Biofabrication strategies for making hematopoietic tissues

Abstract

Hematopoietic stem cells (HSCs) sustain lifelong blood and immune cell production and form the basis of hematopoietic stem cell transplantation (HSCT), one of the most established stem cell therapies. However, HSCT remains constrained by limited donor availability, graft-versus-host disease, and insufficient cell numbers. Although ex vivo HSC expansion could alleviate these limitations, clinically relevant expansion has remained difficult to achieve because conventional culture systems poorly reproduce the complex microenvironments that regulate HSC fate in vivo. In particular, current models often lack the spatial organization, multicellular interactions, dynamic extracellular matrix properties, and mechanical cues characteristic of hematopoietic niches. This thesis develops a modular biofabrication framework for engineering human hematopoietic microenvironments with greater structural and biological fidelity. The work combines stimuli-responsive biomaterials, volumetric and extrusion-based bioprinting, and organoid technologies to recreate key features of adult and developmental hematopoietic niches. First, a photodegradable hyaluronic acid hydrogel was developed to enable spatiotemporal modulation of matrix mechanics. Local light exposure induced controlled matrix softening and altered stromal and endothelial cell morphology and organization, demonstrating the possibility of dynamically rewriting mechanical cues within a 3D microenvironment. Next, the cytocompatibility of volumetric bioprinting (VBP) was assessed beyond conventional viability readouts. Under commonly used printing conditions, VBP did not induce detectable oxidative stress, DNA damage, or major cellular stress responses in mesenchymal stromal cells. To overcome intrinsic limitations of VBP in cell density and material compartmentalization, Embedded Volumetric Printing (EmVP) was subsequently introduced. By integrating extrusion-based bioprinting with VBP, EmVP enabled spatial organization of distinct stromal populations at different densities while preserving construct fidelity and generating mechanically heterogeneous environments. The thesis further explored multicellular organoids as biological building blocks for hematopoietic niche engineering. Stromal Vascular Organoids generated from mesenchymal stromal and endothelial colony-forming cells formed stable microvascular networks that supported hematopoietic stem and progenitor cell homing under static and perfused conditions. In parallel, an induced pluripotent stem cell-derived fetal liver organoid platform was developed to model the developmental niche associated with extensive hematopoietic expansion. Together, these findings show that faithful hematopoietic niche engineering requires integration of cellular complexity with spatial, architectural, and mechanical control. Rather than providing a single complete niche model, this thesis establishes a modular toolbox of responsive materials, biofabrication technologies, and organoid building blocks that can be progressively combined and refined. These approaches provide a foundation for more physiologically relevant human hematopoietic models and may ultimately support the development of platforms for clinically meaningful HSC expansion.