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EVALUATION OF THE IMPORTANCE OF SMOOTH MUSCLE TO THE DEVELOPMENT OF FUNCTIONAL TISSUE ENGINEERED BLOOD VESSELS

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title
EVALUATION OF THE IMPORTANCE OF SMOOTH MUSCLE TO THE DEVELOPMENT OF FUNCTIONAL TISSUE ENGINEERED BLOOD VESSELS
author
Machingal, Masood A.
abstract
Tissue engineered blood vessels (TBEV) represent an attractive method for developing arterial substitutes to address limitations in synthetic scaffolds and lack of suitable autologous vessels. Most studies to date have necessarily focused on incorporating endothelial cells (EC) for TEBV development, with much less attention paid to the potentially important role of smooth muscle cells (SMC). SMC are the predominant cell type in the vessel wall of native vasculature, and as such, they play an essential role in modulating vascular responses to a wide variety of mechanical and biochemical stimuli. Our overarching hypothesis is that incorporation of SMC in TEBV will accelerate tissue formation and remodeling and enhance physiological function following implantation. To this hypothesis we have developed three types of TEBV grafts: 1) acellular arterial scaffolds (AAS) seeded with EC only (i.e., ecTEBV), 2) AAS seeded with both EC and SMC, or dual seeded TEBV (i.e., dsTEBV), and 3) AAS seeded with EC and an enhanced SMC layer, or dual enhanced seeded TEBV (desTEBV). Following bioreactor preconditioning, TEBV grafts were implanted either as arterio-venous fistula (AVF) or as an arterial interposition (AI) graft in an ovine model. Two months post-implantation, retrieved AVF grafts demonstrated no signs of infection, aneurysm or dilation over time indicating their potential clinical utility. During this time frame all grafts remodeled to achieve mechanical properties similar to native vessels. Physiological studies showed desTEBV grafts generated significantly greater contraction to depolarization with 60 mM KCl, as well as receptor-mediated activation with physiologically relevant vasoactive agents (10 µM Serotonin and 100 nM Endothelin-1) than ecTEBV. Additionally, greater α-actin positive cellularity was observed in desTEBV than ecTEBV supporting the physiological observations. In a second set of in vivo studies, the role of SMC in TEBV over a longer maturation period in vivo (four months) was assessed in AI model using ecTEBV and dsTEBV. We demonstrated that dsTEBV generated contractile responses to physiologically relevant vascular contractile agents correlated with increases in both α-actin and smooth muscle myosin immunostaining in the vessel wall. Taken together, seeding SMC significantly enhanced the structural remodeling and physiological properties in comparison to those grafts without SMC. However, the contractile responses were significantly lower in comparison to native vessels possibly due to the limited migration of SMC into medial graft wall. In order to improve the SMC migration into scaffold wall, we developed a more porous acellular venous scaffold (AVS) and evaluated the SMC migration in comparison with the AAS scaffold. SMC after two weeks of bioreactor preconditioning attached, proliferated and migrated into AVS scaffold upto half way into medial layer, farthest migration of SMC as far as we know, achieved using acellular vascular scaffolds in vitro. Significantly greater SMC migration (20 fold, p<0.05) and porosity (2.8 fold, p<0.05) was observed in AVS scaffolds than AAS scaffolds, without a significant difference in circumferential biomechanical properties of AAS scaffolds indicating the potential utility of this scaffolding system. To summarize, these observations unequivocally confirmed the importance of SMC seeding in structural remodeling, enhancing TEBV phenotype and improving physiological responses following implantation in vivo. While further enhancements are required to improve cell seeding, scaffold development and bioreactor preconditioning to develop TEBV with SMC in the medial layers, this study forms a step towards developing TEBV capable of remodeling into a more native like architecture and physiological functional properties upon implantation.
subject
bioreactor
end stage renal disease
regenerative medicine
smooth muscle cells
vascular access
vascular tissue engineering
contributor
Christ, George J (committee chair)
Soker, Shay (committee member)
Geary, Randolph L (committee member)
Goldstein, Aaron S (committee member)
Van Dyke, Mark E (committee member)
date
2011-07-14T20:36:04Z (accessioned)
2011 (issued)
degree
Biomedical Engineering (discipline)
identifier
http://hdl.handle.net/10339/33480 (uri)
language
en (iso)
publisher
Wake Forest University
type
Dissertation

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