Inhibition of Nox-4 activity by plumbagin, a plant-derived bioactive naphthoquinone

NAD(P)H oxiclase contributes to the pathogenesis of cancer and cardiovascular diseases such as hypertension, atherosclerosis, restenosis, cardiac hypertrophy and heart failure. Plumbagin, a plant-derived naphthoquinone, has been shown to exert anticarcinogenic and anti-atherosclerorsis effects in animals. However, the molecular mechanisms underlying these effects remain unknown. it. is possible that the beneficial effect of plumbagin is due to the inhibition of NAD(P)X oxidise. Human embryonic kidney 293 (HEK293) and brain tumour LN229 cells express mainly Nox-4, a renal NAD(P)H oxidase. We have examined the effect of plumbagin on Nox-4 activity in HEK293 and LN229 cells using lucigenin-dependent chemiluminescence assay. Plumbagin inhibited the activity or Nox-4 in a time- and dose-dependent manner in HEK293 and LN229 cells. Production or superoxide in HEK293 cells was inhibited by diphenyleneiodonium (DPI), a MAD(P)H oxiclase inhibitor. the superoxide production in HEK293 cells was NADPH- and NADH-dependent indicating that the superoxide was generated by a NAD(P)H oxiclase in HEK293 cells, but not by the redox-cycling or lucigenin. Furthermore, plumbagin inhibited the superoxide production in Nox-4 transfected COS-7 cells. These results indicated that plumbagin directly interacted with Nox-4 and inhibited its activity.