EPIDEMIOLOGY AND IMMUNOGENETICS OF INSULIN-DEPENDENT DIABETES MELLITUS IN VENEZUELAN CHILDREN (original) (raw)
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Virologic, immunologic, and geneticfactors in insulin-dependent diabetes mellitus
The Journal of Pediatrics, 1983
A 16-month-old girl presented with an episode of fever and acute thrombocytopenic purpura caused by a Coxsackie B5 virus. On days 13 to 23, laboratory evidence of diabetes mellitus was present, followed by a 2 1/2-month remission, then by definitive insulin-dependent diabetes. The involvement of virologic, immunologic, and genetic factors in the pathophysiology was substantiated by the following data: (1) Virus-induced glucose intolerance was produced in selected mouse strains. (2) Islet-cell antibodies were found one week before onset of diabetes; however, circulating lymphocytes of the child at that time suppressed insulin release from islets in vitro. (3) Immunogenetic analysis of the child revealed the presence of high-risk genetic markers. It is suggested that the convergence of an insulotropic variant virus, genetic predisposition, and perhaps some uncontrolled adjuvant factors, e.g. steroid therapy and DPT vaccination, may have determined insular damage and anti-islet autoimmune reactions, leading to insulin-dependent diabetes mellitus.
Diabetes, 2006
C57BL/6 (B6) mice develop glucose intolerance with age, whereas C3H/He (C3H) mice do not. In this study, we examined whether this differential glucose homeostasis was associated with differences of proteolytic activation of pancreatic prohormones. Radioimmunoassays showed comparable levels of fasting plasma insulin between the two strains but a significantly lower glucagon level in B6 mice. Pulse-chase analysis of glucagon biosynthesis in isolated pancreatic islets revealed that proglucagon was less efficiently processed in B6 mice. Because proprotein convertase (PC)2 and its 7B2 helper protein are required for this processing, we quantified islet mRNA levels by RT-PCR and protein levels by immunoblotting. The levels of proPC2 mRNA were similar between the two strains, but B6 protein extracts contained less of the mature PC2. In contrast, 7B2 mRNA and protein levels were both significantly lower in B6 pancreas. Sequencing of the 7B2 gene promoter and cDNA in the two strains revealed seven single nucleotide polymorphisms and one dinucleotide insertion/deletion in the cDNA as well as a single nucleotide polymorphism and two insertions/deletions in the promoter. Differential expression of 7B2 may contribute to the difference between B6 and C3H mice not only in glucagon production and secretion but also in glucose tolerance. Diabetes 55:452-459, 2006 RESEARCH DESIGN AND METHODS Mice, antibodies, and PCR primers. C57BL/6J and C3H/HeJ (8 -12 weeks old) were purchased from The Jackson Laboratory (Bar Harbor, ME). They were housed in a pathogen-free facility, on a 12-h light/dark cycle, with free access to food and water. Experimental manipulations of these animals were approved by the animal care committee of the Ottawa Health Research Institute and performed in accordance with the guidelines of the Canadian Council for Animal Care. The antibodies and the PCR primers used in this study are described in Tables 1 and 2, respectively. Pancreatic islet isolation and culture. Islets of Langerhans were isolated from mouse pancreata by injecting a collagenase buffer (collagenase type XI, 1.5 mg/ml; Sigma-Aldrich, Oakville, ON, Canada) into the pancreatic duct as previously described (20). Briefly, mice were anesthetized and killed by cervical dislocation. Pancreata were digested by injection of a collagenase solution, and islets were handpicked under a microscope. They were either flash frozen for later extraction of RNA and proteins or cultured for 40 h at ϳ100 islets per 1 ml of RPMI-1640 medium supplemented with 10% (vol/vol) fetal bovine serum at 37°C in a 95% air/5% CO 2 humidified atmosphere. Half of the medium was changed 24 h later. For pulse-chase studies, each islet cohort was transferred into 0.5 ml serum-free RPMI-1640 containing 2.6 mmol/l glucose for 30 min to reduce the intracellular content of methionine and then transferred into 0.2 ml of the same medium containing 250 Ci 35 S-methionine and either 2.6 or 16.7 mmol/l glucose. After a 20-min incubation (pulse), the radioactive medium was replaced with fresh, nonradioactive, complete medium and incubation was resumed for 180 min (chase). Spent media and islets were collected at the two time points and kept frozen at Ϫ80°C until analysis.
Molecular Endocrinology, 1987
The structure of the canine prepropancreatic polypeptide (preproPP) cDNA was determined. The nucleotide sequence conservation between human and canine preproPP is very high for the signal peptide (82%) and the region coding for the 36 amino acid pancreatic polypeptide (PP) (92%). The overall sequence homology for the C-terminal portion of proPP containing the icosapeptide and a Cterminal extension peptide is only 63% whereas the 3'-untranslated regions of human and canine PP mRNA share 73% homology after alignment for maximal homology. The only sequence conservation in icosapeptide is the region coding for the last 10 amino acids of the icosapeptide. Comparison of PP immunoactivity and PP mRNA concentrations in extracts of the developmentally distinct uncinate process and splenic lobes of the canine pancreas revealed the same ratio of mRNA concentrations (16 ± 6.5) and PP peptide concentrations (18 ± 7.0) in the uncinate process compared to the splenic lobe (n = 6). However, a similar comparison of insulin C-peptide (CP) immunoactivity and insulin mRNA concentration revealed a smaller ratio of CP immunoactivity (0.37 ± 0.05) than insulin mRNA (0.58 ± 0.10) between the same lobes (P < 0.0074, n = 6). This increased steady state CP concentration relative to insulin mRNA in splenic lobe compared to the uncinate process was not observed for PP peptide and mRNA. (Molecular Endocrinology 1: 413-419, 1987)
Diabetes, 1999
S.M. and G.L. contributed equally to this work. C M V, cytomegalovirus; HPLC, high-performance liquid chromatography; IEF-1, insulin enhancer factor 1; IRI, immunoreactive insulin; KRBH, Krebs-Ringer bicarbonate buffer, pH 7.4; PDX-1, pancreatic duodenal homeobox 1; PLP, proinsulin-like peptides; RIA, radioimmunoassay; RIPE3b1-Act, rat insulin promoter element 3b1 activator; STF-1, somatostatin transcription factor 1.
Identification of Tissue-Restricted Transcripts in Human Islets
Endocrinology, 2004
The purpose of our study was to identify transcripts specific for tissue-restricted, membrane-associated proteins in human islets that, in turn, might serve as markers of healthy or diseased islet cell masses. Using oligonucleotide chips, we obtained gene expression profiles of human islets for comparison with the profiles of exocrine pancreas, liver, and kidney tissue. As periislet presence of type 1 interferon is associated with the development of type 1 diabetes, the expression profile of human islets treated ex vivo with interferon-␣2 (IFN␣2) was also determined. A set of genes encoding transmembrane-or membrane-associated proteins with novel islet-restricted expression was resolved by determining the intersection of the islet set with the complement of datasets obtained from other tissues. Under the influence of IFN␣2, the expression levels of transcripts for several of the identified gene products were up-or down-regulated. One of the islet-restricted gene products identified in this study, vesicular monoamine transporter type 2, was shown to bind 3 H]dihydrotetrabenazine, a ligand with derivatives suitable for positron emission tomography imaging. We report here the first comparison of gene expression profiles of human islets with other tissues and the identification of a target molecule with possible use in determining islet cell masses. (Endocrinology 145: 4513-4521, 2004) Tritiated and cold dihydrotetrabenazine (DTBZ) were purchased from American Radiolabeled Chemicals (St. Louis, MO). ␣-[2-3 H]DTBZ was labeled to a specific activity of 10 -20 Ci/mmol. All other chemicals, protease inhibitors, and biochemicals were purchased from Sigma-Aldrich Corp. (St. Louis, MO).