Structurally and mechanically tuned macroporous hydrogels for scalable mesenchymal stem cell-extracellular matrix spheroid production.

Mesenchymal stem cells (MSCs) are essential in regenerative medicine. However, conventional expansion and harvesting methods often fail to maintain the essential extracellular matrix (ECM) components, which are crucial for their functionality and efficacy in therapeutic applications. Here, we introduce a bone marrow-inspired macroporous hydrogel designed for the large-scale production of MSC-ECM spheroids. Through a soft-templating approach leveraging liquid-liquid phase separation, we engineer macroporous hydrogels with customizable features, including pore size, stiffness, bioactive ligand distribution, and enzyme-responsive degradability. These tailored environments are conducive to optimal MSC proliferation and ease of harvesting. We find that soft hydrogels enhance mechanotransduction in MSCs, establishing a standard for hydrogel-based 3D cell culture. Within these hydrogels, MSCs exist as both cohesive spheroids, preserving their innate vitality, and as migrating entities that actively secrete functional ECM proteins. Additionally, we also introduce a gentle, enzymatic harvesting method that breaks down the hydrogels, allowing MSCs and secreted ECM to naturally form MSC-ECM spheroids. These spheroids display heightened stemness and differentiation capacity, mirroring the benefits of a native ECM milieu. Our research underscores the significance of sophisticated materials design in nurturing distinct MSC subpopulations, facilitating the generation of MSC-ECM spheroids with enhanced therapeutic potential.

Toward Practical Applications of Engineered Living Materials with Advanced Fabrication Techniques.

Engineered Living Materials (ELMs) are materials composed of or incorporating living cells as essential functional units. These materials can be created using bottom-up approaches, where engineered cells spontaneously form well-defined aggregates. Alternatively, top-down methods employ advanced materials science techniques to integrate cells with various kinds of materials, creating hybrids where cells and materials are intricately combined. ELMs blend synthetic biology with materials science, allowing for dynamic responses to environmental stimuli such as stress, pH, humidity, temperature, and light. These materials exhibit unique "living" properties, including self-healing, self-replication, and environmental adaptability, making them highly suitable for a wide range of applications in medicine, environmental conservation, and manufacturing. Their inherent biocompatibility and ability to undergo genetic modifications allow for customized functionalities and prolonged sustainability. This review highlights the transformative impact of ELMs over recent decades, particularly in healthcare and environmental protection. We discuss current preparation methods, including the use of endogenous and exogenous scaffolds, living assembly, 3D bioprinting, and electrospinning. Emphasis is placed on ongoing research and technological advancements necessary to enhance the safety, functionality, and practical applicability of ELMs in real-world contexts.