冠狀動脈繞道手術需要利用小管徑的血管取代物，但是由於小管徑的血管血流速度較慢，無法成功的利用人工合成大管徑血管的材料作為取代物。目前標準的小管徑血管取代物是採用自身的其他血管，但是包含病患已有其他的血管疾病、截肢、以及之前已做過置換手術等因素，限制了自身血管的使用，而組織工程技術則提供了一個製造合適的人工血管取代物的新方法。一個好的血管取代物須具備下列兩個重要條件：完整且具血液相容性的內皮層，以及用來提供機械性強度的平滑肌。在過去的研究中有相當多建構人工血管的方法，但是這些方法具有許多缺點，包含機械強度不足、以及最重要的：不具有完整的內皮細胞層。在本實驗中，我們提出了一個新的方法，利用羊膜作為骨架來建構人工血管。羊膜已經被成功的用於眼角膜移植的骨架，且不具免疫排斥性，並可以提供一層完整且具彈性的基底膜。在初步的結果中，我們從豬的主動脈分離並培養內皮細胞及平滑肌細胞，利用免疫螢光染色確定細胞的純度。我們一般的培養條件下建立了內皮細胞的生長、及細胞與細胞間的接合狀況，並已成功的將內皮細胞培養在羊膜上，並形成完整的單層構造。接著我們利用一個共同培養的系統，將內皮細胞與平滑肌細胞同時培養在一片羊膜上。接著以一根軸心將羊膜捲起，使內皮細胞在此管狀結構的最內層、平滑肌細胞在外層。最後，這個人工血管被置於一個生物反應器中，促進細胞的成熟以接近正常血管的生理狀態。由於內皮細胞可以在羊膜上穩定的生長，因此我們所建構的人工血管應該更接近正常的血管，來解決目前其他人造血管所面臨的問題。 Small-caliber blood vessel grafts are required for coronary artery bypass surgery. But the lower blood flow velocities of small-diameter vessels have led to the failure of synthetic materials that are successful for large-diameter vessel grafts. Autologous vessels are the standard for small-diameter vessel grafts now, but there are many limitations including other vascular diseases, amputation, and previous harvest. Tissue engineering provides a promising way to generate available vessel grafts. There are two important criteria for blood vessel equivalent (BVE): an integral endothelium to create a hemocompatible lining and smooth muscle cells to provide mechanical strength. In previous studies, several ways have been designed to construct tissue engineered blood vessels (TEBVs). But there are several disadvantages in each ways including inadequacy in mechanical strength and most important, no integral and functional endothelium available. Here, we propose a new method to fabricate a biological blood vessel equivalent by amniotic membrane (AM). AM is a nonimmunogenic substrate and can provide a integral, elastic basement membrane. AM has been successfully used as a scaffold in tissue engineered human corneal transplantation. In the preliminary results, porcine endothelial cells (PECs) and smooth muscle cells (PSMCs) were cultured from aorta, and the cells' purities were confirmed by immunofluorescence staining. We have established the growth and junction formation profiles of PECs under normal culture condition. Moreover, we have successfully cultured PECs as a stable monolayer on AM. To fabricate a BVE with expected structure, a co-culture system of PECs and PSMCs on AM has been performed and cells on AM were rolled with ECs at the most inner layer and SMCs at the outer layer by a mandrel. Finally, the BVE were cultured in a bioreactor to promote the maturation of endothelium and SMCs. Because ECs could preserve stable monolayer on AM, it is possible that our BVE will possess the intrinsic properties to circumvent the draw backs encountered by other models.
