB-insulin fed mice significantly reduced the capacity of diabetogenic T cells to transfer diabetes in syngeneic recipients. In vivo antibody blockade of fed animals during adoptive co-transfer experiments indicated that both CD62L and alpha4-integrins pathways were necessary to develop a protective response after oral tolerance induction. In contrast, when antibodies were given to recipient mice, only CD62L was critical for the protection. In vitro stimulated CD62L+CD4+ cells from the spleen of fed animals secreted lower amounts of IL-4 and IL-10 but comparable levels of TGFbeta than CD62L-cells. A reduced IFN-gamma production between the two cell subsets was specifically observed in CTB-insulin fed mice. Furthermore, antibody treatments induced changes in T-cell migration to the spleen, mesenteric and pancreatic lymph nodes. The protective effect was also associated with migration of regulatory T cells into pancreatic islets. Taken together, our results suggest that L-selectin and alpha4-integrin have distinct but complementary roles in the generation and function of regulatory CD4+ T cells following CTB-insulin administration.
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PMID:alpha4 integrins and L-selectin differently orchestrate T-cell activity during diabetes prevention following oral administration of CTB-insulin. 1247 43

The present studies were undertaken to examine if the impaired vascular function observed in diabetes is attributed to the altered levels of G-protein. Diabetes was induced in Sprague Dawley rats by a single intraperitoneal injection of streptozotocin (STZ) (60 mg/kg body wt) and after a period of 5 days, the aorta were used for adenylyl cyclase activity determination and protein quantification. A temporal relationship between the expression of Gialpha proteins and development of diabetes was also examined on day 1, 2, 3, 4 and 5 of injection of STZ. Blood glucose levels were significantly increased from day 1 in STZ-rats as compared to their counterpart control rats and reached to about 20 mM on 3rd day and 30 mM on 5th day. The expression of Gialpha-2 and Gialpha-3 proteins as determined by immunoblotting techniques was decreased by about 70 and 50% respectively in aorta from STZ rats compared to the control rats after 5 days of treatment, whereas 40% decrease in Gialpha-2 and Gialpha-3 was observed after 3rd day of STZ injection. On the other hand, the expression of Gsalpha was unaltered in STZ rats. In addition, the stimulatory effect of cholera toxin (CT) on GTP-mediated stimulation of adenylyl cyclase was not different in STZ as compared to the control group. However, the stimulatory effects of isoproterenol, glucagon, NaF and FSK on adenylyl cyclase activity were significantly enhanced in STZ rats as compared to control rats, whereas basal adenylyl cyclase activity was significantly lower in STZ-rats as compared to control rats. In addition, GTPgammaS inhibited FSK-stimulated adenylyl cyclase activity in concentration-dependent manner (receptor-independent functions of Gialpha) in control rats which was completely attenuated in STZ-rats. In addition, receptor-mediated inhibitions of adenylyl cyclase by angiotensin II, oxotremorine, atrial natriuretic peptide (ANP99-126) and C-ANP4-23 were also attenuated (receptor-dependent functions of Gialpha) in STZ-rats. These results indicate that aorta from diabetic rats exhibit decreased levels of cAMP and decreased expression of Gialpha. The decreased expression of Gialpha may be responsible for the altered responsiveness of adenylyl cyclase to hormonal stimulation and inhibition in STZ-rats. It may thus be suggested that the impaired adenylyl cyclase-Gialpha protein signaling may be one of the possible mechanisms responsible for the impaired vascular functions in diabetes.
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PMID:Streptozotocin-induced diabetes impairs G-protein linked signal transduction in vascular smooth muscle. 1248 72

Interleukin (IL)-6 has recently been shown to be an adipocyte-expressed cytokine. Its serum concentrations are elevated in insulin resistance and obesity. For further evaluation of IL-6 gene expression regulation, fully differentiated 3T3-L1 adipocytes were treated with various hormones known to induce insulin resistance. IL-6 mRNA content was measured by quantitative real-time reverse transcription-polymerase chain reaction. Interestingly, treatment of adipocytes with 100 nM insulin, 10 micro M isoproterenol, 10 ng/ml tumour necrosis factor alpha (TNFalpha), and 500 ng/ml growth hormone (GH) for 16 h stimulated IL-6 mRNA expression 2.3-fold, 47-fold, 74-fold, and 1.4-fold, respectively (p < 0.01). In contrast, treatment with 100 nM dexamethasone significantly decreased IL-6 expression to 32 % of control levels (p < 0.01), whereas triiodothyronine and angiotensin 2 did not have any effect. Furthermore, stimulation of IL-6 expression was time-dependent with maximal stimulatory effects detectable after 1 h of insulin, isoproterenol, and GH addition and 12 h of TNFalpha, respectively. Moreover, isoproterenol's effect could be almost completely reversed by pretreatment of 3T3-L1 cells with the beta-adrenergic antagonist propranolol and mimicked by stimulation of G S -proteins with cholera toxin and adenylyl cyclase with forskolin and dibutyryl cAMP, respectively. Finally, IL-6 strongly induced its own expression in a time-dependent fashion. Taken together, our results demonstrate that IL-6 expression in adipocytes is governed by an autocrine positive feedback loop and upregulated by insulin, isoproterenol, TNFalpha, and GH. In concert with this adipocytokine's upregulation in states of decreased insulin sensitivity such as obesity and diabetes, the data support a possible role of IL-6 as a selectively regulated mediator of insulin resistance.
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PMID:Interleukin (IL)-6 mRNA expression is stimulated by insulin, isoproterenol, tumour necrosis factor alpha, growth hormone, and IL-6 in 3T3-L1 adipocytes. 1273 74

When conjugated to various proteins, the nontoxic B-chain of cholera toxin (CTB) significantly increases the ability of these proteins to induce immunological tolerance after oral administration. Here, we investigated if a nonconjugated form of CTB enhances the induction of immune tolerance after oral insulin administration. Induction of immunological tolerance was studied after oral administration of insulin preparations in three mouse models; an insulin/ovalbumin coimmunization model, a model of virus-induced diabetes in transgenic RIP-LCMV-NP mice and in nonobese diabetic (NOD) mice serving as a model of spontaneous diabetes. In the immunization model, we demonstrate that mixing with CTB increases the tolerogenic potential of insulin, approximately 10 fold. Titration of the CTB concentration in this system revealed that an insulin : CTB ratio of 100 : 1 was optimal for the induction of bystander suppression. Further studies revealed that this insulin : CTB ratio also was optimal for the prevention of diabetes in a virus-induced, transgenic diabetes model. In addition, the administration of this optimal insulin-CTB preparation significantly prevented the onset of diabetes in old NOD mice with established islet infiltration. The data presented here demonstrate that CTB, even in its unconjugated form, functions as a mucosal adjuvant, increasing the specific tolerogenic effect of oral insulin.
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PMID:The cholera toxin B subunit is a mucosal adjuvant for oral tolerance induction in type 1 diabetes. 1275 99

Our earlier investigations have demonstrated a critical difference in the efficacy of orally administered porcine compared to human or mouse insulin (no effect) in preventing type I diabetes in two distinct experimental models. Based on these findings one has to assume that certain insulins might not be suitable for the induction of oral 'tolerance'/bystander suppression, which might be one cause for recent failures in human oral antigen trials. Here we demonstrate that coupling to the non-toxic subunit of cholera toxin (CTB) can abolish these differences in efficacy between human and porcine insulin. As expected, an added benefit was the much smaller oral antigen dose required to induce CD4+ insulin-B specific regulatory cells that bystander-suppress autoaggressive responses. Mechanistically we found that uptake or transport of insulin-CTB conjugates in the gut occurs at least partially via binding to GM-1, which would explain the enhanced clinical efficacy. Both B chains bound well to major histocompatibility complex (MHC) class II, indicating comparable immunological potential once uptake and processing has occurred. Thus, our findings delineate a pathway to overcome issues in oral antigen choice for prevention of type I diabetes.
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PMID:Coupling of oral human or porcine insulin to the B subunit of cholera toxin (CTB) overcomes critical antigenic differences for prevention of type I diabetes. 1297 52

The skin is both an essential barrier for host defense and an important organ of immunity. In this study, we show that the application of cholera toxin to intact mouse skin induces and enhances autoimmune diseases affecting organs at distant anatomic sites, whereas its administration by the mucosal route has been reported to have the opposite effect. First, the CNS autoantigen myelin oligodendrocyte glycoprotein 35-55, when applied repeatedly with cholera toxin to the intact skin of healthy C57BL/6 mice, induced relapsing paralysis with demyelinating immunopathologic features similar to multiple sclerosis. Second, the application of cholera toxin in the absence of autoantigen exacerbated the severity of conventional experimental autoimmune encephalomyelitis induced by myelin oligodendrocyte glycoprotein in CFA. Third, the application of cholera toxin to the intact skin of NOD/Lt mice, with or without insulin B peptide 9-23, exacerbated insulitis and T lymphocyte-derived IFN-gamma and IL-4 production in the islets of Langerhans, resulting in an increased incidence and rate of onset of autoimmune diabetes. The data presented in this study highlight the different outcomes of adjuvant administration by different routes. Because dermal application of cholera toxin, and other bacterial products with similar adjuvant activities, is being developed as a clinical vaccination strategy, these data raise the possibility that it could precipitate autoimmune disease in genetically susceptible humans.
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PMID:Dermal enhancement: bacterial products on intact skin induce and augment organ-specific autoimmune disease. 1468 38


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