Porcine endothelial cells cocultured with smooth muscle cells became procoagulant in vitro.
Abstract
Endothelial cell (EC) seeding represents a promising approach to provide a nonthrombogenic
surface on vascular grafts. In this study, we used a porcine EC/smooth muscle cell
(SMC) coculture model that was previously developed to examine the efficacy of EC
seeding. Expression of tissue factor (TF), a primary initiator in the coagulation
cascade, and TF activity were used as indicators of thrombogenicity. Using immunostaining,
primary cultures of porcine EC showed a low level of TF expression, but a highly heterogeneous
distribution pattern with 14% of ECs expressing TF. Quiescent primary cultures of
porcine SMCs displayed a high level of TF expression and a uniform pattern of staining.
When we used a two-stage amidolytic assay, TF activity of ECs cultured alone was very
low, whereas that of SMCs was high. ECs cocultured with SMCs initially showed low
TF activity, but TF activity of cocultures increased significantly 7-8 days after
EC seeding. The increased TF activity was not due to the activation of nuclear factor
kappa-B on ECs and SMCs, as immunostaining for p65 indicated that nuclear factor kappa-B
was localized in the cytoplasm in an inactive form in both ECs and SMCs. Rather, increased
TF activity appeared to be due to the elevated reactive oxygen species levels and
contraction of the coculture, thereby compromising the integrity of EC monolayer and
exposing TF on SMCs. The incubation of cocultures with N-acetyl-cysteine (2 mM), an
antioxidant, inhibited contraction, suggesting involvement of reactive oxygen species
in regulating the contraction. The results obtained from this study provide useful
information for understanding thrombosis in tissue-engineered vascular grafts.
Type
Journal articleSubject
AnimalsCoculture Techniques
Cytoplasm
Endothelial Cells
Myocytes, Smooth Muscle
Reactive Oxygen Species
Swine
Thromboplastin
Tissue Engineering
Transcription Factor RelA
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https://hdl.handle.net/10161/3327Published Version (Please cite this version)
10.1089/ten.TEA.2009.0448Publication Info
Pang, Zhengyu; Niklason, Laura E; & Truskey, George A (2010). Porcine endothelial cells cocultured with smooth muscle cells became procoagulant
in vitro. Tissue Eng Part A, 16(6). pp. 1835-1844. 10.1089/ten.TEA.2009.0448. Retrieved from https://hdl.handle.net/10161/3327.This is constructed from limited available data and may be imprecise. To cite this
article, please review & use the official citation provided by the journal.
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Show full item recordScholars@Duke
George A. Truskey
R. Eugene and Susie E. Goodson Distinguished Professor of Biomedical Engineering
My research interests focus upon the effect of physical forces on the function of
vascular cells and skeletal muscle, cell adhesion, and the design of engineered tissues.
Current research projects examine the effect of endothelial cell senescence upon
permeability to macromolecules and the response to fluid shear stress, the development
of microphysiological blood vessels and muscles for evaluation of drug toxicity and
the design of engineered endothelialized blood vessels and skelet

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