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Domain CR9 of low-density lipoprotein receptor-related protein 1 (LRP1) is critical for aggregated LDL-induced foam cell formation from human vascular smooth muscle cells.
2015, J Biol Chem. 2015 Apr 27. pii: jbc.M115.638361. [Epub ahead of print]
Costales P1, Fuentes-Prior P2, Castellano J1, Revuelta-Lopez E1, Corral-Rodriguez MÁ2, Nasarre L1, Badimon L1, Llorente-Cortes V3.
Autors del centre relacionats: Badimon Lina, Llorente-Cortés Vicenta, Nasarre Laura, Revuelta Elena .
Costales P1, Fuentes-Prior P2, Castellano J1, Revuelta-Lopez E1, Corral-Rodriguez MÁ2, Nasarre L1, Badimon L1, Llorente-Cortes V3.
Autors del centre relacionats: Badimon Lina, Llorente-Cortés Vicenta, Nasarre Laura, Revuelta Elena .
1CSIC-ICCC, Spain;
2Biomedical Research Institute Sant Pau (IIB Sant Pau), Hospital de la S, Spain.
3CSIC-ICCC, Spain; [email protected].
Abstract
Low-density lipoprotein receptor-related protein (LRP1) mediates the internalization of aggregated LDL (AgLDL), which in turn increases the expression of LRP1 in human vascular smooth muscle cells (hVSMC). This positive feedback mechanism is thus highly efficient to promote the formation of hVSMC-foam cells, a crucial vascular component determining the susceptibility of atherosclerotic plaque to rupture. Here we have determined LRP1 domains involved in AgLDL recognition with the aim of specifically blocking AgLDL internalization in hVSMC. The capacity of fluorescently labeled AgLDL to bind to functional LRP1 clusters was tested in a receptor-ligand fluorometric assay made by immobilizing soluble LRP1 minireceptors (sLRP1-II, sLRP1-III and sLRP1-IV) recombinantly expressed in CHO cells. This assay showed that AgLDL binds to cluster II. We predicted three well exposed and potentially immunogenic peptides in CR7-CR9 domains of this cluster [termed P1, Cys1051-Glu1066, P2 (Asp1090-Cys1104) and P3 (Gly1127-Cys1140)]. AgLDL, but not native LDL, bound specifically and tightly to P3-coated wells. Rabbit polyclonal antibodies raised against P3 prevented AgLDL uptake by hVSMC and were almost twice more effective than anti-P1 and anti-P2 Abs in reducing intracellular cholesteryl ester accumulation. Moreover, anti-P3 Abs highly efficiently prevented AgLDL-induced LRP1 upregulation and counteracted the downregulatory effect of AgLDL on hVSMC migration. In conclusion, domain CR9 appears to be critical for LRP1-mediated AgLDL binding and internalization in hVSMC. Our results open new avenues for an innovative anti-VSMC-foam cell-based strategy for the treatment of vascular lipid deposition in atherosclerosis.
Copyright © 2015, The American Society for Biochemistry and Molecular Biology.
KEYWORDS:
atherosclerosis; lipid droplet; lipoprotein structure; receptor structure-function; vascular smooth muscle cells
PMID: 25918169 [PubMed - as supplied by publisher]
2Biomedical Research Institute Sant Pau (IIB Sant Pau), Hospital de la S, Spain.
3CSIC-ICCC, Spain; [email protected].
Abstract
Low-density lipoprotein receptor-related protein (LRP1) mediates the internalization of aggregated LDL (AgLDL), which in turn increases the expression of LRP1 in human vascular smooth muscle cells (hVSMC). This positive feedback mechanism is thus highly efficient to promote the formation of hVSMC-foam cells, a crucial vascular component determining the susceptibility of atherosclerotic plaque to rupture. Here we have determined LRP1 domains involved in AgLDL recognition with the aim of specifically blocking AgLDL internalization in hVSMC. The capacity of fluorescently labeled AgLDL to bind to functional LRP1 clusters was tested in a receptor-ligand fluorometric assay made by immobilizing soluble LRP1 minireceptors (sLRP1-II, sLRP1-III and sLRP1-IV) recombinantly expressed in CHO cells. This assay showed that AgLDL binds to cluster II. We predicted three well exposed and potentially immunogenic peptides in CR7-CR9 domains of this cluster [termed P1, Cys1051-Glu1066, P2 (Asp1090-Cys1104) and P3 (Gly1127-Cys1140)]. AgLDL, but not native LDL, bound specifically and tightly to P3-coated wells. Rabbit polyclonal antibodies raised against P3 prevented AgLDL uptake by hVSMC and were almost twice more effective than anti-P1 and anti-P2 Abs in reducing intracellular cholesteryl ester accumulation. Moreover, anti-P3 Abs highly efficiently prevented AgLDL-induced LRP1 upregulation and counteracted the downregulatory effect of AgLDL on hVSMC migration. In conclusion, domain CR9 appears to be critical for LRP1-mediated AgLDL binding and internalization in hVSMC. Our results open new avenues for an innovative anti-VSMC-foam cell-based strategy for the treatment of vascular lipid deposition in atherosclerosis.
Copyright © 2015, The American Society for Biochemistry and Molecular Biology.
KEYWORDS:
atherosclerosis; lipid droplet; lipoprotein structure; receptor structure-function; vascular smooth muscle cells
PMID: 25918169 [PubMed - as supplied by publisher]
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