Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winson-Salem, NC, USA.
J Vasc Surg. 2010 Jan;51(1):174-83. doi: 10.1016/j.jvs.2009.08.074.
A strategy in minimizing thrombotic events of vascular constructs is to seed the luminal surface with autologous endothelial cells (ECs). The task of seeding ECs can be achieved via bioreactors, which induce mechanical forces (shear stress, strain, pressure) onto the ECs. Although bioreactors can achieve a confluent layer of ECs in vitro, their acute response to blood remains unclear. Moreover, the necessary mechanical conditions that will increase EC adhesion and function remain unclear. We hypothesize that preconditioning seeded endothelium under physiological flow will enhance their retention and function.
To determine the role of varying preconditioning protocols on seeded ECs in vitro and in vivo.
Scaffolds derived from decelluarized arteries seeded with autologous ECs were preconditioned for 9 days. Three specific protocols, low steady shear stress (SS), high SS, and cyclic SS were investigated. After preconditioning, the seeded grafts were exposed to 15 minutes of blood via an ex vivo arteriovenous shunt model or alternately an in vivo arteriovenous bypass graft model.
The shunt model demonstrated ECs remained intact for all conditions. In the arteriovenous bypass model, only the cyclic preconditioned grafts remained intact, maintained morphology, and resisted the attachment of circulating blood elements such as platelets, red blood cells, and leukocytes. Western blotting analysis demonstrated an increase in the protein expression of eNOS and prostaglandin I synthase for the cyclic high shear stress-conditioned cells relative to cells conditioned with high shear stress alone.
Cyclic preconditioning has been shown here to increase the ECs ability to resist blood flow-induced shear stress and the attachment of circulating blood elements, key attributes in minimizing thrombotic events. These studies may ultimately establish protocols for the formation of a more durable endothelial monolayer that may be useful in the context of small vessel arterial reconstruction.
为了最大限度地减少血管构建物的血栓事件，一种策略是在腔表面种植自体内皮细胞（ECs）。种植 ECs 的任务可以通过生物反应器来完成，生物反应器会对 ECs 施加机械力（剪切应力、应变、压力）。虽然生物反应器可以在体外实现 EC 层的融合，但它们对血液的急性反应尚不清楚。此外，增加 EC 黏附和功能的必要机械条件尚不清楚。我们假设在生理流动下对种植的内皮细胞进行预处理可以增强其保留和功能。
确定不同预处理方案对体外和体内种植 ECs 的作用。
从脱细胞动脉中提取的支架中种植自体 ECs，进行 9 天的预处理。研究了三种特定的方案，低稳态剪切应力（SS）、高 SS 和循环 SS。预处理后，通过体外动静脉分流模型或体内动静脉旁路移植模型将接种的移植物暴露于 15 分钟的血液中。
分流模型显示所有条件下 ECs 均保持完整。在动静脉旁路模型中，只有循环预处理的移植物保持完整，保持形态，并抵抗循环血液成分（如血小板、红细胞和白细胞）的附着。Western blot 分析表明，与仅接受高剪切应力预处理的细胞相比，循环高剪切应力条件下的细胞中 eNOS 和前列环素 I 合酶的蛋白表达增加。
这里已经证明循环预处理可以提高 ECs 抵抗血流诱导的剪切应力和循环血液成分附着的能力，这是减少血栓事件的关键属性。这些研究最终可能会建立形成更持久的内皮单层的方案，这可能在小血管动脉重建的背景下有用。
