Nanocomposite Hydrogel Bioinks for 3D Printing of Living Cells

活细胞的3D打印，或3D生物打印，在构建3D组织和最终器官方面有着巨大的希望。然而，尽管最近在非生物材料的3D打印方面取得了进展，但活细胞的3D打印仍然是一个挑战。其中一个关键的挑战是缺乏具有3D打印性和生物相容性的生物墨水。该奖项支持在3D打印过程中封装活细胞的纳米复合水凝胶生物墨水的基础研究。本研究结果不仅可以合理设计具有3D打印性和生物相容性的细胞包封生物墨水，还可以实现组织结构的高分辨率3D打印。这项研究对生物医学的许多领域都有潜在的益处，包括组织和器官模型的发展、组织工程和再生医学。本项目的研究目标是：(1)了解负载细胞的纳米复合生物墨水的组成对其动态力学性能（剪切变薄和自愈性能）和细胞活力的影响；(2)建立负载细胞的纳米复合生物墨水的动态力学性能与其3D打印能力和细胞相容性（打印组织中的细胞活力）之间的关系。为了实现第一个目标，将合成由表面功能化的单壁碳纳米管、甲基丙烯酸明胶（水凝胶基质）和活细胞组成的凝胶相纳米复合生物墨水。纳米管和甲基丙烯酸明胶的浓度在0 ~ 10%之间变化，总固相分数小于10%。成纤维细胞和间充质干细胞（10^6至10^8个细胞/mL）将被用作模型细胞。这些凝胶相生物墨水的储存和损失剪切模量将通过振荡应变扫描和应变步骤（触变性）来测量，使用流变仪来确定剪切变薄和自愈特性。将测量包裹在生物墨水中的细胞的活力（每小时和每天，最多7天）。为了实现第二个目标，将使用100和200微米的喷嘴打印多层3D组织结构。凝胶相生物墨水的3D可打印性将通过使用光学显微镜表征打印结构（形状定义和结构完整性）来确定。生物墨水可以产生直径小于200微米的自支撑3D结构，将被定义为可3D打印的。纳米复合生物墨水的细胞相容性将通过测量打印组织中细胞的活力来确定（每天最多7天）。

3D printing of living cells, or 3D bioprinting, holds great promise to build 3D tissues and, ultimately, organs. However, despite recent advances in 3D printing of non-biological materials, 3D printing of living cells remains a challenge. One of the key challenges is the lack of bioinks with 3D printability and biocompatibility. This award supports fundamental research on nanocomposite hydrogel bioinks that encapsulate living cells in 3D printing process. Results from this research will enable not only the rational design of cell-encapsulating bioinks with 3D printability and biocompatibility, but also the high resolution 3D printing of tissue structures. This research potentially benefits many areas in biomedicine, including the development of tissue and organ models, tissue engineering, and regenerative medicine.The research objectives of this project are (1) to understand the effects of compositions of cell-laden nanocomposite bioinks on their dynamic mechanical properties (shear-thinning and self-healing properties) and cell viability; and (2) to establish the relationship between the dynamic mechanical properties of cell-laden nanocomposite bioinks and their 3D printability and cell-compatibility (cell viability in printed tissues). To achieve the first objective, gel-phase nanocomposite bioinks that consist of surface-functionalized single-walled carbon nanotubes, gelatin methacrylate (hydrogel matrix), and living cells will be synthesized. The concentration of nanotubes and gelatin methacrylate will be varied from 0 to 10% with the total solid fraction of less than 10%. Fibroblasts and mesenchymal stem cells (10^6 to 10^8 cells/mL) will be used as model cells. The storage and loss shear moduli of these gel-phase bioinks will be measured with oscillatory strain sweeps and strain steps (thixotropy) using a rheometer to determine the shear-thinning and self-healing properties. The viability of cells encapsulated in bioinks will be measured (hourly and daily up to 7 days). To achieve the second objective, multilayer 3D tissue structures will be printed with 100 and 200 micron nozzles. The 3D printability of gel-phase bioinks will be determined by characterizing printed structures (shape definition and structural integrity) using optical microscopy. Bioinks that produce self-supporting 3D structures with a filament diameter of less than 200 microns will be defined as 3D printable. The cell-compatibility of nanocomposite bioinks will be determined by measuring the viability of cells in printed tissues (daily up to 7 days).