Expression profiles in surgically‐induced carotid stenosis: a combined transcriptomic and proteomic investigation

Abstract

Vascular injury aimed at stenosis removal induces local reactions often leading to restenosis. The aim of this study was a concerted transcriptomic-proteomics analysis of molecular variations in a model of rat carotid arteriotomy, to dissect the molecular pathways triggered by vascular surgical injury and to identify new potential anti-restenosis targets. RNA and proteins extracted from inbred Wistar Kyoro (WKY) rat carotids harvested 4 hrs, 48 hrs and 7 days after arteriotomy were analysed by Affymetrix rat microarrays and by bidi-mensional electrophoresis followed by liquid chromatography and tandem mass spectrometry, using as reference the RNA and the proteins extracted from uninjured rat carotids. Results were classified according to their biological function, and the most significant Kyoro Encyclopedia of Genes and Genomes (KEGG) pathways were identified. A total of 1163 mRNAs were differentially regulated in arteriotomy-injured carotids 4 hrs, 48 hrs and 7 days after injury (P < 0.0001, fold-change ≥2), while 48 spots exhibited significant changes after carotid arteriotomy (P < 0.05, fold-change ≥2). Among them, 16 spots were successfully identified and resulted to correspond to a set of 19 proteins. mRNAs were mainly involved in signal transduction, oxidative stress/inflammation and remodelling, including many new potential targets for limitation of surgically induced (re)stenosis (e.g. Arginase I, Kruppel like factors). Proteome analysis confirmed and extended the microrarray data, revealing time-dependent post-translational modifications of Hsp27, haptoglobin and contrapsin-like protease inhibitor 6, and the differential expression of proteins mainly involved in contractility. Transcriptomic and proteomic methods revealed functional categories with different preferences, related to the experimental sensitivity and to mechanisms of regulation. The comparative analysis revealed correlation between transcriptional and translational expression for 47% of identified proteins. Exceptions from this correlation confirm the complementarities of these approaches.

Stem Cell Therapy for Arterial Restenosis: Potential Parameters Contributing to the Success of Bone Marrow-Derived Mesenchymal Stromal Cells

“…In addition, internal and external elastic lamina and media disruptions at the injury site were evident, often with media substitution by fibrotic tissue. Our previous gene expression microarray data obtained in this model [12] revealed the early and marked upregulation of inflammation-related genes, thus supporting the expected picture of inflammatory cell infiltration in the acute phase after arteriotomy (mainly at 4 h and 3 days after arteriotomy).…”

“…TLRs are known to link innate and adaptive immunity. The activation and the role of TLR2 and TLR4 in inflammation and neointima proliferation associated to stenosis progression after vascular injury has already been highlighted by other studies [12][13][14]. The decrease of mRNAs coding for TLR2 and TLR4 in injured carotids from MSC-treated rats in comparison to DMEM-treated rats (Fig.…”

“…Nonetheless, it has also been reported that MSCs do not produce SDF-1α but they consume it in vitro [33]. Most importantly, SDF-1α constitutive expression is upregulated in injured arteries [12,34,35], playing a role in the recruitment of stem cells for tissue repair and in inflammation. In agreement with this observation, the ELISA assay revealed a significant increase of SDF-1α in the serum of rats at 7 days after carotid arteriotomy.…”

The Polyamine Pathway as a Potential Target for Vascular Diseases: Focus on Restenosis

“…Innovative strategies, e.g. based on local gene delivery, targeted inhibition of gene expression, or stem cell-mediated repair of injured vessels [36,37] have been successfully tested in animal models of vascular stenosis. Additional studies will be necessary to verify the effectiveness of these strategies also in patients.…”

“…Microarraybased studies of gene expression in a rat model of carotid arteriotomy revealed that vascular injury induced an acutely increased expression of genes involved in the polyamine metabolic pathway, including ODC, SSAT, AdoMetDC and arginase I [36].…”