Regulation of glucokinase gene expression in cultured rat islet cells: the inhibitory effects of T3 and glucagon, and the stimulatory effect of glibenclamide.
Hormonal and non-hormonal regulation of glucokinase gene expression was investigsted in cultured rat islet cells. To measure glucokinase mRNA in pancreatic islet cells, the competitive PCR method was adopted. With this method, GKmRNA levels can be measured using only 0.1-1.0 microgram of total RNA isolated from cultured rat islet cells. Following 24 h preculture with 5.5 mM glucose, islet cells were cultured for 24 or 8 h with hormonal or non-hormonal factors. Glucokinase mRNA levels tended to increase, but not significantly, at 16.7 mM glucose compared to those at 5.5 mM glucose. Treatment with either 1 microM T3 or 1 microM glucagon resulted in a decrease in the glucokinase mRNA level with 16.7 mM glucose, whereas 1 microM insulin had no effect on glucokinase mRNA. Five mM dibutyryl cyclic AMP decreased the glucokinase mRNA level with 16.7 mM glucose, but cycloheximide did not block this inhibitory effect, suggesting that the effect of glucagon may be mediated by cyclic AMP and that protein synthesis is not involved in the response. Furthermore, the islet glucokinase mRNA level increased in response to 1 microM glibenclamide with 5.5 mM glucose and the response was abolished by cycloheximide, which indicates the involvement of protein synthesis in the glibenclamide-induced mRNA change. An 8-bromo-cyclic GMP (1 microM) and vanadate (1 microM) did not affect the islet GKmRNA level. These findings suggested that thyroid hormone and glucagon-cyclic AMP suppress, and glibenclamide increases the GKmRNA level in cultured rat islet cells, and that insulin, cyclic GMP and vanadate differentially affect glucokinase gene expression in pancreatic islet cells and in the liver.
Hinata
S
S
Second Department of Internal Medicine, Hamamatsu University School of Medicine, Japan.
Nishi
S
S
Matsukage
T
T
Funai
T
T
Ichiyama
A
A
Yoshimi
T
T
D000818:Animals; D001483:Base Sequence; D003994:Bucladesine / Q000494:pharmacology; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000031:analogs & derivatives / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D018076:DNA, Complementary / Q000032:analysis; D015971:Gene Expression Regulation, Enzymologic; D005934:Glucagon / Q000494:pharmacology*; D005941:Glucokinase / Q000096:biosynthesis / Q000235:genetics*; D005947:Glucose / Q000494:pharmacology; D005905:Glyburide / Q000494:pharmacology*; D007328:Insulin / Q000494:pharmacology; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D014284:Triiodothyronine / Q000494:pharmacology*; D014638:Vanadates / Q000494:pharmacology
D018076:DNA, Complementary; D007328:Insulin; D012333:RNA, Messenger; D014284:Triiodothyronine; C016276:8-bromocyclic GMP; D014638:Vanadates; D003994:Bucladesine; D005934:Glucagon; D003513:Cycloheximide; D005941:Glucokinase; D006152:Cyclic GMP; D005947:Glucose; D005905:Glyburide