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Article
Thrombomodulin Induces a Quiescent Phenotype and Inhibits Migration in Vascular Smooth Muscle Cells In Vitro
Annals of Vascular Surgery
(2016)
Abstract
Background: Loss of critical endothelial cell function and subsequent vascular smooth muscle cell (VSMC) migration is central to the pathology of injury-induced neointimal hyperplasia and recurrent stenosis. Thrombomodulin (TM), well known for its function as an endothelial surface anticoagulant, may have an unknown direct effect on VSMC physiology that would be lost after injury. Here, we examined a novel effect of TM on VSMC by testing the hypothesis that direct application of TM induces favorable changes to the morphology of VSMC and inhibits their migration.
Methods: Primary human VSMC were harvested using the explant technique and used in early passage (1e4) for all experiments. Laser-scanning confocal fluorescent imaging was performed to assess the effect of soluble TM on VSMC morphology. In vitro, migration of VSMC was measured using: (1) a 4-hr modified Boyden chemotaxis assay and (2) a 24-hr electric cellsubstrate impedance sensing injury migration assay. Migration experiments were conducted with VSMC exposed to increasing doses of soluble recombinant TM. Recombinant thrombin served as a positive control and serum-free media as a negative control for all experimentation. Data were analyzed using a Student’s t-test or repeated measures analysis of variance where appropriate (α < 0.05).
Results: VSMC exposed to TM clearly demonstrated a quiescent morphology with organized stress fibers consistent with a quiescent, differentiated, contractile phenotype; whereas, thrombin stimulation led to an activated, dedifferentiated, synthetic phenotype. VSMC demonstrated a low, baseline level of migration in unstimulated serum-free conditions. Thrombin significantly stimulated VSMC migration as expected. TM, independent of thrombin, significantly inhibited baseline VSMC migration in a dose-response fashion. The maximal inhibition was observed at (5 μg/mL) with 70% reduction (56 ± 1.7 vs. 18 ± 3.5 cells/5 high-power fields, P = 0.0005).
Conclusions: TM has a direct effect on VSMC resulting in a quiescent, differentiated and contractile phenotype, and inhibition of migration. This effect is independent of the presence of thrombin. These findings provide new knowledge in understanding the pathophysiology of vascular injury and support a strategy focused on restoring key endothelial function to prevent intimal hyperplasia.
Disciplines
Publication Date
January, 2016
DOI
10.1016/j.avsg.2015.10.002
Citation Information
Heather M. Bass, Richard S. Beard, Byeong J. Cha, Sarah Y. Yuan, et al.. "Thrombomodulin Induces a Quiescent Phenotype and Inhibits Migration in Vascular Smooth Muscle Cells In Vitro" Annals of Vascular Surgery Vol. 30 (2016) p. 149 - 156 ISSN: 08905096 Available at: http://works.bepress.com/richard-beard/3/