Angiotensin II increases vascular proteoglycan content preceding and contributing to atherosclerosis development

Abstract

Angiotensin II (angII) is known to promote atherosclerosis; however, the mechanisms involved are not fully understood. To determine whether angII stimulates proteoglycan production and LDL retention, LDL receptordeficient mice were infused with angII (1,000 ng/kg/min) or saline via osmotic minipumps. To control for the hypertensive effect of angII, a parallel group received norepinephrine (NE; 5.6 mg/kg/day). Arterial lipid accumulation was evaluated by measuring the retention rate of LDL in isolated carotid arteries perfused ex vivo. Mice infused with angII had increased vascular content of biglycan and perlecan and retained twice as much LDL as saline-or NEinfused mice, although no group developed atherosclerosis at this time. To determine whether this increase in biglycan and perlecan content predisposed to atherosclerosis development, mice were infused with angII, saline, or NE for 4 weeks, then pumps were removed and mice received an atherogenic Western diet for another 6 weeks. Mice that had received angII infusions had 3-fold increased atherosclerosis compared with mice that had received saline or NE, and apolipoprotein B colocalized with both proteoglycans. Thus, one mechanism by which angII promotes atherosclerosis is increased proteoglycan synthesis and increased arterial LDL retention, which precedes and contributes to atherosclerosis

Serum Amyloid A, but Not C-Reactive Protein, Stimulates Vascular Proteoglycan Synthesis in a Pro-Atherogenic Manner

“…14,50 -52 ApoB has also been reported to co-localize with perlecan in mice, 33,50 and decorin in humans. 51 However, decorin overexpression in mice was associated with decreased atherosclerosis development, 53 and studies examining atherosclerosis in mice heterozygous for perlecan (perlecan deficiency is lethal) are complex, with only male apoE Ϫ/Ϫ mice showing decreased atherosclerosis, and no differences in atherosclerosis extent in female apoE Ϫ/Ϫ or either gender of LDLR Ϫ/Ϫ mice.…”

“…Twenty-eight days after injections, aortas were collected, stripped of adventitia, and total protein was extracted and used in Western blot analyses to quantify proteoglycan content as previously described. 33 ␤-Actin was used as the loading control. Atherosclerosis was quantified in the aortic root, en face aorta, and brachiocephalic artery as previously described.…”

“…The mechanisms by which SAA exerts its effects are not clear; however, there is striking similarity between observed effects of SAA on biglycan with that previously observed with TGF-␤. 29,30,33,[42][43][44] Thus, TGF-␤ levels were quantified to determine whether SAA stimulated biglycan expression via stimulation of endogenous TGF-␤. Conditioned media from vascular smooth muscle cells exposed to SAA had increased amounts of both total TGF-␤ (not shown) and endogenously active TGF-␤ compared to media from unexposed cells (165 Ϯ 11 pg/ml versus 68 Ϯ 24 pg/ml, respectively; P ϭ 0.04).…”

“…29,32 Dried gels were exposed to Fuji Imaging Plates for 48 to 72 hours and then visualized using FLA-5000 (Fujifilm, Tokyo, Japan) and analyzed using Multi Gauge software (Fujifilm). To evaluate proteoglycan synthesis total proteins were precipitated from the culture media of each condition by trichloroacetic acid, and biglycan, versican, and decorin were evaluated by Western blots as previously described 33 using antibodies to biglycan (R&D Systems); decorin (generously provided by Dr. Larry Fisher, National Institutes of Health, Bethesda, MD), and versican (Chemicon, Temecula, CA). ␤-Actin was used as the loading control (Abcam, Cambridge, MA).…”

“…Atherosclerosis was quantified in the aortic root, en face aorta, and brachiocephalic artery as previously described. 14,39,40 Immunohistochemical staining was performed on adjacent aortic root sections as previously described. 33 …”