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Biological and Medical Applications of Materials and Interfaces
Rapid and Versatile Cell Aggregate Formation Using Lipid-Conjugated Heparin Eunsol Kim, Jong Chul Kim, Kiyoon Min, MeeiChyn Goh, and Giyoong Tae ACS Appl. Mater. Interfaces, Just Accepted Manuscript • Publication Date (Web): 28 Jun 2018 Downloaded from http://pubs.acs.org on June 28, 2018
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ACS Applied Materials & Interfaces
Rapid and Versatile Cell Aggregate Formation Using Lipid-Conjugated Heparin Eunsol Kim†, Jong Chul Kim†, Kiyoon Min†, MeeiChyn Goh, and Giyoong Tae* School of Materials Science and Engineering, Gwangju Institute of Science and Technology, 123 Cheomdan-gwagiro, Buk-gu, Gwangju 61005, Republic of Korea.
KEYWORDS: cell surface engineering; lipid-conjugated heparin; instant cell aggregation; multi-compartment; cell thread
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ABSTRACT
Cell aggregates hold significant therapeutic promise for in vitro cell analysis, ex vivo tissue models, and in vivo cell therapy and tissue engineering. Traditional methods of making cell aggregates require long incubation times and can only produce 3D-spheroid-shaped aggregates. We propose a novel method of making cell aggregates of diverse sizes and shapes using lipid-conjugated heparin. Shaking the cell suspension containing a small amount of lipid-conjugated heparin for approximately 30 min produced cell aggregates. This approach can be applied to any cell type, including stem cells, fibroblast cells, and T lymphocytes. The shape of biocompatible templates could modulate the shape of cell aggregates. In addition to layered, multi-compartmental cell aggregates on template, template-free, tube-shaped cell aggregates could also be made. The cell aggregates formed were alive and maintained biological activities.
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ACS Applied Materials & Interfaces
1. INTRODUCTION The construction of a three-dimensional (3D) cell aggregation structure is widely applicable as a powerful approach to analyzing in vitro cell behaviors, creating a specific ex vivo tissue model such as a tumor, and enhancing the in vivo efficiency of cell therapy and tissue engineering. Cell aggregation mediates cell-cell interactions that determine a variety of vital processes in living cells, such as differentiation, migration, mitosis, and apoptosis signaling. Indeed, 3D cell aggregates enhance the regenerative capacity of mesenchymal stem cells (MSCs) through the action of anti-inflammatory cytokines, chemotaxis factors, and proangiogenic factors.1 Additionally, compared to single cell transplantation, cell aggregates provide improved biological properties, including multilineage potential, secretion of therapeutic factors, and resistance of MSCs against ischemic conditions.2 Various methods have been developed to generate 3D cell aggregates. Traditional strategies for making cell aggregates include pellet culture,3,4 hanging drop culture,5,6 non-adhesive surface culture,7 and rotational culture.8,9 However, these conventional methods have limitations, such as their time-consuming processes (from several days to weeks) and difficulty in controlling the size and shape of the aggregates. Recently, several new approaches for obtaining 3D cell aggregates have been reported. The shape or surface of the cell culture plate can be modified to provide more efficient formation of the cell aggregate or with better control. For example, by introducing concave wells on the surface, the shape and size of the cell aggregates can be better controlled.10 Similarly, micropatterning of scaffold can increase the differentiation efficiency of MSCs.11 Micropatterned culture of embryoid bodies can control the growth and differentiation by separation distance of them.12 The patterning of super-hydrophobic domains13or coating with functional polymers on a cell 3
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culture plate14, 15 was also effective in forming cell aggregates. 3D bioprinting of spheroids by needle-array system can assemble cell aggregates to a desired size and shape.16 Although all of these approaches are effective in controlling the formation of cell aggregates, they require specialized processes, functional coatings, or a relatively long time (>3 days) to obtain cell aggregates. In contrast, if cell aggregates can be formed in a short time, it is possible to apply the formed cell aggregates to a personalized, patient-tailored therapy in a single operation. To shorten cell aggregation time, it is possible to induce bridging among cells by directly modifying the cell surface instead of the cell culture substrate. Methods that have reported using this approach typically use synthetic polymers containing the affinity group on the cell membrane. For example, cell crosslinking can be induced by hyperbranched C18,17 polyvalent choline phosphate,18 nanofilm coating with fibronectin/gelatin19, or polymer containing chemically reactive groups with the cell membrane.20 Using these polymers, cell aggregates are formed instantaneously and can be applied to blood clotting. However, they are synthetic polymers, and the induced intercellular interactions are too strong not to distort the cell membrane after cell aggregation, which might interfere with or hinder the biological activity of cell aggregates. In the present work, we propose a simple method for generating cell aggregates with a reproducible size and shape in a controlled and rapid manner (
