Saeki Kotone, Hiramatsu Hisataka, Hori Ayaka, Hirai Yu, Yamada Masumi, Utoh Rie, Seki Minoru
Department of Applied Chemistry and Biotechnology, Graduate School of Engineering, Chiba University, 1-33 Yayoi-cho, Inage-ku, Chiba 263-8522, Japan.
ACS Omega. 2020 Aug 20;5(34):21641-21650. doi: 10.1021/acsomega.0c02385. eCollection 2020 Sep 1.
Although many types of technologies for hydrogel-based cell cultivation have recently been developed, strategies to integrate cell-adhesive micrometer-sized supports with bulk-scale hydrogel platforms have not been fully established. Here, we present a highly unique approach to produce cell-adhesive, protein-based microfibers assisted by the sacrificial template of alginate; we applied these fibers as microengineered scaffolds for hydrogel-based cell encapsulation. Two types of microfluidic devices were designed and fabricated: a single-layered device for producing relatively thick (Φ of 10-60 μm) alginate-protein composite fibers with a uniform cross-sectional morphology and a four-layered device for preparing thinner (Φ of ∼4 μm) ones through the formation of patterned microfibers with eight distinct alginate-protein composite regions. Following chemical cross-linking of protein molecules and the subsequent removal of the sacrificial alginate from the double-network matrices, microfibers composed only of cross-linked proteins were obtained. We used gelatin, albumin, and hemoglobin as the protein material, and the gelatin-based cell-adhesive fibers were further encapsulated in hydrogels together with the mammalian cells. We clarified that the thinner fibers were especially effective in promoting cell proliferation, and the shape of the constructs was maintained even after removing the hydrogel matrices. The presented approach offers cells with biocompatible solid supports that enhance cell adhesion and proliferation, paving the way for the next generation of techniques for tissue engineering and multicellular organoid formation.
尽管近年来已开发出多种基于水凝胶的细胞培养技术，但将细胞粘附性微米级支撑物与大规模水凝胶平台相结合的策略尚未完全确立。在此，我们提出了一种极具独特性的方法，借助藻酸盐的牺牲模板来制备细胞粘附性的、基于蛋白质的微纤维；我们将这些纤维用作基于水凝胶的细胞封装的微工程支架。设计并制造了两种类型的微流控装置：一种单层装置，用于生产具有均匀横截面形态的相对较粗（直径为10 - 60μm）的藻酸盐 - 蛋白质复合纤维；另一种四层装置，用于通过形成具有八个不同藻酸盐 - 蛋白质复合区域的图案化微纤维来制备更细（直径约为4μm）的纤维。在蛋白质分子进行化学交联并随后从双网络基质中去除牺牲性藻酸盐之后，获得了仅由交联蛋白质组成的微纤维。我们使用明胶、白蛋白和血红蛋白作为蛋白质材料，并将基于明胶的细胞粘附纤维与哺乳动物细胞一起进一步封装在水凝胶中。我们阐明，较细的纤维在促进细胞增殖方面特别有效，并且即使在去除水凝胶基质后，构建体的形状仍能保持。所提出的方法为细胞提供了生物相容性固体支撑物，增强了细胞粘附和增殖，为下一代组织工程和多细胞类器官形成技术铺平了道路。
