Attenuation of expression of gamma-glutamylcysteine synthetase by ribozyme transfection enhance insulin secretion by pancreatic beta cell line, MIN6.
Low levels of intracellular antioxidant enzyme activities as well as glutathione (GSH) concentrations have been described in pancreatic beta cells. We examined the effects of intracellular GSH depletion on insulin secretion and the role of intracellular GSH in signal transduction in beta cell line, MIN6 cells. Anti-gamma-glutamylcysteine synthetase (gamma-GCS) heavy subunit ribozyme was stably transfected to MIN6 cells to reduce intracellular GSH concentration. In the presence of 10 mM glucose, ribozyme-transfected cells (RTC) increased insulin secretion from 0.58 microg/10(6) cells/h in control cells (CC) to 1.48 microg/10(6) cells/h. This was associated with increased intracellular Ca(2+) concentration in RTC, detected by fluo-3 staining. Our results demonstrated that intracellular GSH concentration might influence insulin secretion by MIN6 cells, and suggest that enhanced insulin secretion by beta cells conditioned by chronic depletion of GSH is mediated by increased intracellular Ca(2+) concentration.
Kondo
H
H
First Department of Internal Medicine, Nagasaki University School of Medicine, 1-7-1 Sakamoto, Nagasaki 852-8501, Japan.
Mori
S
S
Takino
H
H
Kijima
H
H
Yamasaki
H
H
Ozaki
M
M
Tetsuya
I
I
Urata
Y
Y
Abe
T
T
Sera
Y
Y
Yamakawa
K
K
Kawasaki
E
E
Yamaguchi
Y
Y
Kondo
T
T
Eguchi
K
K
D000255:Adenosine Triphosphate / Q000378:metabolism; D000814:Aniline Compounds / Q000494:pharmacology; D000818:Animals; D015152:Blotting, Northern; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D005456:Fluorescent Dyes / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology; D005721:Glutamate-Cysteine Ligase / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000166:cytology / Q000201:enzymology*; D051379:Mice; D008856:Microscopy, Fluorescence; D010957:Plasmids / Q000378:metabolism; D016337:RNA, Catalytic / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D014162:Transfection; D014966:Xanthenes / Q000494:pharmacology
D000814:Aniline Compounds; D005456:Fluorescent Dyes; D007328:Insulin; D016337:RNA, Catalytic; D012333:RNA, Messenger; D014966:Xanthenes; C059715:Fluo-3; D000255:Adenosine Triphosphate; D005721:Glutamate-Cysteine Ligase; D005978:Glutathione; D005947:Glucose; D002118:Calcium