Researchers from莱斯大学已经开发了一种使用3D打印的新方法，从糖粉中创建人造血管网络。
Replacing traditional production methods with Selective Laser Sintering (SLS) 3D printing, the team created sacrificial templates made from laser-sintered carbohydrate powders. These sugar-based constructs enable cell-laden hydrogels to be patterned with dendritic vessel networks, without the use of support materials. The newly-devised technique could improve the speed and scale of biomaterial production.
“One of the biggest hurdles to engineering clinically relevant tissues is packing a large tissue structure with hundreds of millions of living cells,” said Ian Kinstlinger, lead author and graduate student at Rice’s Brown School of Engineering. “Selective Laser Sintering gives us far more control in all three dimensions, allowing us to easily access complex topologies while still preserving the utility of the sugar material.”
“A major benefit of this approach is the speed at which we can generate each tissue structure. We can create some of the largest tissue models yet demonstrated in under five minutes.”
Devising a new and improved 3D printing method
Metabolic function in human tissues is sustained by the delivery of oxygen and nutrients, as well as the removal of waste, through complex 3D networks of blood vessels. Understanding vascular systems was essential for scientists in creating multicellular organisms, and reproducing them is equally vital to enabling 3D printed tissues to be developed. According to the research team, these tissues need to be supported using biocompatible matrices, in order to survive and supply the required nutrients to the tissue’s host.
先前的研究已经见过软光刻和needle moulding methods used to create these matrices, but 3D printing advances have led to the increased adoption of methods such as direct extrusion and inkjet-created polymers. In other research, light has been harnessed to generate sophisticated microchannel architectures, but ‘sacrificial templating’ has emerged as the dominant and most widely-used method.
The templating technique involves creating a temporary sacrificial gelatin in the shape of the desired vascular network, in which cells are encased, and then selectively removed. While extrusion-based 3D printing techniques have led to an increased adoption of this method, the features and complexity of the sacrificially templated networks have remained limited. “There are certain architectures, such as overhanging structures, branched networks and multivascular networks, which you really can’t do well with extrusion printing,” explained Jordan Miller, co-author of the study, and Assistant Professor of Bioengineering at Rice.
As a result, vascular systems printed using extrusion methods, are often subject to deformation or collapse under their own weight, and their viscosity and surface tension make precise dispensing of small volumes difficult. Moreover, printing the cells with support materials may mitigate these issues, but at the expense of longer print times and additional post-processing steps, which become increasingly difficult with increasing vascular complexity.
SLS 3D printing meanwhile, utilizes a fully supported and powder-based build volume, which enables the fabrication of objects with complicated overhangs and unsupported geometries. “Selective laser sintering gives us far more control in all three dimensions, allowing us to easily access complex topologies while still preserving the utility of the sugar material,” added Miller.
研究小组推测,使用SLS 3 d printing to produce the sacrificial materials instead of extrusion techniques, could allow vascular networks in hydrogels to be readily patterned in the presence of fragile human cells. By creating extensively branched carbohydrate filament networks via SLS, and applying them sacrificially to pattern volumetric vascular networks, the team aimed to create a faster and more stable bioprinting process.
建立基于糖的血管网络
Isomalt是一种常用于无糖肠含糖的糖醇，与SLS兼容，并且该团队设计了一个工作流程，用于自动制造Isomalt粉末的3D结构。虽然可以烧结一层纯同层，但粉末的凝聚力强，相对较差的流动性使其非常适合根据需要扩散到光滑的薄层中。发现进一步将粉末与玉米淀粉混合在一起，可以有效地增加粉末的流量，同时保持烧结的质量。使用此混合物，研究团队成功地制造了3D分支和不支持的几何形状的结构。
Beginning with the patterning of a simple branched architecture, the researchers went on to cast a series of elastomers, stiff plastics and hydrogels around post-processed carbohydrates. During the process, the hydrogel became semi-solid within minutes, and the original template was then sacrificed, being dissolved in water or phosphate buffered saline (PBS). In each case, perfusion through the patterned channel network demonstrated channel patency and the connectivity of branched filaments. Despite the opacity of the sintered carbohydrates, polyethylene glycol and diacrylate gels were successfully polymerized by incident light from various angles, demonstrating the team’s methodology.
Moreover, the researchers’ newly-developed iteration of OpenSLS hardware and firmware, optimized the process for carbohydrate SLS, and an updated software toolchain was used to prepare 3D models. By manipulating the multi-extruder capabilities of an open-source slicing software designed for extrusion 3D printers, the team were able to encode specific sintering parameters, enabling them to fine tune the model’s final geometry.
与科学家合作华盛顿大学他的研究小组专门研究delicate cells, the team later demonstrated the seeding of endothelial cells in rodent liver cells called hepatocytes. “We showed that perfusion through 3D vascular networks allows us to sustain these large liverlike tissues,” said Miller. “While there are still long-standing challenges associated with maintaining hepatocyte function, the ability to both generate large volumes of tissue and sustain the cells in those volumes for sufficient time to assess their function is an exciting step forward.”
The Rice team’s new methodology enabled them to overcome the drawbacks of previous 3D printing techniques, and produce elaborate fluidic networks within engineered living tissues. Moreover, while the researchers’ OpenSLS technique allowed them to effectively create the carbohydrates with diameters as small as 300μm, the higher quality optical components in commercial SLS printers could yet yield higher resolution templates. This opens the opportunity for further upgrades for the process. Nonetheless, the rapid nature of the manufacturing process, with none of the experiments taking longer than 15 minutes, could yet enable the process to be utilized in a range of bioprinting applications.
“这种方法可以与许多其他生物打印技术一起使用多种材料鸡尾酒。”Kelly Stevens, study co-author and Bioengineer at University of Washington. “This makes it incredibly versatile.”
3D打印中的血管应用
A range of additive manufacturing techniques have been developed by companies and researchers in recent years, with the goal of producing vascular-like structures. Researchers from诺丁汉大学andQueen Mary University of Londonfor instance, have 3D printed graphene oxide with a protein which can organise intostructures that replicate vascular tissues。
波士顿大学工程学院另一方面，科学家开发了一种治疗缺血的新方法，3D printing a vascular patch这鼓励血管生长。对啮齿动物进行了测试，并被证明能够在其体内运输营养。
Swedish manufacturerCELLINKand Texas-based biomanufacturing company体积, meanwhile, introduced a 3D printer, which is designed to produce large vascular structures. TheLumen X数字光处理（DLP）生物生产商works with bioinks to print high-resolution, macroporous, and vasculature structures.
研究人员的发现在其论文中详细介绍了“Generation of model tissues with dendritic vascular networks via sacrificial laser-sintered carbohydrate templates” published in the自然生物医学工程日记于2020年6月29日。该报告由伊恩·金斯特林格（Ian S.路易斯·罗森伯格（Louis-Rosenberg），弗雷德里克·约翰逊（Fredrik Johansson），凯文·D·詹森（Kevin D.
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Featured image shows graduate student Ian Kinstlinger preparing the selective laser sintering system in the Miller bioengineering lab at Rice University. Photo via Rice University.
