Uzarski Joseph S, Cores Jhon, McFetridge Peter S
J. Crayton Pruitt Family Department of Biomedical Engineering, University of Florida , Gainesville, Florida.
Tissue Eng Part C Methods. 2015 Nov;21(11):1125-34. doi: 10.1089/ten.TEC.2015.0110. Epub 2015 Jun 24.
The lack of a functional endothelium on small-diameter vascular grafts leads to intimal hyperplasia and thrombotic occlusion. Shear stress conditioning through controlled hydrodynamics within in vitro perfusion bioreactors has shown promise as a mechanism to drive endothelial cell (EC) phenotype from an activated, pro-inflammatory wound state toward a quiescent functional state that inhibits responses that lead to occlusive failure. As part of an overall design strategy to engineer functional vascular grafts, we present a novel two-phase shear conditioning approach to improve graft endothelialization. Axial rotation was first used to seed uniform EC monolayers onto the lumenal surface of decellularized scaffolds derived from the human umbilical vein. Using computer-controlled perfusion circuits, a flow-ramping paradigm was applied to adapt endothelia to arterial levels of fluid shear stress and pressure without graft denudation. The effects of constant pulse frequencies (CF) on EC quiescence were then compared with pulse frequencies modeled from temporal fluctuations in blood flow observed in vivo, termed physiologically modeled pulse dynamics (PMPD). Constructs exposed to PMPD for 72 h expressed a more functional transcriptional profile, lower metabolic activity (39.8% ± 8.4% vs. 62.5% ± 11.5% reduction, p = 0.012), and higher nitric oxide production (80.42 ± 23.93 vs. 48.75 ± 6.93 nmol/10(5) cells, p = 0.028) than those exposed to CF. By manipulating in vitro flow conditions to mimic natural physiology, endothelialized vascular grafts can be stimulated to express a nonactivated phenotype that would better inhibit peripheral cell adhesion and smooth muscle cell hyperplasia, conditions that typically lead to occlusive failure. Development of robust, functional endothelia on vascular grafts by modulation of environmental conditions within perfusion bioreactors may ultimately improve clinical outcomes in vascular bypass grafting.
小口径血管移植物缺乏功能性内皮会导致内膜增生和血栓闭塞。通过体外灌注生物反应器内的可控流体动力学进行剪切应力调节，已显示出有望作为一种机制，促使内皮细胞（EC）表型从活化的促炎伤口状态转变为静止的功能状态，从而抑制导致闭塞性失败的反应。作为构建功能性血管移植物整体设计策略的一部分，我们提出了一种新颖的两阶段剪切调节方法来改善移植物内皮化。首先使用轴向旋转将均匀的内皮细胞单层接种到源自人脐静脉的脱细胞支架的管腔表面。使用计算机控制的灌注回路，应用流量递增模式使内皮适应动脉水平的流体剪切应力和压力，而不会导致移植物剥脱。然后将恒定脉冲频率（CF）对内皮细胞静止的影响与根据体内观察到的血流时间波动建模的脉冲频率进行比较，后者称为生理建模脉冲动力学（PMPD）。暴露于PMPD 72小时的构建体表现出更具功能性的转录谱、更低的代谢活性（降低39.8%±8.4% vs. 62.5%±11.5%，p = 0.012）以及比暴露于CF的构建体更高的一氧化氮产生量（80.42±23.93 vs. 48.75±6.93 nmol/10(5)细胞，p = 0.028）。通过操纵体外流动条件以模拟自然生理状态，可以刺激内皮化血管移植物表达非活化表型，从而更好地抑制外周细胞粘附和平滑肌细胞增生，这些情况通常会导致闭塞性失败。通过调节灌注生物反应器内的环境条件在血管移植物上培养强健的功能性内皮，最终可能改善血管旁路移植的临床结果。
