Omega-3 fatty acid supplement reduces activation of NADPH oxidase in intracranial atherosclerosis stenosis

ABSTRACT

Objectives Intracranial atherosclerotic stenosis (ICAS) is one of the most common causes of stroke worldwide. We adapted a rat model of atherosclerosis to study brain intracranial atherosclerosis, and further investigated how omega-3 fatty acids (O3FA) attenuated the development of ICAS by reducing the generation of reactive oxygen species (ROS) and the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) activity.

Methods Adult male Sprague-Dawley rats were divided into control normal-cholesterol or high-cholesterol diet groups with or without O3FA for up to 6 weeks. NG-nitro-L-arginine methyl ester (L-NAME, 3 mg/mL), a nitric oxide synthase inhibitor, was added to the drinking water of the high-cholesterol groups during the first 2 weeks. The rats received supplementation with O3FA (5 mg/kg/day) by gavage. At 3 and 6 weeks, we measured blood lipid levels, including low-density lipoprotein (LDL), cholesterol (CHO), triglycerides (TG), and high-density lipoprotein (HDL) as atherosclerotic blood markers. The lumen of middle cerebral artery (MCA) and the thickness of the vessel wall were assessed histologically. ROS production was measured. NOX activity and mRNA and protein expression of NOX subunits (p47^phox, gp91^phox, p22^phox, and p67^phox) were measured.

Results A high-cholesterol diet exhibited a significant increase in the classic blood markers (LDL, CHO, and TG) for atherosclerosis, as well as a decrease in HDL. These markers were found to be progressively more severe with time. Additionally, increased lumen stenosis and intimal thickening were observed in the MCA for this group. Rats given O3FA demonstrated attenuation of blood lipid levels with an absence of morphological changes.O3FA significantly reduced ROS production and NOX activity in the brain. Moreover, O3FA decreased the mRNA and protein expression of the NOX subunits p47^phox, gp91^phox, and p67^phox.

Conclusions Long-term O3FA dietary supplementation prevents the development of intracranial atherosclerosis. This O3FA effect appears to be mediated by its attenuation of NOX subunit expression and NOX activity, therefore reducing ROS production. O3FA dietary supplement shows promising results in the prevention of ICAS.

KEYWORDS:

• High-cholesterol diet
• NG-nitro-L-arginine methyl ester (L-NAME)
• brain
• middle cerebral artery (MCA)
• reactive oxygen species (ROS)

Acknowledgments

This work was partially supported by the Wayne State University Neurosurgery Fund; the Merit Review Award (I01RX-001964-01) from the U.S. Department of Veterans Affairs Rehabilitation R&D Service; the National Natural Science Foundation of China (81501141); Science and Technology Project of Beijing Municipal Education Commission (KM201610025028),Talent Training Program of Beijing Ministry of Organization (2013D008011000002),the Beijing NOVA program (xx2016061), and Science and Technology Plan of Beijing Tongzhou District (KJ2017CX043).