Understanding the Printing Dynamics during Inkjetting of Cell-Laden Bioink for 3D Biomanufacturing Applications

喷墨打印是一种将液滴以计算机控制的模式沉积在基板上的技术，是制造3D活体组织和器官最有前途的技术之一。所采用的载体液，被称为生物墨水，包含活细胞。由于细胞与细胞之间的相互作用，活细胞在打印喷嘴中相互粘附，形成细胞聚集体。这些聚集体可能堵塞喷嘴或导致意外的液滴沉积在基材上，这两种情况都会严重影响喷墨打印的可靠性。本项目涉及基础科学研究，旨在了解喷墨打印过程中活细胞粘附和聚集的机制，以及如何控制打印结果。如果成功，这项研究将连接基于喷气的生物打印制造科学和生物医学应用，为3D功能组织和器官的再生开辟新的途径，并显著推动美国医疗保健行业的发展。研究成果将无缝整合到目前在制造工程和生物工程项目中提供的生物制造课程中，以增加女性和少数民族学生的参与。该研究还将通过区域夏令营（探索工程）和国家大众媒体传播，以教育和激励公众和下一代生物工程师。研究生参与研究活动将加强未来生物工程和先进制造业的劳动力。研究目的是获得关于细胞负载生物墨水喷墨打印过程中打印头喷嘴内细胞聚集和喷嘴外卫星液滴形成的基本新知识。在打印过程中，生物墨水从压电打印头喷射出来形成液滴，这些液滴精确地沉积在基板上，形成一层一层的3D结构。据信，从喷嘴中排出生物墨水所需的压力波的性质影响细胞聚集和卫星滴形成的可能性。为了研究细胞聚集，将使用多相方法模拟细胞粘附，该方法将生物链接建模为悬液，将活细胞的粘弹性流体模型与固体模型耦合。在不同的激励电压和细胞浓度下，对质量和动量守恒的控制方程进行了数值求解。由于卫星液滴的形成取决于韧带径向变薄和轴向收缩的竞争作用，因此将对这两个过程进行分析模拟，以预测卫星液滴的形成。一旦了解了核心基础知识，就可以利用这些知识以无量纲数字生成工艺阶段图，最终确定最佳印刷条件。此外，在打印和培养7天后，将立即评估细胞活力和增殖能力，以确认所选择的喷墨打印条件对细胞没有负面影响。由此产生的科学知识高度适用于其他生物制造技术。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

Inkjet printing, where droplets of liquid are deposited onto a substrate in a computer-controlled pattern, is one of the most promising technologies for creation of 3D living tissues and organs. The adopted carrier fluid, referred to as bioink, incorporates living cells. Due to cell-cell interaction, living cells adhere to each other in the print nozzle to form cell aggregates. These aggregates can clog the nozzle or result in unplanned droplets deposited on the substrate, both of which can significantly affect inkjet printing reliability. This project involves fundamental scientific research to understand the mechanisms of living cell adhesion and aggregation during inkjet printing, and how to control the printing outcomes. If successful, this research can bridge jet-based bioprinting manufacturing science and biomedical applications, open new avenues in regeneration of 3D functional tissues and organs, and significantly advance the U.S. healthcare industry. The research outcomes will be seamlessly integrated into a biomanufacturing course currently offered in the Manufacturing Engineering and Bioengineering programs to increase participation of female and minority students. The research will also be disseminated through the regional summer camp (Explore Engineering) and national mass media to educate and inspire the general public and the next generation of bioengineers. Engagement of graduate students in the research activities will strengthen the future workforce in bioengineering and advanced manufacturing industries.The research objective is to gain fundamental new knowledge regarding cell aggregation inside the printhead nozzle, and the satellite droplet formation outside the nozzle during inkjet printing of cell-laden bioink. During printing, the bioink is ejected from a piezoelectric printhead to form droplets, which are precisely deposited on the substrate to form 3D constructs layer by layer. It is believed that the nature of pressure wave required to expel the bioink from the nozzle affects both the likelihood of cell aggregation and satellite drop formation. To investigate cell aggregation, cell adhesion will be simulated using a multiphase approach, which models the bioink as a suspension coupling a viscoelastic fluid model with a solid model for living cells. The governing equations for the conservation of mass and momentum will be numerically solved under different excitation voltages and cell concentrations. As the satellite droplet formation depends on the competing actions of ligament radial thinning and axial contraction, both processes will be modeled analytically to predict satellite droplet formation. Once the core fundamentals are understood, the knowledge will be leveraged to generate process phase diagrams in terms of non-dimensional numbers to ultimately identify optimal printing conditions. Additionally, cell viability and proliferation will be assessed immediately after printing and after 7-day incubation to confirm the chosen inkjet printing conditions have no negative impact on the cells. The resulting scientific knowledge is highly adoptable to other biofabrication technologies.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.