Department of Supramolecular and Biomaterials Chemistry, Leiden Institute of Chemistry, Leiden University, P.O. Box 9502, Leiden, 2300 RA, Netherlands.
Division of Drug Discovery and Safety, Leiden Academic Centre for Drug Research, Leiden University, P.O. Box 9502, Leiden, 2300 RA, Netherlands.
Adv Healthc Mater. 2021 Jun;10(11):e2001903. doi: 10.1002/adhm.202001903. Epub 2021 Apr 30.
A major challenge in the use of HepG2 cell culture models for drug toxicity screening is their lack of maturity in 2D culture. 3D culture in Matrigel promotes the formation of spheroids that express liver-relevant markers, yet they still lack various primary hepatocyte functions. Therefore, alternative matrices where chemical composition and materials properties are controlled to steer maturation of HepG2 spheroids remain desired. Herein, a modular approach is taken based on a fully synthetic and minimalistic supramolecular matrix based on squaramide synthons outfitted with a cell-adhesive peptide, RGD for 3D HepG2 spheroid culture. Co-assemblies of RGD-functionalized squaramide-based and native monomers resulted in soft and self-recovering supramolecular hydrogels with a tunable RGD concentration. HepG2 spheroids are self-assembled and grown (≈150 µm) within the supramolecular hydrogels with high cell viability and differentiation over 21 days of culture. Importantly, significantly higher mRNA and protein expression levels of phase I and II metabolic enzymes, drug transporters, and liver markers are found for the squaramide hydrogels in comparison to Matrigel. Overall, the fully synthetic squaramide hydrogels are proven to be synthetically accessible and effective for HepG2 differentiation showcasing the potential of this supramolecular matrix to rival and replace naturally-derived materials classically used in high-throughput toxicity screening.
在使用 HepG2 细胞培养模型进行药物毒性筛选时，面临的一个主要挑战是其在 2D 培养中的不成熟。在 Matrigel 中进行 3D 培养可促进球体的形成，这些球体表达与肝脏相关的标志物，但它们仍然缺乏各种原代肝细胞功能。因此，仍然需要替代基质，这些基质的化学成分和材料特性可得到控制，以促进 HepG2 球体的成熟。在此，提出了一种基于完全合成的最小超分子基质的模块化方法，该基质基于带有细胞黏附肽 RGD 的 squaramide 合成子。RGD 功能化的 squaramide 基和天然单体的共组装导致具有可调 RGD 浓度的柔软且可自我修复的超分子水凝胶。RGD 功能化的 squaramide 基和天然单体的共组装导致具有可调 RGD 浓度的柔软且可自我修复的超分子水凝胶。RGD 功能化的 squaramide 基和天然单体的共组装导致具有可调 RGD 浓度的柔软且可自我修复的超分子水凝胶。HepG2 球体在超分子水凝胶中自组装并生长（≈150 µm），在 21 天的培养过程中具有高细胞活力和分化。重要的是，与 Matrigel 相比，超分子水凝胶中 I 期和 II 期代谢酶、药物转运体和肝脏标志物的 mRNA 和蛋白表达水平显著更高。总体而言，完全合成的 squaramide 水凝胶具有可合成性，并且可有效促进 HepG2 分化，证明了这种超分子基质具有替代经典用于高通量毒性筛选的天然衍生材料的潜力。
