title: Regulation of GAD expression in islets of Langerhans occurs both at the mRNA and protein level.
abstract: The expression of the autoantigen glutamate decarboxylase in islets of Langerhans was investigated under different culture conditions, which affect the functional activity of the beta-cell. Using immunoprecipitations and analyses of enzyme activity, an increase in glutamate decarboxylase was detected in rat islets cultured at a glucose concentration of 11 mmol/l compared with those cultured at 5.6 mmol/l glucose. To determine whether the change was induced at the level of mRNA expression, total RNA was extracted from rat islets cultured at 5.6 or 11 mmol/l glucose, reverse transcribed and amplified by the polymerase chain reaction. Comparative quantitation in a phosphor imager revealed a significantly higher (82%, P < 0.005) content of glutamate decarboxylase mRNA in islets cultured at 11 mmol/l glucose. In parallel, human recombinant interleukin-1 beta, and diazoxide were tested for their effects on the expression of glutamate decarboxylase. Islets cultured at 11 mmol/l glucose in the presence of 40 U/ml of interleukin-1 beta, showed a 63% decrease (P < 0.005) in enzyme activity compared with those cultured at 11 mmol/l glucose alone, and similar decreases were noted on analysis of glutamate decarboxylase biosynthesis and mRNA. Islets cultured at 11 mmol/l glucose in the presence of 22.5 mg/ml diazoxide exhibited a significant reduction in enzyme activity (59%; P < 0.001) compared with those cultured at 11 mmol/l glucose only. This reduction, however, was not accompanied by a decrease in the content of glutamate decarboxylase mRNA.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Internal Medicine, University Hospital, Uppsala, Sweden.
authors.forename: L A
authors.initials: LA
authors.lastname: Velloso
authors.forename: E
authors.initials: E
authors.lastname: Björk
authors.forename: A E
authors.initials: AE
authors.lastname: Ballagi
authors.forename: K
authors.initials: K
authors.lastname: Funa
authors.forename: A
authors.initials: A
authors.lastname: Andersson
authors.forename: O
authors.initials: O
authors.lastname: Kämpe
authors.forename: F A
authors.initials: FA
authors.lastname: Karlsson
authors.forename: D L
authors.initials: DL
authors.lastname: Eizirik
mesh_terms: D000818:Animals; D001483:Base Sequence; D015342:DNA Probes; D015971:Gene Expression Regulation, Enzymologic; D005947:Glucose / Q000494:pharmacology; D005968:Glutamate Decarboxylase / Q000096:biosynthesis* / Q000187:drug effects / Q000235:genetics; D006801:Humans; D007328:Insulin / Q000096:biosynthesis; D007375:Interleukin-1 / Q000494:pharmacology; D007515:Islets of Langerhans / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D008297:Male; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D015342:DNA Probes; D007328:Insulin; D007375:Interleukin-1; D012333:RNA, Messenger; D005968:Glutamate Decarboxylase; D005947:Glucose