The importance of effective ligand concentration to direct epithelial cell polarity in dynamic hydrogels.

Epithelial cysts and organoids are multicellular hollow structures formed by correctly polarized epithelial cells. Important in steering these cysts from single cells is the dynamic regulation of extracellular matrix presented ligands, and matrix dynamics. Here, we introduce control over the effective ligand concentration, decoupled from bulk and local mechanical properties, in synthetic dynamic supramolecular hydrogels formed through non-covalent crosslinking of supramolecular fibers. Control over the effective ligand concentration is realized by (1) keeping the ligand concentration constant, but changing the concentration of non-functionalized molecules or by (2) varying the ligand concentration, while keeping the concentration of non-functionalized molecules constant. Our results show that in 2D, the effective ligand concentration within our supramolecular fibers rather than gel stiffness (from 0.1 to 8 kPa) regulates epithelial polarity. In 3D, increasing the effective ligand concentration from 0.5 to 2 mM strengthens the effect of increased gel stiffness from 0.1 to 2 kPa, to synergistically yield more correctly polarized cysts. Through integrin manipulation, we show that epithelial polarity is regulated by tension-based homeostasis between cells and matrix. Our results reveal the effective ligand concentration as influential factor in regulating epithelial polarity and provide insights on engineering of synthetic biomaterials for cell and organoid culture. This article is protected by copyright. All rights reserved.