Novel, engineered bio-inks for 3D printing of complex, perfusable structures

非技术摘要利用活细胞打印三维组织的生物打印具有巨大的未来潜力，但这项新兴技术的进步受到生物材料缺乏的限制，这些生物材料既具有可打印所需的机械性能，又具有与活细胞接口的适当的生化性能。特别是，能够灌流、运输氧气、营养物质和其他维持生命的成分的生物材料严重缺乏。血管网络可能是对可灌流结构的需求的最好证明，血管网络是向活组织输送氧气和营养物质所必需的。可灌流的结构对全身许多其他组织至关重要，包括淋巴系统、呼吸道和胃肠道。为了实现下一代打印组织，该项目将开发一系列生物材料，实现一种新的生物制造策略，称为嵌入式3D打印中均匀界面扩散剂的凝胶(GUIDE-3DP)。GUIDE-3DP材料将允许快速制造可灌装的互联通道网络，并精确控制其形状和大小。在目标1中，将打印具有复杂分支点的流体可灌流结构，例如分支血管的模拟。在目标2中，将开发用于气体可灌流结构的材料。作为一个案例研究，人类的肠道组织将被打印出来，并评估氧气通过打印的生物材料的运输。在目标3中，将开发能够制造内径精确变化的连续容器的材料。作为案例研究，将打印(1)血管狭窄(其中血管的一个区域被收缩)和(2)大肠(具有重复的袋状结构)的打印模型。这些生物材料将使未来能够制造用于促进生物材料、生物技术、国民健康的各种应用的活组织模拟材料，并将进一步使美国在新兴的生物制造领域拥有全球领先地位。技术摘要无论是在组织工程、器官发育和疾病的体外模型，还是在细胞行为的基础研究中，灌流，特别是灌流生物材料，仍然是形成三维(3D)多细胞结构的最关键的挑战之一。制造具有特定几何形状的通道的挑战对于全身的多个组织来说是重要的，包括血管、淋巴管、呼吸道和胃肠道。为了应对这一挑战，该项目将开发一系列生物材料，实现一种新的生物制造策略，称为嵌入式3D打印中均匀界面扩散剂的凝胶(GUIDE-3DP)。嵌入式3D打印包括在支撑材料中制造所需的结构，减少由于重力造成的变形和屈曲，并使复杂结构能够打印。GUIDE-3DP方法建立在这种方法的基础上，开发了一种界面扩散策略，通过精确控制分支几何形状和血管直径，快速制造可灌装的互联通道网络。在目标1中，制造了具有复杂分支点的流体可灌流结构。作为一个生物学案例研究，将描述生物打印分支结构中内皮细胞的形态和表型，重点是基质力学如何改变细胞对流体剪应力的反应。在目标2中，将制造用于控制氧浓度的气体可灌流结构。作为一个案例研究，将打印一个3D人类肠道器官培养模型，并评估氧合在调节肠道干细胞命运中的作用。在目标3中，将制造管腔直径精确变化的连续血管，因为这种几何形状通常存在于体内的许多结构中。作为案例研究，将打印(1)血管狭窄(血管区域收缩)和(2)大肠(具有重复的、袋状结构)的体外模型。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

Non-technical AbstractBioprinting, printing with living cells, of three-dimensional tissue has immense future potential, but progress of this emerging technology is limited by a lack of biomaterials that have the necessary mechanical properties to be printable while also having the appropriate biochemical properties to interface with living cells. In particular, biomaterials that enable perfusion, the transport of oxygen, nutrients, and other life-sustaining components, is critically lacking. The need for perfusable structures is perhaps best demonstrated through blood vessel networks, which are required to deliver oxygen and nutrients to living tissue. Perfusable structures are critically important for many other tissues throughout the body including the lymphatic system, airways, and the gastrointestinal tract. To achieve the next generation of printed tissue, this project will develop a family of biomaterials that enable a new biofabrication strategy termed Gelation of Uniform Interfacial Diffusant in Embedded 3D Printing (GUIDE-3DP). The GUIDE-3DP materials will allow rapid fabrication of perfusable networks of interconnected channels with precise control over their shapes and sizes. In Aim 1, fluid-perfusable structures with complex branch points, such as mimics of branched blood vessels will be printed. In Aim 2, materials for gas-perfusable structures will be developed. As a case study, a human intestinal tissue will be printed and evaluated for the transport of oxygen through the printed living material. In Aim 3, materials that enable fabrication of continuous vessels with precise variation of the internal diameters will be developed. As case studies, printed models of (1) vascular stenosis (in which a region of the blood vessel is constricted) and (2) the large intestine (which has a repetitive, pouch-like structure) will be printed. These biomaterials will enable the future fabrication of living tissue mimics for a variety of applications that advance, biomaterials, biotechnology, national health and will further position the US to have global leadership in the emerging field of biomanufacturing. Technical AbstractPerfusion, and specifically perfusion-enabling biomaterials, remains one of the most critical challenges in the formation of three-dimensional (3D) multicellular structures, whether for the purposes of tissue engineering, in vitro models of organ development and disease, or fundamental studies of cell behavior. The challenge of fabricating channels with specified geometry are important for multiple tissues throughout the body, including blood vessels, lymphatics, airways, and the gastrointestinal tract. To address this challenge, this project will develop a family of biomaterials that enable a new biofabrication strategy termed Gelation of Uniform Interfacial Diffusant in Embedded 3D Printing (GUIDE-3DP). Embedded 3D printing involves the fabrication of desired structures within a support material, reducing deformation and buckling due to gravity and enabling the printing of complex structures. The GUIDE-3DP method builds upon this approach by developing an interfacial diffusant strategy to rapidly fabricate perfusable networks of interconnected channels with precise control over the branching geometry and vessel diameters. In Aim 1, fluid-perfusable structures with complex branch points are fabricated. As a biological case study, endothelial cell morphology and phenotype in the bioprinted branch structures will be characterized, with a focus on how matrix mechanics alters cellular response to fluid shear stress. In Aim 2, gas-perfusable structures for controlled oxygen concentration will be fabricated. As a case study, a 3D human intestinal organoid culture model will be printed and evaluated for the role of oxygenation in regulating intestinal stem cell fate. In Aim 3, continuous vessels with precise variation of luminal diameters will be fabricated, as this geometry commonly occurs in many structures in vivo. As case studies, in vitro models of (1) vascular stenosis (in which a region of the blood vessel is constricted), and (2) the large intestine (which has a repetitive, pouch-like structure) will be printed.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.