Congzhu Shi

MicroRNA-10a regulation of proinflammatory phenotype in athero-susceptible endothelium in vivo and in vitro

A chronic proinflammatory state precedes pathological change in arterial endothelial cells located within regions of susceptibility to atherosclerosis. The potential contributions of regulatory microRNAs to this disequilibrium were investigated by artery site-specific profiling in normal adult swine. Expression of endothelial microRNA10a (miR-10a) was lower in the athero-susceptible regions of the inner aortic arch and aorto-renal branches than elsewhere. Expression of Homeobox A1 (HOXA1), a known miR-10a target, was up-regulated in the same locations. Endothelial transcriptome microarray analysis of miR-10a knockdown in cultured human aortic endothelial cells (HAEC) identified IκB/NF-κB–mediated inflammation as the top category of up-regulated biological processes. Phosphorylation of IκBα, a prerequisite for IκBα proteolysis and NF-κB activation, was significantly up-regulated in miR-10a knockdown HAEC and was accompanied by increased nuclear expression of NF-κB p65. The inflammatory biomarkers monocyte chemotactic protein 1 (MCP-1), IL-6, IL-8, vascular cell adhesion molecule 1 (VCAM-1), and E-selectin were elevated following miR-10a knockdown. Conversely, knockin of miR-10a (a conservative 25-fold increase) inhibited the basal expression of VCAM-1 and E-selectin in HAEC. Two key regulators of IκBα degradation—mitogen-activated kinase kinase kinase 7 (MAP3K7; TAK1) and β-transducin repeat-containing gene (βTRC)—contain a highly conserved miR-10a binding site in the 3′ UTR. Both molecules were up-regulated by miR-10a knockdown and suppressed by miR-10a knockin, and evidence of direct miR-10a binding to the 3′ UTR was demonstrated by luciferase assay. Comparative expression studies of endothelium located in athero-susceptible aortic arch and athero-protected descending thoracic aorta identified significantly up-regulated MAP3K7, βTRC, phopho-IκBα, and nuclear p65 expression suggesting that the differential expression of miR-10a contributes to the regulation of proinflammatory endothelial phenotypes in athero-susceptible regions in vivo.

Hemodynamics and the focal origin of atherosclerosis: a spatial approach to endothelial structure, gene expression, and function.

Abstract: Atherosclerosis originates at predictable focal and regional sites that are associated with complex flow disturbances and flow separations in large arteries. The spatial relationships associated with hemodynamic shear stress forces acting on the endothelial monolayer are considered in experiments that model regions susceptible to atherosclerosis (flow disturbance) and resistant to atherosclerosis (undisturbed flow). Flow disturbance in vitro induced differential expression at the single gene level as illustrated for the intercellular communication gene and protein, connexin 43. Transcription profiles of individual endothelial cells isolated from both disturbed and undisturbed flow regions exhibited more expression heterogeneity in disturbed than in undisturbed flow. We propose that within highly heterogeneous populations of endothelial cells located in disturbed flow regions, proatherosclerotic gene expression may occur within the range of expression profiles induced by the local hemodynamics. These may be sites of initiation of focal atherosclerosis. Mechanisms are proposed to account for heterogeneous endothelial responses to shear stress by reference to the decentralized model of endothelial mechano‐transduction. Length scales ranging from centimeters to nanometers are useful in describing regional, single cell, and intracellular mechanotransduction mechanisms.

Prelesional arterial endothelial phenotypes in hypercholesterolemia: universal ABCA1 upregulation contrasts with region-specific gene expression in vivo

Atherosclerosis originates as focal arterial lesions having a predictable distribution to regions of bifurcations, branches, and inner curvatures where blood flow characteristics are complex. Distinct endothelial phenotypes correlate with regional hemodynamics. We propose that systemic risk factors modify regional endothelial phenotype to influence focal susceptibility to atherosclerosis. Transcript profiles of freshly isolated endothelial cells from three atherosusceptible and three atheroprotected arterial regions in adult swine were analyzed to determine the initial prelesional effects of hypercholesterolemia on endothelial phenotypes in vivo. Cholesterol efflux transporter ATP-binding cassette transporter A1 (ABCA1) was upregulated at all sites in response to short-term high-fat diet. Proinflammatory and antioxidative endothelial gene expression profiles were induced in atherosusceptible and atheroprotected regions, respectively. However, markers for endoplasmic reticulum stress, a signature of susceptible endothelial phenotype, were not further enhanced by brief hypercholesterolemia. Both region-specific and ubiquitous (ABCA1) phenotype changes were identified as early prelesional responses of the endothelium to hypercholesterolemia.