Bone tissue engineers are facing a daunting challenge when attempting to fabricate bigger constructs intended for use in the treatment of large bone defects, which is the vascularization of the graft. Cell-based approaches and, in particular, the use of in vitro coculture of human umbilical vein endothelial cells (HUVECs) and human mesenchymal stem cells (hMSCs) has been one of the most explored options. We present in this paper an alternative method to mimic the spatial pattern of HUVECs and hMSCs found in native osteons based on the use of extrusion-based 3D bioprinting (3DP). We developed a 3DP biphasic osteon-like scaffold, containing two separate osteogenic and vasculogenic cell populations encapsulated in a fibrin bioink in order to improve neovascularization. To this end, we optimized the fibrin bioink to improve the resolution of printed strands and ensure a reproducible printing process; the influence of printing parameters on extruded strand diameter and cell survival was also investigated. The mechanical strength of the construct was improved by co-printing the fibrin bioink along a supporting PCL carrier scaffold. Compressive mechanical testing showed improved mechanical properties with an average compressive modulus of 131 ± 23 MPa, which falls in the range of cortical bone. HUVEC and hMSC laden fibrin hydrogels were printed in osteon-like patterns and cultured in vitro. A significant increase in gene expression of angiogenic markers was observed for the biomimetic scaffolds. Finally, biphasic scaffolds were implanted subcutaneously in rats. Histological analysis of explanted scaffolds showed a significant increase in the number of blood vessels per area in the 3D printed osteon-like scaffolds. The utilization of these scaffolds in constructing biomimetic osteons for bone regeneration demonstrated a promising capacity to improve neovascularization of the construct. These results indicates that proper cell orientation and scaffold design could play a critical role in neovascularization.

中文翻译：

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生物打印的骨样支架增强体内新血管形成。


骨组织工程师在尝试制造用于治疗大骨缺损的更大结构时面临着艰巨的挑战，即移植物的血管化。基于细胞的方法，特别是使用人脐静脉内皮细胞（HUVEC）和人间充质干细胞（hMSC）的体外共培养是探索最多的选择之一。我们在本文中提出了一种替代方法，基于使用基于挤出的 3D 生物打印 (3DP) 来模拟天然骨中发现的 HUVEC 和 hMSC 的空间模式。我们开发了一种 3DP 双相骨样支架，包含封装在纤维蛋白生物墨水中的两个独立的成骨细胞和血管生成细胞群，以改善新血管形成。为此，我们优化了纤维蛋白生物墨水，以提高打印线的分辨率并确保打印过程的可重复性；还研究了打印参数对挤出链直径和细胞存活的影响。通过沿着支撑 PCL 载体支架共同打印纤维蛋白生物墨水，提高了构建体的机械强度。压缩机械测试显示机械性能得到改善，平均压缩模量为 131 ± 23 MPa，属于皮质骨范围。将 HUVEC 和 hMSC 负载的纤维蛋白水凝胶打印成骨样图案并在体外培养。观察到仿生支架的血管生成标记物的基因表达显着增加。最后，将双相支架植入大鼠皮下。外植支架的组织学分析显示，3D 打印的骨样支架中每个区域的血管数量显着增加。 利用这些支架构建用于骨再生的仿生骨，证明了其在改善该结构的新血管形成方面具有良好的能力。这些结果表明，正确的细胞方向和支架设计可以在新血管形成中发挥关键作用。