Characterization of novel interactions with membrane NEU1 highlights new regulatory functions for the Elastin Receptor Complex in monocyte interaction with endothelial cells

Background Vascular aging is associated with remodeling of elastin, one of the main extracellular matrix component of the arterial wall, and production of elastin-derived peptides (EDP). These extracellular matrix degradation products have been shown to trigger biological activities through the elastin receptor complex (ERC) and data from the last decade have brought significant insights on the critical role played by its NEU1 subunit in the biological effects mediated by EDP and the ERC in vascular and metabolic diseases. Results Using a proteomic approach, we previously identified new potential interaction partners of membrane NEU1. Here, we validated the interaction between NEU1 and the β2 integrin in human monocytes and show that binding of EDP to the ERC leads to desialylation of β2 integrin through NEU1. A similar action mechanism was identified in human umbilical vein endothelial cells (HUVEC) for intercellular cell adhesion molecule-1 (ICAM-1). Importantly, these effects were associated with a significant increase in monocyte adhesion to endothelial cells and monocyte transendothelial migration. Conclusions These results demonstrate that membrane NEU1 sialidase interacts and modulates the sialylation levels of the β2 integrin and ICAM-1 through the ERC in monocytes and endothelial cells, respectively, and suggest that EDP and the ERC, through this newly identified common mode of action governed by NEU1, may be important regulators of circulating monocyte recruitment to inflamed vascular sites. Moreover, by its ability to interact with and to modulate the sialylation of key membrane glycoproteins through NEU1, new biological functions are anticipated for EDP and the ERC in elastin remodeling-associated disorders.

Structural analysis of nonapeptides derived from elastin

Elastin-derived peptides are released from the extracellular matrix remodeling by numerous proteases and seem to regulate many biological processes, notably cancer progression. The canonical elastin peptide is VGVAPG which harbors the XGXXPG consensus pattern allowing interaction with the elastin receptor complex located at the surface of cells. Besides these elastokines, another class of peptides has been identified. This group of bioactive elastin peptides presents the XGXPGXGXG consensus sequence but the reason for their bioactivity remains unexplained. In order to better understand their nature and structure-function relationships, herein we searched the current databases for this nonapeptide motif and observed that the XGXPGXGXG elastin peptides define a specific group of tandemly repeated patterns. Further, we focused on four tandemly repeated human elastin nonapeptides, i.e. AGIPGLGVG, VGVPGLGVG, AGVPGLGVG and AGVPGFGAG. These peptides were analysed by means of optical spectroscopies and molecular dynamics. UV-circular dichroism and Raman spectra are consistent with a conformational equilibrium between β-turn, β-strand and random chain secondary elements in aqueous media. This equilibrium was found to be concentration-independent. Quantitative analysis of their conformations suggested that turns corresponded to half of the total population of structural elements while the remaining half was equally distributed between β-strand and unordered chains. These distributions were confirmed by molecular dynamics simulations. Altogether, our data suggest that these peptides harbor a type II β-turn located in their central part. We hypothesize that this structural element could explain their specific bioactivity.Statement of SignificanceElastin fragmentation products, the so-called elastin peptides, may exhibit a bioactivity towards normal and tumor cells. This phenomenon depends on the sequence motif they harbor. While XGXXPG sequences bioactivity is explained by the presence of a type VIII β-turn allowing interaction with the elastin receptor complex, the structural reasons for XGXPGXGXG specific activity remain unexplained. Using data mining, we show that elastin nonapeptides define a specific class of tandemly repeated features. Further, spectroscopic and numerical simulations methods suggest the presence of a type II β-turn in their conformation. This structural element could explain their bioactivity.

Abstract 480: Aortic Microcalcification Associates With Aortic Elastin Fragmentation in Marfan Syndrome

Marfan syndrome (MFS) is a genetic connective tissue disorder, in which aortic rupture is the major cause of death. MFS patients with an aortic diameter below the advised limit for prophylactic surgery (<5cm) may unexpectedly experience an aortic dissection or rupture, despite yearly monitoring. Hence, there is a clear need for improved prognostic markers to predict such aortic events. We hypothesize that elastin fragments play a causal role in aortic calcification in MFS and that microcalcification serves as a novel marker for aortic disease severity. To address this hypothesis, we analyzed MFS patient and mouse aortas. MFS patient aortic tissue showed enhanced microcalcification in areas with extensive elastic lamina fragmentation in the media. A causal relationship between medial injury and microcalcification was revealed by studies in vascular smooth muscle cells (SMCs); elastin peptides were shown to increase the activity of the calcification marker alkaline phosphatase (ALP) and reduce the expression of the calcification inhibitor matrix gla protein (MGP) in human SMCs. In murine Fbn1 C1039G/+ MFS aortic SMCs, ALP mRNA and activity was upregulated when compared to wildtype SMCs. The elastin peptide-induced ALP activity was prevented by incubation with lactose as inhibitor of the elastin receptor complex, and a MEK1/2 kinase inhibitor, indicating downstream involvement of ERK1/2 phosphorylation. Histological analyses in MFS mice revealed macrocalcification in the aortic root, while the ascending aorta contained microcalcification, as identified with the near-infrared fluorescent bisphosphonate probe OsteoSense-800. Significantly, microcalcification correlated strongly with aortic diameter, aortic distensibility, elastin breaks and phosphorylated ERK1/2. In conclusion, microcalcification co-localizes with aortic elastin degradation in MFS aorta of man and mice, where elastin-derived peptides induce a calcification process in SMCs via the elastin receptor complex and ERK1/2 activation. We propose microcalcification as a novel imaging marker to monitor local elastin degradation and thus predict aortic events in MFS patients.