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Vascular smooth muscle cells (vSMC) exert a critical role in sensing and maintaining vascular integrity. These cells abundantly express the low-density lipoprotein receptor-related protein 1 (LRP1), a large endocytic and signaling receptor that recognizes numerous ligands including ApoE-rich lipoproteins, proteases, and protease-inhibitor complexes. We observed the spontaneous formation of aneurysms in the superior mesenteric artery (SMA) of both male and female mice in which LRP1 was genetically deleted in v SMC (smLRP1-/- mice). Quantitative proteomics revealed elevated abundance of several proteins in smLRP1-/- mice that are known to be induced by angiotensin II (AngII)-mediated signaling, suggesting that this pathway is dysregulated. Administration of losartan, an AngII type I receptor antagonist, or an angiotensinogen antisense oligonucleotide to reduce plasma angiotensinogen concentrations restored the normal SMA phenotype in smLRP1-/- mice and prevented aneurysm formation. Additionally, employing a vascular injury model, we noted excessive vascular remodeling and neointima formation in smLRP1-/- mice that was restored by losartan administration. Together, these findings reveal that LRP1 regulates vascular integrity and remodeling of the SMA by attenuating excessive AngII-mediated signaling.

LRP1 (low density lipoprotein receptor-related protein 1) is a large endocytic receptor that binds over 100 structurally unrelated ligands. Our lab has demonstrated that a sm22 promoter-driven, smooth muscle specific knock-out of LRP1 (smLRP1-/-) results in aortic aneurysms in mice due to fragmentation and degradation of the elastic fibers, further implicating LRP1 in vascular development. Furthermore, the identification of a patient with abdominal aortic aneurysm (AAA) harboring missense mutations in LRP1 has allowed for investigations into the role of LRP1 in aneurysm formation. Utilizing a combination of biochemical and proteomic techniques, we sought to understand the role of these mutations in cellular function. By means of a receptor-ligand binding assay, we quantified the internalization and degradation of 125 I-activated alpha-2-macroglobulin (a2M*) in LRP1-deficient cells transfected with mutant LRP1 and aortic smooth muscle cells isolated from a patient with an abdominal aortic aneurysm and LRP1 missense mutations. The results showed defects in LRP1-mediated internalization of a 2 M* in LRP1 variants when compared to wild type LRP1. Furthermore, through extensive proteomic analysis of primary patient cells harboring these mutations, multiple dysregulated pathways of interest have been identified along with elevated levels of MMP-1 in the cultured media. In total, our data highlights the biochemical deficits in different variants of LRP1 that may contribute to the pathogenesis of AAA. Pinpointing the role of LRP1 in aneurysm mechanisms will allow for non-invasive interventions to be employed before aortic rupture.

Matrix metalloprotease (MMP) activation contributes to the degradation of the extracellular matrix (ECM), resulting in a multitude of pathologies. Low-density lipoprotein receptor-related protein 1 (LRP1) is a multifaceted endocytic and signaling receptor that is responsible for internalization and lysosomal degradation of diverse proteases, protease inhibitors, and lipoproteins along with numerous other proteins. In this study, we identified MMP-1 as a novel LRP1 ligand. Binding studies employing surface plasmon resonance revealed that both proMMP-1 and active MMP-1 bind to purified LRP1 with equilibrium dissociation constants (KD) of 19 and 25 nM, respectively. We observed that human aortic smooth muscle cells readily internalize and degrade 125I-labeled proMMP-1 in an LRP1-mediated process. Our binding data also revealed that all tissue inhibitors of metalloproteases (TIMPs) bind to LRP1 with KD values ranging from 23 to 33 nM. Interestingly, the MMP-1/TIMP-1 complex bound to LRP1 with an affinity (KD = 0.6 nM) that was 30-fold higher than that of either component alone, revealing that LRP1 prefers the protease:inhibitor complex as a ligand. Of note, modification of lysine residues on either proMMP-1 or TIMP-1 ablated the ability of the MMP-1/TIMP-1 complex to bind to LRP1. LRP1's preferential binding to enzyme:inhibitor complexes was further supported by the higher binding affinity for proMMP-9/TIMP-1 complexes than for either of these two components alone. LRP1 has four clusters of ligand-binding repeats, and MMP-1, TIMP-1, and MMP-1/TIMP-1 complexes bound to cluster III most avidly. Our results reveal an important role for LRP1 in controlling ECM homeostasis by regulating MMP-1 and MMP-9 levels.

Objective— Mutations affecting contractile-related proteins in the ECM (extracellular matrix), microfibrils, or vascular smooth muscle cells can predispose the aorta to aneurysms. We reported previously that the LRP1 (low-density lipoprotein receptor–related protein 1) maintains vessel wall integrity, and smLRP1 −/− mice exhibited aortic dilatation. The current study focused on defining the mechanisms by which LRP1 regulates vessel wall function and integrity. Approach and Results— Isometric contraction assays demonstrated that vasoreactivity of LRP1-deficient aortic rings was significantly attenuated when stimulated with vasoconstrictors, including phenylephrine, thromboxane receptor agonist U-46619, increased potassium, and L-type Ca 2+ channel ligand FPL-64176. Quantitative proteomics revealed proteins involved in actin polymerization and contraction were significantly downregulated in aortas of smLRP1 −/− mice. However, studies with calyculin A indicated that although aortic muscle from smLRP1 −/− mice can contract in response to calyculin A, a role for LRP1 in regulating the contractile machinery is not revealed. Furthermore, intracellular calcium imaging experiments identified defects in calcium release in response to a RyR (ryanodine receptor) agonist in smLRP1 −/− aortic rings and cultured vascular smooth muscle cells. Conclusions— These results identify a critical role for LRP1 in modulating vascular smooth muscle cell contraction by regulating calcium signaling events that potentially protect against aneurysm development.

Aortic aneurysms represent a significant clinical problem as they largely go undetected until a rupture occurs. Currently, an understanding of mechanisms leading to aneurysm formation is limited. Numerous studies clearly indicate that vascular smooth muscle cells play a major role in the development and response of the vasculature to hemodynamic changes and defects in these responses can lead to aneurysm formation. The LDL receptor-related protein 1 (LRP1) is major smooth muscle cell receptor that has the capacity to mediate the endocytosis of numerous ligands and to initiate and regulate signaling pathways. Genetic evidence in humans and mouse models reveal a critical role for LRP1 in maintaining the integrity of the vasculature. Understanding the mechanisms by which this is accomplished represents an important area of research, and likely involves LRP1’s ability to regulate levels of proteases known to degrade the extracellular matrix as well as its ability to modulate signaling events.

Objective- Mutations affecting contractile-related proteins in the ECM (extracellular matrix), microfibrils, or vascular smooth muscle cells can predispose the aorta to aneurysms. We reported previously that the LRP1 (low-density lipoprotein receptor-related protein 1) maintains vessel wall integrity, and smLRP1

Objective: Mutations affecting proteins in the extracellular matrix (ECM), microfibrils, or vascular smooth muscle cells (VSMCs) that impact contractility can predispose individuals to thoracic aortic aneurysms. We reported previously that the low-density lipoprotein receptor-related protein 1 (LRP1) maintains vessel wall integrity, and smooth muscle LRP1-deficient ( smLRP1 -/- ) mice exhibited aortic dilatation. The current study focused on the descending thoracic aorta (DTA) and examined the role of LRP1 in VSMC contractility and its potential effect on the vascular ECM. Approach and Results: LRP1-deficient VSMCs exhibited a synthetic phenotype characterized by higher proliferation rates and an increase in synthetic organelles, mitochondria, multivesicular bodies, and macropinocytotic vesicles. LRP1-deficient VSMCs also displayed changes in their microfilament and actin structure that result in an inadequate interaction with the ECM. Quantitative proteomics identified proteins involved in actin polymerization and contraction that were downregulated significantly in the DTA of smLRP1 -/- mice. Further analysis by qRT-PCR revealed attenuated mRNA levels for α-1D adrenergic receptor ( adra1d ) and calcium voltage-gated channel subunit α1 C ( cacna1c ) in smLRP1 -/- aortas. Isometric contraction assays confirmed aberrant contraction of smLRP1 -/- aortic rings when stimulated with vasoconstrictors. Furthermore, intracellular calcium imaging identified defects in response to a ryanodine receptor agonist in smLRP1 -/- aortic rings. Conclusions: These results suggest that LRP1 is required for maintaining the VSMC contractile phenotype and identifies a novel role for LRP1 in calcium homeostasis that potentially protects against aneurysm development.

Hepatic inflammation is associated with the development of insulin resistance, which can perpetuate the disease state and may increase the risk of metabolic syndrome and diabetes. Despite recent advances, mechanisms linking hepatic inflammation and insulin resistance are still unclear. The low-density lipoprotein receptor-related protein 1 (LRP1) is a large endocytic and signaling receptor that is highly expressed in macrophages, adipocytes, hepatocytes, and vascular smooth muscle cells. To investigate the potential role of macrophage LRP1 in hepatic inflammation and insulin resistance, we conducted experiments using macrophage-specific LRP1-deficient mice (macLRP1−/−) generated on a low-density lipoprotein receptor knockout (LDLR−/−) background and fed a Western diet. LDLR−/−; macLRP1−/− mice gained less body weight and had improved glucose tolerance compared to LDLR−/− mice. Livers from LDLR−/−; macLRP1−/− mice displayed lower levels of gene expression for several inflammatory cytokines, including Ccl3, Ccl4, Ccl8, Ccr1, Ccr2, Cxcl9, and Tnf, and reduced phosphorylation of GSK3α and p38 MAPK proteins. Furthermore, LRP1-deficient peritoneal macrophages displayed altered cholesterol metabolism. Finally, circulating levels of sFRP-5, a potent anti-inflammatory adipokine that functions as a decoy receptor for Wnt5a, were elevated in LDLR−/−; macLRP1−/− mice. Surface plasmon resonance experiments revealed that sFRP-5 is a novel high affinity ligand for LRP1, revealing that LRP1 regulates levels of this inhibitor of Wnt5a-mediated signaling. Collectively, our results suggest that LRP1 expression in macrophages promotes hepatic inflammation and the development of glucose intolerance and insulin resistance by modulating Wnt signaling.

Low-density lipoprotein receptor–related protein-1 (LRP1) is a large endocytic and signaling receptor that is widely expressed. In the liver, LRP1 plays an important role in regulating the plasma levels of blood coagulation factor VIII (fVIII) by mediating its uptake and subsequent degradation. fVIII is a key plasma protein that is deficient in hemophilia A and circulates in complex with von Willebrand factor. Because von Willebrand factor blocks binding of fVIII to LRP1, questions remain on the molecular mechanisms by which LRP1 removes fVIII from the circulation. LRP1 also regulates cell surface levels of tissue factor, a component of the extrinsic blood coagulation pathway. This occurs when tissue factor pathway inhibitor bridges the fVII/tissue factor complex to LRP1, resulting in rapid LRP1-mediated internalization and downregulation of coagulant activity. In the vasculature LRP1 also plays protective role from the development of aneurysms. Mice in which the lrp1 gene is selectively deleted in vascular smooth muscle cells develop a phenotype similar to the progression of aneurysm formation in human patient, revealing that these mice are ideal for investigating molecular mechanisms associated with aneurysm formation. Studies suggest that LRP1 protects against elastin fiber fragmentation by reducing excess protease activity in the vessel wall. These proteases include high-temperature requirement factor A1, matrix metalloproteinase 2, matrix metalloproteinase-9, and membrane associated type 1-matrix metalloproteinase. In addition, LRP1 regulates matrix deposition, in part, by modulating levels of connective tissue growth factor. Defining pathways modulated by LRP1 that lead to aneurysm formation and defining its role in thrombosis may allow for more effective intervention in patients.

The metabolic syndrome is an escalating worldwide public health concern. Defined by a combination of physiological, metabolic, and biochemical factors, the metabolic syndrome is used as a clinical guideline to identify individuals with a higher risk for type 2 diabetes and cardiovascular disease. Although risk factors for type 2 diabetes and cardiovascular disease have been known for decades, the molecular mechanisms involved in the pathophysiology of these diseases and their interrelationship remain unclear. The LDL receptor-related protein 1 (LRP1) is a large endocytic and signaling receptor that is widely expressed in several tissues. As a member of the LDL receptor family, LRP1 is involved in the clearance of chylomicron remnants from the circulation and has been demonstrated to be atheroprotective. Recently, studies have shown that LRP1 is involved in insulin receptor trafficking and regulation and glucose metabolism. This review summarizes the role of tissue-specific LRP1 in insulin signaling and its potential role as a link between lipoprotein and glucose metabolism in diabetes.

The authors issue the following correction in order to cite and discuss previously published in vitro studies on the role of LRP1 that are relevant to this article. The sentence in the Abstract should read “To date, several macrophage receptors have been identified that contribute to the uptake of modified forms of lipoproteins leading to foam cell formation, but the in vivo contribution of the LDL receptor-related protein 1 (LRP1) to this process is not known.” In addition, the following paragraphs should be added to the Discussion: In the current study it is not clear what form of lipoprotein is recognized by macrophages in vivo. The potential of LRP1 to mediate the uptake of lipoprotein particles was first suggested in cell-based studies demonstrating that LRP1 mediates the uptake of cholesterol esters derived from apolipoprotein E enriched β-VLDL lipoprotein particles [2]. This early study established two important principals regarding the role of LRP1 in mediating lipoprotein catabolism. First, the study demonstrated a requirement for enrichment of the β-VLDL particles with apolipoprotein E in order to be recognized by LRP1 which led to the sequestration model for LRP1 mediated hepatic uptake of lipoproteins [3]. The in vivo role of LRP1 in chylomicron remnant metabolism was firmly established in 1998 when Rohlmann et al. [4] used a genetic approach to reveal LRP1's role as a chylomicron receptor. A second observation from early studies was the finding that unlike the LDL receptor, LRP1 levels are not reduced when cells are incubated with excess hydroxycholesterol [2]. These results suggested that LRP1-mediated uptake of lipoproteins could lead to foam cell formation, and indeed this was demonstrated to be the case when human monocyte-derived macrophages or vascular smooth muscle cells were incubated with aggregated LDL[5,6]. However, it is highly unlikely that aggregated LDL represents the physiological ligand for LRP1 in the macLRP1-/- / LDLR-/- mice fed a Western diet, since we observed accumulation of triglyceride-rich VLDL particles in the plasma of these mice. This result suggests that some form of VLDL particle is the physiological ligand for macrophage LRP1. Indeed, studies have suggested that LRP1-deficient macrophages are defective in mediating the internalization of VLDL particles [7], although we were unable to reproduce this observation in the current study using thioglycollate-elicited peritoneal macrophages. Very likely, the in vivo uptake of lipoproteins by LRP1 in macrophages is complex, and difficult to reproduce in cell culture experiments.