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- Jiahong Zue and
- Canzhan Zhu
ATP binding cassette transporter G1 (ABCG1), a regulator of cholesterol efflux to HDL, has been found its up-regulation reversed the high glucose-induced oxidative stress in endothelial cells in our previous study. The present study aimed to further implore whether the endogenous antioxidant gene regulator, nuclear factor erythroid-2-related factor 2(Nrf2) was involved in the effect of ABCG1 against oxidative stress.
Human umbilical vein endothelial cells(HUVECs) were treated with or without D-glucose (5.6 mM and 30.0 mM) and LXR agonist T0901307 for 24-72 h. Real time PCR and Western blot were employed to measure the expression of ABCG1 and antioxidative target gene of Nrf2, such as heme oxygenase 1 (HO-1). Furthermore, Nrf2-antioxidant response element (ARE) binding activity as well as nuclear translocation of Nrf2 was evaluated by electromobility shift assay and Western blotting. Intracellular reactive oxygen species (ROS) were measured using dihydroethidium (DHE) fluorescence. Also, HUVECs were transfected with siRNAs of ABCG1 or Nrf2 and ABCG1 overexpression plasmid to further observed the relationship between ABCG1 and Nrf2 activity.
Compared with normal glucose, high glucose reduced the expression of ABCG1 and induced ROS production in HUVECs in a time-dependent manner. Furthermore, high glucose increased the Nrf2 nuclear translocation and Nrf2-ARE binding activity, which was along with increased HO-1 mRNA and protein expression by 20% at the culture of 1 day. However, Nrf2 nuclear expression and Nrf2-ARE binding activity were inhibited by 20% and 30% in hyperglycemia-cultured ECs for next culture of 3 and 5 days. Subsequently, HO-1 expression was significantly decreased by 20% and a markedly increased ROS level was found. In addition, upregulation of ABCG1 using LXR agonist T0901307 or ABCG1 overexpression plasmid both markedly reversed the high glucose-decreased nuclear Nrf2 translocation and ARE-regulated HO-1 expression at 3 and 5 days, which was accompanied with decreased levels of ROS. On the contrary, ABCG1 siRNA was showed to block the Nrf2/HO-1 expression and subsequently to increase ROS levels. Finally, Nrf2 siRNA abrogated ABCG1-mediatiated suppression of high glucose-induced oxidative stress in ECs.
It is suggested that endothelial protection regulated by ABCG1 is associated with Nrf2/HO-1-dependent antioxidative passway.