Haeme-oxygenase 1 expression in rat pancreatic beta cells is stimulated by supraphysiological glucose concentrations and by cyclic AMP.
OBJECTIVE
Increased expression of haeme-oxygenase 1 (HO1) and other antioxidant enzymes could improve pancreatic beta-cell survival under stressful conditions, including hyperglycaemia. However, how hyperglycaemia increases islet HO1 expression is not known.


METHODS
Rat islets were pre-cultured for 1 week in RPMI medium containing 10 mmol x l(-1) glucose (G10), and further cultured overnight in G5-G30 plus various test substances. Islet HO1 mRNA and protein expression was measured by semiquantitative RT-PCR, western blot, and immunohistochemistry.


RESULTS
Islet HO1 mRNA expression was minimal after overnight culture in G10, slightly increased in G5, and increased by five- to ten-fold in G30 in parallel with a heterogeneous increase in beta-cell HO1 protein expression. The effect of G30 was fully inhibited by agents decreasing cytosolic Ca2+ (diazoxide, nimodipine), but was only slightly reproduced by agents raising Ca2+ (tolbutamide, 30 mmol x l(-1) potassium). It was also suppressed by the alpha2-adrenoceptor agonist clonidine, whereas dibutyryl-cyclic-AMP largely increased beta-cell HO1 expression. The induction of HO1 mRNA expression by G30 was independent from changes in medium insulin concentration, but was completely inhibited by a cocktail of antioxidants. In contrast to HO1, islet mRNA expression of glutathione peroxidase and constitutive haeme-oxygenase 2 were not affected by G30, nor by dibutyryl-cyclic-AMP.


CONCLUSIONS
High glucose and dibutyryl-cyclic-AMP stimulate expression of HO1 in rat pancreatic beta cells. The inhibition of HO1 expression in G30 by nimodipine, clonidine, and antioxidants, suggests that Ca2+ influx and cyclic-AMP are necessary for the generation of oxidative stress by G30, or for the stimulation of beta-cell HO1 expression by increased oxidative stress.
Jonas
J C
JC
Unit of Endocrinology and Metabolism, Faculty of Medicine, University of Louvain, Brussels, Belgium. jonas@endo.ucl.ac.be [corrected]
Guiot
Y
Y
Rahier
J
J
Henquin
J C
JC
D000818:Animals; D001483:Base Sequence; D002118:Calcium / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism / Q000494:pharmacology*; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D005947:Glucose / Q000494:pharmacology*; D006207:Half-Life; D006419:Heme Oxygenase (Decyclizing) / Q000235:genetics*; D051547:Heme Oxygenase-1; D007515:Islets of Langerhans / Q000187:drug effects / Q000201:enzymology*; D007700:Kinetics; D008297:Male; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods
D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D000242:Cyclic AMP; D006419:Heme Oxygenase (Decyclizing); D051547:Heme Oxygenase-1; D005947:Glucose; D002118:Calcium