Mehanna Lauren E, Boyd James D, Remus-Williams Shelley, Racca Nicole M, Spraggins Dawson P, Grady Martha E, Berron Brad J
Department of Chemical and Materials Engineering, University of Kentucky, Lexington, KY, United States of America.
Department of Mechanical Engineering, University of Kentucky, Lexington, KY, United States of America.
Biomed Mater. 2025 Feb 13;20(2):025025. doi: 10.1088/1748-605X/ada508.
Rapid and strategic cell placement is necessary for high throughput tissue fabrication. Current adhesive cell patterning systems rely on fluidic shear flow to remove cells outside of the patterned regions, but limitations in washing complexity and uniformity prevent adhesive patterns from being widely applied. Centrifugation is commonly used to study the adhesive strength of cells to various substrates; however, the approach has not been applied to selective cell adhesion systems to create highly organized cell patterns. This study shows centrifugation as a promising method to wash cellular patterns after selective binding of cells to the surface has taken place. After patterning H9C2 cells using biotin-streptavidin as a model adhesive patterning system and washing with centrifugation, there is a significant number of cells removed outside of the patterned areas of the substrate compared to the initial seeding, while there is not a significant number removed from the desired patterned areas. This method is effective in patterning multiple size and linear structures from line widths of 50-200 μm without compromising immediate cell viability below 80%. We also test this procedure on a variety of tube-forming cell lines (MPCs, HUVECs) on various tissue-like surface materials (collagen 1 and Matrigel) with no significant differences in their respective tube formation metrics when the cells were seeded directly on their unconjugated surface versus patterned and washed through centrifugation. This result demonstrates that our patterning and centrifugation system can be adapted to a variety of cell types and substrates to create patterns tailored to many biological applications.
快速且具策略性的细胞放置对于高通量组织制造而言是必要的。当前的粘附细胞图案化系统依靠流体剪切流来去除图案化区域之外的细胞，但在洗涤复杂性和均匀性方面的限制阻碍了粘附图案的广泛应用。离心法通常用于研究细胞与各种底物的粘附强度；然而，该方法尚未应用于选择性细胞粘附系统以创建高度有序的细胞图案。本研究表明，在细胞与表面发生选择性结合后，离心法是一种用于洗涤细胞图案的有前景的方法。使用生物素 - 链霉亲和素作为模型粘附图案化系统对H9C2细胞进行图案化并通过离心洗涤后，与初始接种相比，在底物图案化区域之外有大量细胞被去除，而在所需图案化区域中没有大量细胞被去除。该方法对于形成线宽为50 - 200μm的多种尺寸和线性结构的图案有效，且不会使即时细胞活力低于80%。我们还在各种组织样表面材料（胶原蛋白1和基质胶）上对多种形成管的细胞系（MPCs、HUVECs）测试了该程序，当细胞直接接种在未结合的表面上与图案化并通过离心洗涤时，它们各自的成管指标没有显著差异。这一结果表明，我们所构建的图案化和离心系统能够适用于多种细胞类型和底物，以创建适合众多生物学应用的图案。
