Bone Morphogenic Protein 4 Produced in Endothelial Cells by Oscillatory Shear Stress Induces Monocyte Adhesion by Stimulating Reactive Oxygen Species Production From a Nox1-Based NADPH Oxidase

Abstract

Abstract-Atherosclerosis is an inflammatory disease occurring preferentially in arterial regions exposed to disturbed flow conditions including oscillatory shear stress (OS). OS exposure induces endothelial expression of bone morphogenic protein 4 (BMP4), which in turn may activate intercellular adhesion molecule-1 (ICAM-1) expression and monocyte adhesion. OS is also known to induce monocyte adhesion by producing reactive oxygen species (ROS) from reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidases, raising the possibility that BMP4 may stimulate the inflammatory response by ROS-dependent mechanisms. Here we show that ROS scavengers blocked ICAM-1 expression and monocyte adhesion induced by BMP4 or OS in endothelial cells (ECs). Similar to OS, BMP4 stimulated H 2 O 2 and O 2 Ϫ production in ECs. Next, we used ECs obtained from p47phox Ϫ/Ϫ mice (MAE-p47 Ϫ/Ϫ ), which do not produce ROS in response to OS, to determine the role of NADPH oxidases. Similar to OS, BMP4 failed to induce monocyte adhesion in MAE-p47 Ϫ/Ϫ , but it was restored when the cells were transfected with p47 phox plasmid. Moreover, OS-induced O 2 Ϫ production was blocked by noggin (a BMP antagonist), suggesting a role for BMP. Furthermore, OS increased gp91phox (nox2) and nox1 mRNA levels while decreasing nox4. In contrast, BMP4 induced nox1 mRNA expression, whereas nox2 and nox4 were decreased or not affected, respectively. Also, OS-induced monocyte adhesion was blocked by knocking down nox1 with the small interfering RNA (siRNA). Finally, BMP4 siRNA inhibited OS-induced ROS production and monocyte adhesion. Together, these results suggest that BMP4 produced in ECs by OS stimulates ROS release from the nox1-dependent NADPH oxidase leading to inflammation, a critical early atherogenic step. Key words: BMP4 Ⅲ oscillatory shear Ⅲ reactive oxygen species Ⅲ monocyte adhesion Ⅲ endothelial cells Ⅲ NADPH oxidase V ascular endothelial cells (ECs) are constantly exposed to fluid shear stress, the frictional force generated by blood flow over the vascular endothelium. The importance of shear stress in vascular biology and pathophysiology has been highlighted by the focal development patterns of atherosclerosis in hemodynamically defined regions. For example, the regions of branched and curved arteries exposed to disturbed flow conditions including oscillatory shear stress (OS) correspond to "lesion-prone areas" that preferentially develop atherosclerosis. 1,2 In contrast, straight arteries exposed to steady, high levels of laminar shear stress (LS) are relatively well protected from atherosclerotic plaque development. 1,2 Atherosclerosis is an inflammatory disease preferentially occurring in lesion-prone areas. 2,3 The earliest measurable markers of atherogenesis include expression of inflammatory adhesion molecules such as E-selectin, vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1), and subsequent monocyte adhesion and recruitment into the lesion-prone areas. 2,4,5 Additional critical atherogen...

Oxidized-1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine induces vascular endothelial superoxide production: Implication of NADPH oxidase

“…We have previously shown that Nox4 is more abundantly expressed than Nox2 in cultured BAEC in response to fluid shear stress [25]. In static cultured endothelial cells obtained from the thoracic aortas of C57BL6, Nox4 mRNA was reported to be far more abundant than Nox1 and Nox2 [68]. To show intracellular O 2 −• production, we used dihydroethidium and 2′,7′-dichlorodihydrofluorescein diacetate to account for Nox4 activity in BAEC [69].…”

Nox enzymes, ROS, and chronic disease: An example of antagonistic pleiotropy

“…Oscillatory sheer stress results in several-fold induction of Nox1, Nox2 and Nox4 in vascular endothelium (with opposite effects of the anti-atherogenic laminar flow that occurs in straight portions of vessels) [80,81]. Oscillatory sheer stress is associated with induction of bone morphogenic protein 4 (BMP4) [82], which induces Nox1 and p47phox, resulting in an oxidative stress that leads to ICAM-1 expression and monocyte adhesion [81]. Nox1 expression increases ~3-fold following balloon injury and precedes restenosis and atherosclerosis [83].…”

Basic Mechanisms of Oxidative Stress and Reactive Oxygen Species in Cardiovascular Injury

“…Importantly, oscillatory shear stress leads to continuous O 2 •− production in a NADPH oxidase-dependent manner (Hwang et al 2003), resulting in nuclear factor-κB (NF-κB)-mediated monocyte adhesion. Recent work has shown that bone morphogenic protein-4 (BMP-4) plays a significant role in the response to oscillatory shear stress by stimulating ROS from NADPH oxidases and regulating the expression of ICAM-1 (Sorescu et al 2004). …”