In-situ Bioengineering of Arterial Vein Grafts

The autogenous saphenous vein remains the graft of choice for both coronary (500,000 annually in the US) and peripheral (80,000 annually) arterial bypass procedures. Failure of arterial vein grafts (AVGs) remains a major problem, and patients with failed grafts will die or require re-operation. Intimal hyperplasia (IH) accounts for 20% to 40% of all AVG failures. It is believed that this adverse pathological response by AVGs is largely due to their abrupt exposure to the significantly elevated circumferential wall stress (CWS) associated with the arterial system. We believe that if an AVG is given an ample opportunity to adapt and remodel to the stresses of its new environment, cellular injury may be reduced, thus limiting the initiating mechanisms of IH. The goal of this work was to develop a new mechanical conditioning paradigm, in the form of a peri-adventitially placed, biodegradable polymer wrap, to safely and functionally "arterialize" AVGs in situ. The polymer wrap was tuned so that as it degraded over a desired period of time, the mechanical support offered by it was reduced and the vein was exposed to gradually increasing levels of CWS in situ. To investigate the effects of mechanical conditioning on AVGs, we utilized both our well established, validated ex vivo vascular perfusion system (EVPS) as well as an appropriate preclinical animal model. The "engineering" component of this bioengineering study was to enhance our EVPS capabilities. Enhancements were made in the form of rigorous mathematical modeling, via subspace system identification, and automatic feedback control, via proportional integral and derivative control, of the arterial CWS and shear stress waveform generation capabilities of the EVPS. Pairs of freshly harvested porcine internal jugular veins (PIJVs) were perfused ex vivo under several biomechanical conditions. The acute hyperplastic response of PIJVs abruptly exposed to arterial hemodynamic conditions was compared to PIJVs perfused under normal venous conditions. In an attempt to attenuate this acute hyperplastic response, an ex vivo mechanical conditioning paradigm was imposed onto the PIJVs both via manual adjustment of EVPS parameters and via an adventitially placed tuned electrospun biodegradable polymer wrap. Early markers of IH were evaluated post-perfusion, and they included vascular smooth muscle cell apoptosis, proliferation, and phenotypic modulation. Quantification of these markers via immunohistochemical techniques provided the foundation for the final stage of this work. To assess the efficacy of the tuned electrospun biodegradable polymer wrap in attenuating the development of intimal hyperplasia in AVGs, a series of preclinical studies was performed in a pig model.PIJVs abruptly exposed to arterial levels of CWS showed a significant increase in apoptosis and in the number of synthetic smooth muscle cells, as well as a decrease in proliferation. Mechanical conditioning, via both manual adjustment of the EVPS parameters and placement of the biodegradable adventitial wrap, appeared to have beneficial effects on the acute hyperplastic response of PIJVs perfused ex vivo. The beneficial effects of the adventitially placed polymer wrap was also observed in vivo, however the results did not achieve significance over unwrapped controls. Future work should be aimed at enhancing the beneficial effects of the electrospun biodegradable polymer wrap by incorporating the delivery of drugs and/or stem cells in addition to the delivery of structural support to AVGs

Age-Specific Acute Changes in Carotid-Femoral Pulse Wave Velocity With Head-up Tilt

BACKGROUND: Aortic stiffness as measured by carotid-femoral pulse wave velocity (cfPWV) is known to depend on blood pressure (BP), and this dependency may change with age. Therefore, the hydrostatic BP gradient resulting from a change in body posture may elicit a cfPWV change that is age-dependent. We aimed to analyze the relationship between BP gradient-induced by head-up body tilting-and related changes in cfPWV in individuals of varying age. METHODS: cfPWV and other hemodynamic parameters were measured in 30 healthy individuals at a head-up tilt of 0° (supine), 30°, and 60°. At each angle, the PWV gradient and resulting cfPWV were also estimated (predicted) by assuming a global nonlinear, exponential, pressure-diameter relationship characterized by a constant β0, and taking into account that (diastolic) foot-to-foot cfPWV acutely depends on diastolic BP. RESULTS: cfPWV significantly increased upon body tilting (8.0 ± 2.0 m/s supine, 9.1 ± 2.6 m/s at 30°, 9.5 ± 3.2 m/s at 60°, P for trend <0.01); a positive trend was also observed for heart rate (HR; P < 0.01). When the observed, tilt-induced cfPWV change measured by applanation tonometry was compared with that predicted from the estimated BP hydrostatic gradient, the difference in observed-vs.-predicted PWV change increased nonlinearly as a function of age (R2 for quadratic trend = 0.38, P < 0.01, P vs. linear = 0.04). This result was unaffected by HR tilt-related variations (R2 for quadratic trend = 0.37, P < 0.01, P vs. linear = 0.04). CONCLUSIONS: Under a hydrostatic pressure gradient, the pulse wave traveling along the aorta undergoes an age-related, nonlinear PWV increase exceeding the increase predicted from BP dependency

Effects of Vascular Nitric Oxide Bioactivity and Vascular Ageing on Arterial Blood Pressure and Flow Waveforms

Analysis of blood pressure and flow waveforms may lead to improved diagnosis of arterial dysfunction and disease. This thesis describes experiments to investigate the characteristic alteration of peripheral waveforms produced by stimulated release of vascular nitric oxide (an effect that is attenuated by endothelial dysfunction) and the changes that occur with age. In vivo experiments were conducted in anaesthetised rabbits and in vitro experiments employed a polyurethane model of the human aorta and its principal branches. Blood pressure, blood flow, pulse wave velocities and vessel diameter were recorded in the rabbit abdominal aorta. Equivalent recordings were obtained from the model aorta. Data were analysed in the time domain using wave intensity analysis after separation of reservoir (Windkessel) pressure and wave pressure. Effects of acetylcholine (Ach) and NG-nitro-L-arginine methyl ester (L-NAME) were investigated in vivo. Ach (which promotes endothelial production of nitric oxide) increased wave reflection, whilst L-NAME (which inhibits nitric oxide production) decreased it. These trends were opposite to the expected ones, and do not account for the established effects of nitric oxide on peripheral arterial waveforms. Further work is required to investigate these contradictions. Arterial stiffening in immature and mature rabbits was attempted by supplementing their diet with fructose. Fructose is known to form advanced glycation end-products in arterial walls and hence to stiffen arteries. Unexpectedly, fructose did not affect haemodynamic function. However, immature control rabbits had markedly reduced aortic wave reflection compared to mature control rabbits, indicating that the former have arterial impedances that are better matched for incident wave propagation. Wrapping the model aorta in Clingfilm was used to simulate age- or drug-induced stiffening of the aorta in vivo. Increased pulse pressure was observed, resembling the isolated systolic hypertension prevalent in aged populations. The model has potential for modelling haemodynamic function in health and disease

Investigation of blood flow in the superior mesenteric artery and its potential influence on atheroma and gut ischaemia.

Atherosclerosis is the underlying process in coronary heart disease leading to myocardial infarction, and in arterial damage leading to cerebrovascular accidents. It accounts for almost 50% of deaths in the western world. Atherosclerosis is characterised by the presence of fibro-lipid plaques (atheroma) within the vessel wall. Whilst the initiation and progression of atheroma are not fully understood, it is generally accepted that the time-varying haemodynamic wall shear stress (WSS) that the vessel wall is exposed to is important in determining the likelihood of development of an atherosclerotic plaque The superior mesenteric artery (SMA) is the major blood vessel feeding the small intestine; compared to other vessels of similar size, it is largely spared the effects of atherosclerosis

Magnetic Resonance Imaging of the Neonatal Cardiovascular System : Impact of Patent Ductus Arteriosus

The incidence of premature birth is increasing in absolute number and as a proportion of all births around the world. Many pathologies seen in this cohort are related to abnormal blood supply. Fetal and premature cardiovascular systems differ greatly as to maintain adequate blood flow to the developing organs in the uterine and extra-uterine environments require very different circulations. Subsequently following preterm birth the immature cardiovascular system undergoes abrupt adaptations, often resulting in the prolonged patency of the fetal shunt, ductus arteriosus. The impact of a patent ductus arteriosus (PDA) is poorly understood. However it is thought that large ductal shunt volumes may result in congestive cardiac failure and systemic hypo-­‐perfusion. Cardiac MRI has contributed greatly to the understanding of many cardiovascular diseases and congenital defects in paediatric and adult patients. Translating these imaging techniques to assess the preterm cardiovascular system requires careful optimization due to their condition, size and significantly increased heart rate. The work presented in this thesis employs multiple functional CMR techniques to investigate the preterm cardiovascular system in the presence and absence of PDA and the resultant cardiac function. A novel technique utilizing PC MRI to quantify PDA shunt volume and its impact on flow distribution is presented. Despite large shunt volumes, systemic circulation remained within normal range, although slight reduction is detectable when assessed at group level. Subsequently the impact of PDA and associated increased work load on left ventricular dimensions and function was then investigated using SSFP imaging. Results indicated that cardiac function was maintained even in the presence of large shunt volumes. Finally 4D PC sequences were employed to evaluate pulse wave velocity and flow regime within the preterm aorta, demonstrating the feasibility of hemodynamic assessment in this cohort. The findings of these studies provide insight into the impact of PDA. The reliable measurement and assessment of preterm cardiovascular system provides the potential to improve the understanding of the development and effects of certain pathologies seen in this cohort.Open Acces

Development of a Stem Cell-Based Tissue Engineered Vascular Graft

Limited autologous vascular graft availability and poor patency rates of synthetic grafts for small-diameter revascularization (e.g., coronary artery bypass, peripheral bypass, arteriovenous graft for hemodyalisis access, etc.) remain a concern in the surgical community. A tissue engineering vascular graft (TEVG), including suitable cell source, scaffold, seeding, and culture methods can potentially solve these limitations. Muscle-derived stem cells (MDSCs) are multipotent cells, with long-term proliferation and self-renewal capabilities, which represent a valid candidate for vascular tissue engineering applications due to their plasticity/heterogeneity. The poly(ester urethane) urea (PEUU) is also an attractive potential candidate for use as a TEVG due to its elasticity and tunable mechanical and degradation properties. We hypothesized that a novel scaffold optimally seeded with stem cells, acutely cultured and stimulated in vitro, and ultimately implanted in vivo will remodel into a functional vascular tissue. To test this hypothesis, we developed an innovative, multidisciplinary framework to fabricate and culture a TEVG in a timeframe compatible with clinical practice. In this approach, MDSCs were incorporated into a newly-designed and characterized PEUU-based scaffold via a novel seeding device, which was tested quantitatively for cell seeding uniformity and viability. The seeded TEVGs were acutely cultured in dynamic conditions and assessed for cell phenotype, proliferation, and spreading. The conduits were then implanted systemically in a small and a large animal model and assessed, at different time points, for patency rate, remodeling, and cellular engraftment and phenotype. The seeding technology demonstrated a rapid, efficient, reproducible, and quantitatively uniform seeding without affecting cell viability. The PEUU scaffold that was developed is suitable for arterial applications, exhibiting appropriate strength, compliance, and suture retention properties. The dynamic culture resulted in cell proliferation and spreading within the 3D scaffold environment. Rat preclinical studies suggested a role of the seeded MDSCs in the maintenance of patency and in the remodeling of the TEVG toward a native-like structure. Pig studies were inconclusive due to a poor pre-implantation cell density. Future work should address this and other issues encountered during the large animal study, and should test longer time points in both models. Finally, this approach might benefit from a more readily available cell source such as the bone marrow