Comparison of Natural Polysaccharides and Synthetic Polymers in Facilitating Macromolecular Crowding to Create Customized Extracellular Matrix-Rich Supramolecular Assemblies from Fibroblasts

Regenerative medicine relies on tissue engineering to create functional tissue constructs that can repair or replace damaged organs. This process requires both structural support, such as physical scaffolds, cell adhesion sites, and spatial organization of cells, and biochemical support, including reservoirs of growth factors and cytokines provided by the extracellular matrix (ECM). The ECM plays a crucial role in regulating cell proliferation, differentiation, and migration, thereby guiding tissue formation and regeneration. Tissue engineering requires efficient ECM deposition. Macromolecular crowding (MMC) has emerged as a powerful strategy to accelerate the ECM deposition. MMC accelerates metabolic events and protein assembly to imitate the congested intracellular milieu and increase ECM deposition. Various literature has documented the utility of dextran sulfate (DxS), Ficoll, polyvinylpyrrolidone (PVP), and carrageenan (CR) for collagen deposition and ECM construction. Our work reevaluated macromolecular crowders, including synthetic polymers Ficoll 400, PVP 40, and PVP 360, as well as natural polysaccharides like CR. We determined the ECM deposition pattern of variable macromolecular crowder by measuring fibronectin, collagen, and laminin. These crowders rely on the biophysical excluded volume effect, which limits the space available for other molecules to diffuse. This stabilizes folded proteins properly and makes it easier for molecules to bind and interact. Crowding also increases the effective concentration of molecules, thereby accelerating biochemical reactions within the cell. Empirical evidence shows that MMC greatly promotes ECM deposition. Importantly, different crowders cause cells to deposit the ECM in unique patterns under MMC conditions. After decellularization, these customized supramolecular assemblies form biologically relevant three-dimensional ECM scaffolds that serve as substrates for reseeding with cells such as MDA-MB-231. In tissue engineering, macromolecular crowders mimic biological tissues' dense molecular environment, speeding up ECM deposition. Improved cell growth, proliferation, and differentiation are essential for functional tissue constructs. We critically examined the role of macromolecular crowders in increasing ECM deposition for tissue engineering, highlighting their biophysical principles.