Cytoprotective effect of β-lapachone by inducing heme oxygenase-1 expression and AMP-activated protein kinase activation in human endothelial cells.
OBJECTIVE
AMP-activated protein kinase (AMPK) is suggested to exert cytoprotective and anti-inflammatory effects in endothelial cells, but the precise mechanisms are not fully understood. It has been reported that pharmacological activation of AMPK induces endothelial heme oxygenase-1 (HO-1) expression. β-Lapachone (BL), a well-known substrate of
quinone oxidoreductase (NQO1), stimulates AMPK activation via NQO1 activation. Here we examined whether AMPK activation by BL would be linked to HO-1 expression in ECV304 endothelial cells and whether HO-1 expression could mediate the cytoprotective effect of BL.


METHODS
Endothelial cells were pre-incubated for 6 h with BL or 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR) in the absence or presence of dicoumarol (DC), compound C (CC), or tin protoporphyrin-IX (SnPP), and then challenged with tumor necrosis factor-α (TNF-α) for 24 h. Cell viability was evaluated by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide (MTT) assay. AMPK phosphorylation and HO-1 expression were detected by Western blot analysis.


RESULTS
At non-cytotoxic concentrations, BL induced AMPK phosphorylation and HO-1 expression. AICAR, an AMPK activator, also induced HO-1 expression. In contrast, CC, an inhibitor of AMPK activation, and DC, an inhibitor of NQO1, prevented the increase in BL-induced HO-1 expression. Pretreatment with BL or AICAR reduced TNF-α-induced endothelial cell death. Cytoprotection by BL was almost completely abolished by CC and DC and partly by SnPP, a competitive inhibitor of HO-1.


CONCLUSIONS
Our results suggest that BL induces cytoprotective HO-1 expression in endothelial cells via AMPK activation, providing one of possible mechanisms by which BL can exert beneficial effects.
Department of Surgery, Wonkwang University School of Medicine, Iksan, Republic of Korea. hopae@wku.ac.kr.
S J
SJ
Byun
Y
Y
Son
H-O
HO
Pae
D055372:AMP-Activated Protein Kinases / Q000378:metabolism*; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D019610:Cytoprotection; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D051547:Heme Oxygenase-1 / Q000378:metabolism*; D006801:Humans; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000378:metabolism; D009285:Naphthoquinones / Q000494:pharmacology*; D020011:Protective Agents / Q000494:pharmacology*; D014409:Tumor Necrosis Factor-alpha / Q000633:toxicity
D009285:Naphthoquinones; D020011:Protective Agents; D014409:Tumor Necrosis Factor-alpha; C014638:beta-lapachone; C496541:HMOX1 protein, human; D051547:Heme Oxygenase-1; D016660:NAD(P)H Dehydrogenase (Quinone); C447479:NQO1 protein, human; D055372:AMP-Activated Protein Kinases