Phosphorylation of p38 mitogen-activated protein kinase downstream of bax-caspase-3 pathway leads to cell death induced by high D-glucose in human endothelial cells.
Because high D-glucose significantly stimulates endothelial cell death, we examined the molecular mechanisms of high D-glucose-induced endothelial apoptosis. Treatment of human aortic endothelial cells with high D-glucose (25 mmol/l), but not mannitol and L-glucose, resulted in a significant decrease in cell number and a significant increase in apoptotic cells as compared with a physiological concentration (5 mmol/l). Interestingly, high D-glucose treatment significantly increased bax protein, accompanied by translocation of bax protein from cytosol to mitochondria-enriched heavy membrane fraction. In contrast, the expression and distribution of bcl-2 protein were not altered by high D-glucose. In addition, the activity of caspase-3 proteases was increased after exposure to high glucose, whereas caspase inhibitors prevented endothelial cell death induced by high D-glucose. On the other hand, p38 mitogen-activated protein kinase (MAPK) was markedly phosphorylated and showed sustained phosphorylation after stimulation. A specific inhibitor of p38 MAPK, SB 203580, and the overexpression of kinase-inactive p38 MAPK significantly attenuated cell death induced by high D-glucose in human aortic endothelial cells, whereas at 6 h after high D-glucose treatment, SB 203580 and overexpression of kinase-inactive p38 MAPK did not attenuate caspase-3 activation induced by high D-glucose. Importantly, caspase inhibitors significantly attenuated the sustained phosphorylation of p38 MAPK induced by high D-glucose. Thus, we finally focused the MAPK kinase (MEK) kinase 1 (MEKK1) to further examine the cross-talk between p38 MAPK and the bax-caspase proteases pathway. High D-glucose treatment induced MEKK1 cleavage, whereas caspase inhibitors significantly attenuated the cleavage. Importantly, kinase-inactive MEKK1 also blocked the phosphorylation of p38 MAPK induced by high D-glucose. Here, we demonstrated that high D-glucose induced apoptosis in human endothelial cells through activation of the bax-caspase proteases pathway and through phosphorylation of p38 MAPK mediated by MEKK1. Phosphorylation of p38 MAPK downstream of the bax-caspase pathway may play a pivotal role in endothelial apoptosis mediated by high D-glucose.
Nakagami
H
H
Department of Geriatric Medicine, Osaka University Medical School, Yamada-oka, Suita, Osaka, Japan.
Morishita
R
R
Yamamoto
K
K
Yoshimura
S I
SI
Taniyama
Y
Y
Aoki
M
M
Matsubara
H
H
Kim
S
S
Kaneda
Y
Y
Ogihara
T
T
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D002417:Cattle; D016923:Cell Death / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D004789:Enzyme Activation / Q000502:physiology; D005947:Glucose / Q000494:pharmacology*; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D048051:p38 Mitogen-Activated Protein Kinases
C494826:BAX protein, human; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D005947:Glucose