Molecular Dissection of Angiotensin II–Activated Human LOX-1 Promoter

Abstract

Objective-LOX-1, a receptor for oxidized low-density lipoprotein, plays a critical role in atherosclerosis. Its expression is upregulated by pro-atherogenic stimuli, such as angiotensin II (Ang II). In this study, we explored LOX-1 transcriptional promoter activation in response to Ang II in human coronary artery endothelial cells (HCAECs). Methods and Results-We constructed full-length and deletion LOX-1 promoter mutants and examined their activation in response to Ang II in HCAECs. The Ang II (1 mol/L for 24 hours) markedly induced LOX-1 promoter activity beyond the basal level, and a 116-bp fragment (between nt Ϫ2247 and Ϫ2131) was necessary for this induction. Key Words: LOX-1 Ⅲ Ang II Ⅲ oxLDL Ⅲ transcription factor Ⅲ NF-B Ⅲ oxidative stress A therosclerosis and associated diseases are major causes of death in the Western world. High plasma level of low-density lipoprotein (LDL), especially in the form of oxidized LDL (oxLDL), is a major risk factor for atherogenesis. 1 oxLDL plays a critical role in endothelial dysfunction, which appears at the early stages of atherogenesis. 2 In vitro studies have demonstrated that during oxLDL-induced endothelial dysfunction, vascular endothelial cells internalize ox-LDL through a receptor-mediated pathway. Although several scavenger receptors are involved in the internalization of oxLDL in macrophages, these receptors are absent or present only in a very small amount on endothelial cells. 3 Recently, an endothelial oxLDL receptor, LOX-1, was identified on cultured bovine aortic endothelial cells 4 and human coronary artery endothelial cells (HCAECs). 5 Many of the proatherogenic effects of oxLDL in endothelial cells appear to be mediated through LOX-1. 6 In vitro and in vivo studies have shown that LOX-1 expression is upregulated by oxLDL. 7,8 Angiotensin II (Ang II), which participates in atherogenesis, 9 also upregulates the expression of LOX-1. 10 Singh and Mehta 11 recently proposed an interaction between hyperlipidemia and renin-angiotensin system (RAS) activation in the development of atherosclerosis, during which LOX-1 plays a critical role.The nucleotide sequence of human LOX-1 promoter has been identified. 12 Although only a few details are known about the transcription factors that modulate LOX-1 gene expression, a number of potential cis-regulatory elements, such as nuclear factor-B (NF-B) and activator protein-1 (AP-1) binding motif, are found within the LOX-1 promoter sequence. 12 Nonetheless, human LOX-1 transcriptional promoter has not been thoroughly examined on the molecular level.In a recent study, we examined LOX-1 promoter activation in response to oxLDL in HCAECs, and found the transcription factor octamer-1(Oct-1) is essential for this process. 13 To delineate whether Ang II activates LOX-1 promoter through this (Oct-1) or another pathway, we analyzed Ang II-mediated activation of human LOX-1 promoter in HCAECs. Methods and Materials LOX-1 Promoter-Luciferase Reporter Plasmid ConstructionThe promoterless luciferase reporter vector (pGL3 Basic) was o...

Modulation of Angiotensin II–Mediated Hypertension and Cardiac Remodeling by Lectin-Like Oxidized Low-Density Lipoprotein Receptor-1 Deletion

“…In keeping with previous in vitro studies, 2 Ang II infusion enhanced LOX-1 expression in the wild-type mice ( Figure 5D). As expected, LOX-1 was not detectable in LOX-1 KO mice hearts.…”

“…5,6 Although LOX-1 mRNA expression is minimal in normal arterial tissues, it is markedly upregulated in vascular tissues of spontaneously hypertensive animals, suggesting a correlation between LOX-1 and hypertension. 7 This concept is supported by in vitro observations that Ang II upregulates LOX-1 expression, 2 and angiotensin-converting enzyme inhibitors and AT1R blockers decrease LOX-1 expression. 8 These findings suggest that LOX-1 overexpression may contribute to the pathological states induced by Ang II.…”

“…16 Ang II also upregulates LOX-1 expression. 2 Therefore, we determined nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase (p47 phox and p22 phox subunits) expression, ROS production, phosphorylation of oxidative stress-sensitive mitogen-activated protein kinases (MAPKs; p38 and p44/42 MAPK isoforms), and LOX-1 expression. In keeping with previous in vitro studies, 17 and p22 phox subunits, Figure 5A through 5D) in the wild-type mice, but much less so in the LOX-1 KO mice.…”

“…Previous studies have shown that Ang II via type 1 receptor (AT1R) activation stimulates the expression of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1). 2 In turn, activation of LOX-1 upregulates AT1R expression. 3 Activation of both AT1R and LOX-1 induces a state of oxidative stress.…”

Combination of fluvastatin and losartan relieves atherosclerosis and macrophage infiltration in atherosclerotic plaques in rabbits

“…Previous studies showed that statins (eg, simvastatin) significantly reduced the MCP-1 level in monocytes in vitro [12] . In addition, angiotensin II receptor blockers (eg, irbesartan) can also decrease the expression of MCP-1 and relieve the extent of atherosclerotic plaques [22,23] . However, less synergistic effects on the expression of MCP-1 were observed by the combination of statins and ARB.…”