Current projects
Molecular mechanism involved in the regulation of endothelin-1 synthesis in human endothelial cells
Endothelium has an important role in the maintenance of vascular homeostasis. Endothelin-1 (ET-1) is a powerful vasoconstrictor and its up-regulation has been associated with endothelial dysfunction and cardiovascular disorders, including atherosclerosis, hypertension and diabetes. The underlying mechanisms accountable for the increased ET-1 expression are not completely defined.
High glucose induces endothelin-1 expression via a functional CCAAT-enhanced binding protein β.
Lysosomal enzymes in vascular cells involved in atherosclerosis
In normal conditions, LDL particles are internalized via receptor–mediated endocytosis and delivered to the lysosomes, an important process in monitoring the cholesterol homeostasis by a negative feed-back, with the prevention of cellular lipid overload. Upon LDL modification, oxidized LDL (oxLDL) or glycated LDL (gLDL), are internalized via scavenger receptors, without a feed-back inhibition. LDL-derived cholesteryl esters as (±)9-hydroxyoctadecadienoic acid-cholesteryl ester (HODE) accumulate and leads to foam cells formation, a major component of atherosclerotic lesions. Lysosomal acid lipase (LAL) is the only enzyme involved in hydrolysis of LDL-cholesteryl esters.
We searched the effect of. oxLDL, HODE and gLDL on LAL activity and expression in endothelial cells (EC) and smooth muscle cells (SMC) and the role of LXR and PPAR-gamma. We have found that the oxLDL- or HODE-exposed EC exhibited a decrease in LAL activity and expression. In SMC, gLDL down-regulated LAL activity and mRNA. LXR agonist was the most effector in reversing the decreased EC-LAL expression induced by oxLDL or HODE, and SMC-LAL expression produced by gLDL. PPAR agonist, RSG, has a lower effect on EC-LAL expression exposed to oxLDL or HODE but not in gLDL-exposed SMC. The therapeutic role of LAL and the identification of a new marker molecule that will trigger the warning before the appearance of irreversible lesions are our future interest.
Junctional proteins at the endothelial cell-cell contact
The permeability of the tight junctions in endothelial cell is controlled among others by the tight junctional protein occludin, which is present in these structures. The occludin's function, to seal the junctional structure and control the paracellular flux of solutes, is likely to be regulated by occludin phosphorylation.
We recently observed that histamine induces relocation of occludin in venous endothelium (see figure below). Currently, we investigate the correlations between occludin and other tight junctional proteins including claudin 5 in endothelial cells in culture and rat macrovasculature (veins and arteries) by immunofluorescence microscopy and western blotting. Differences in the expression of the tight junctional proteins are expected to underlie functional variations in permeability across the vascular bed. A map of expression for tight junctional molecules in the endothelium of different origins in an animal model will contribute to our understanding of the functional significances for topological variations in the expression of tight junctional molecules. We also envisage a therapeutical approach, hoping to selectively increase the permeability of various epithelia without affecting endothelium. (project of C. Neagoe, PhD)
Occludin redistributes to the endothelial cell periphery after histamine treatment (left), immunofluorescence micrograph, scale bar: 25 μm (C. Neagoe, et al.,manuscript in preparation )
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