Ang II–stimulated migration of vascular smooth muscle cells is dependent on LR11 in mice
Meizi Jiang, Hideaki Bujo, Kenji Ohwaki, Hiroyuki Unoki, Hiroyuki Yamazaki, Tatsuro Kanaki, Manabu Shibasaki, Kazuhiko Azuma, Kenichi Harigaya, Wolfgang J. Schneider, Yasushi Saito
Meizi Jiang, Hideaki Bujo, Kenji Ohwaki, Hiroyuki Unoki, Hiroyuki Yamazaki, Tatsuro Kanaki, Manabu Shibasaki, Kazuhiko Azuma, Kenichi Harigaya, Wolfgang J. Schneider, Yasushi Saito
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Research Article Cardiology

Ang II–stimulated migration of vascular smooth muscle cells is dependent on LR11 in mice

Abstract

Medial-to-intimal migration of SMCs is critical to atherosclerotic plaque formation and remodeling of injured arteries. Considerable amounts of the shed soluble form of the LDL receptor relative LR11 (sLR11) produced by intimal SMCs enhance SMC migration in vitro via upregulation of urokinase-type plasminogen activator receptor (uPAR) expression. Here, we show that circulating sLR11 is a novel marker of carotid intima-media thickness (IMT) and that targeted disruption of the LR11 gene greatly reduces intimal thickening of arteries through attenuation of Ang II–induced migration of SMCs. Serum concentrations of sLR11 were positively correlated with IMT in dyslipidemic subjects, and multivariable regression analysis suggested sLR11 levels as an index of IMT, independent of classical atherosclerosis risk factors. In Lr11–/– mice, femoral artery intimal thickness after cuff placement was decreased, and Ang II–stimulated migration and attachment of SMCs from these mice were largely abolished. In isolated murine SMCs, sLR11 caused membrane ruffle formation via activation of focal adhesion kinase/ERK/Rac1 accompanied by complex formation between uPAR and integrin αvβ3, a process accelerated by Ang II. Overproduction of sLR11 decreased the sensitivity of Ang II–induced activation pathways to inhibition by an Ang II type 1 receptor blocker in mice. Thus, we demonstrate a requirement for sLR11 in Ang II–induced SMC migration and propose what we believe is a novel role for sLR11 as a biomarker of carotid IMT.

Authors

Meizi Jiang, Hideaki Bujo, Kenji Ohwaki, Hiroyuki Unoki, Hiroyuki Yamazaki, Tatsuro Kanaki, Manabu Shibasaki, Kazuhiko Azuma, Kenichi Harigaya, Wolfgang J. Schneider, Yasushi Saito

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Figure 7

sLR11-mediated intracellular signals related to cytoskeleton reorganization.

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sLR11-mediated intracellular signals related to cytoskeleton reorganizat...
(A) sLR11-induced FAK activation. Cell lysates of rabbit SMCs were incubated with or without sLR11 (1 μg/ml) in the presence or absence of antibody against LR11 or integrin αvβ3 (MAB1976), immunoprecipitated with anti-FAK antibody, and subjected to immunoblot analysis using anti-FAK (~130 kDa) or anti–phospho-FAK (~130 kDa) antibody. (B) sLR11–induced phosphorylation of ERK1/2. Cell lysates (10 μg protein) of rabbit SMCs were incubated with sLR11 (1 μg/ml) for the indicated times in the presence or absence of anti–integrin αvβ3 antibody (MAB1976) and subjected to immunoblot analysis using antibody against (phospho) p42/44 MAP kinase. Upper and lower signals represent ERK1 (~44 kDa) and ERK2 (~42 kDa), respectively (13). Blot shown is representative of 3 independent experiments. Data of p-ERK1/2 are presented as mean ± SD (n = 3). *P < 0.05. (C) Rac1 activation in Lr11–/– SMCs. Cell lysates (60 μg protein) of Lr11+/+ or Lr11–/– SMCs were incubated with sLR11 (1 μg/ml) in the presence or absence of antibody against integrin αvβ3 (RMV-7), immunoprecipitated with PAK-1 PBD Protein GST beads, and subjected to immunoblot analysis with anti-Rac1 (~21 kDa) or anti–GTP-Rac1 (~21 kDa) antibody. (D) sLR11-induced Rac1 activation. Cell lysates (60 μg protein) of rabbit SMCs were incubated with sLR11 (1 μg/ml) for the indicated times in the presence or absence of anti–integrin αvβ3 antibody (MAB1976) and subjected to immunoblot analysis with anti-Rac1 (Rac1 ~21 kDa and GST-Rac1 ~21 kDa) antibody with (top) or without (bottom) prior immunoprecipitation with PAK-1 PBD Protein GST beads.