Amylin-induced cytotoxicity is associated with activation of caspase-3 and MAP kinases.
Nanomolar concentrations of human amylin promote death of RINm5F cells in a time- and concentrationdependent manner. Morphological changes of chromatin integrity suggest that cells are predominantly undergoing apoptosis. Human amylin induces significant activation of caspase-3 and strong and sustained phosphorylation of stress-activated protein kinases, c-Jun N-terminal kinase (JNK) and p38, that precedes cell death. Extracellular signal-regulated kinase (ERK) activation was not concomitant with JNK and/or p38 activation. Activation of caspase-3 and mitogen-activated protein kinases (MAPKs) was detected by Western blot analysis. Addition of the MEK1 inhibitor PD 98059 had no effect on amylin-induced apoptosis, suggesting that ERK activation does not play a role in this apoptotic scenario. A correlative inhibition of JNK activation by the immunosuppressive drug FK506, as well as a selective inhibition of p38 MAPK activation by SB 203580, significantly suppressed procaspase-3 processing and the extent of amylin-induced cell death. Moreover, simultaneous pretreatment with both FK506 and SB 203580, or with the caspase-3 inhibitor Ac-DEVD-CHO alone, almost completely abolished procaspase-3 processing and cell death. Thus, our results suggest that amylin-induced apoptosis proceeds through sustained activation of JNK and p38 MAPK followed by caspase-3 activation.
Rumora
Lada
L
Department of Medical Biochemistry and Haematology, Faculty of Pharmacy and Biochemistry, Domagojeva 2,10000 Zagreb, Croatia.
Hadzija
Mirko
M
Barisić
Karmela
K
Maysinger
Dusica
D
Grubiić
Tihana Zanić
TZ
D000682:Amyloid / Q000494:pharmacology*; D000818:Animals; D015153:Blotting, Western; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D016923:Cell Death / Q000187:drug effects; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003124:Colorimetry; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007340:Insulinoma / Q000473:pathology; D058228:Islet Amyloid Polypeptide; D007515:Islets of Langerhans / Q000187:drug effects*; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010190:Pancreatic Neoplasms / Q000473:pathology; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects; D013778:Tetrazolium Salts; D013844:Thiazoles
D000682:Amyloid; D058228:Islet Amyloid Polypeptide; D013778:Tetrazolium Salts; D013844:Thiazoles; D007770:L-Lactate Dehydrogenase; D020928:Mitogen-Activated Protein Kinases; C505407:CASP3 protein, human; C505410:Casp3 protein, rat; D053148:Caspase 3; D020169:Caspases; C022616:thiazolyl blue