Modulation of in vitro angiogenesis in a three-dimensional spheroidal coculture model for bone tissue engineering
Modulation of in vitro angiogenesis in a three-dimensional spheroidal coculture model for bone tissue engineering
复制标题
DOI：
10.1089/ten.2004.10.1536
复制
发表时间：
2004-09-01
影响因子：
--
通讯作者：
Kneser, U
中科院分区：
文献类型：
--
作者：
Wenger, A;Stahl, A;Kneser, U
骨替代物组织工程的主要挑战之一仍然是移植物的血管化。我们开发了一种基于三维胶原蛋白的共培养系统，用于在体外评估人内皮细胞（hEC）和人成骨细胞（hOB）之间的相互作用。人脐静脉内皮细胞 (HUVEC) 生长为三维多细胞球体，并接种在胶原基质中，以评估球体的发芽，即类似于早期毛细血管的管状结构的形成。通过将 hOB 掺入 EC 球体中，建立 hOB 和 HUVEC 之间的直接细胞接触，从而形成异质球体。通过免疫组织化学整体染色技术和共焦激光显微镜评估胶球体的空间组织和芽的构型。通过数字成像平面测量法定量分析球体的累积发芽长度。在此模型中，HUVEC 和 hOB 形成具有不同空间组织的异质球体。在异质 HUVEC/hOB 球状体中，HUVEC 球状体在血管内皮生长因子和碱性成纤维细胞生长因子的血管生成刺激下形成管状结构的能力受到抑制。本研究中引入的模型系统可能有助于评估骨形成过程中调节血管生成的机制，并进一步研究异型细胞间相互作用抑制内皮管形成的机制，以应用于骨组织工程。
One of the major challenges in tissue engineering of bone substitutes remains vascularization of the transplant. We have developed a three-dimensional collagen-based coculture system to assess interactions between human endothelial cells (hECs) and human osteoblasts (hOBs) in vitro. Human umbilical vein endothelial cells (HUVECs) were grown as three-dimensional multicellular spheroids and seeded in a collagen matrix to assess sprouting of the spheroids, that is, formation of tubelike structures resembling early capillaries. Direct cell contact between hOBs and HUVECs was established by incorporating hOBs into the EC spheroids, thus forming heterogeneous cospheroids. Spatial organization of cospheroids and sprout configuration were assessed by immunohistochemical wholemount staining techniques and confocal laser microscopy. Cumulative sprout length of spheroids was quantitatively analyzed by digital imaging planimetry. In this model HUVECs and hOBs formed heterogeneous cospheroids with distinct spatial organization. The ability of HUVEC spheroids to form tubelike structures on angiogenic stimulation with vascular endothelial growth factor and basic fibroblast growth factor was suppressed in heterogeneous HUVEC/hOB cospheroids. The model system introduced in this study may be useful to assess the mechanisms involved in regulating angiogenesis during bone formation and to further investigate the mechanisms by which heterotypic cell-cell interactions inhibit endothelial tube formation for applications in bone tissue engineering.