Additive-free hyaluronic acid-based bioink for 3D bioprinting of bone marrow microenvironments

Bioprinting of soft tissues is an emerging technology with significant potential in regenerative medicine. It requires bioinks that mimic biochemical and physical properties of natural tissues and enable precise positioning of multiple cell types to allow their physiological interplay. Here, we describe the development of a bioink for bone marrow as an example of soft tissue. Bone marrow is the site of blood regeneration, driven by hematopoietic stem cells (HSCs) and relying on the orchestrated interplay of hematopoietic and stromal cells in a soft microenvironment. The bioink is based on hyaluronic acid (HA), dual-functionalized in a one-pot synthesis with alkyl side chains enhancing physical crosslinking via hydrophobic interactions and methacrylamide groups allowing covalent photo-crosslinking. Polymers are synthesized with HA of differing molecular weights, alkyl side chain lengths and modification degrees. Their gelling behavior, shear-thinning and self-healing properties deem them suitable for extrusion-based bioprinting. The ink allows two bioprinting approaches: cell encapsulation pre-printing and cell injection post-printing, both yielding excellent cell viability. The latter approach allows precise placement of hematopoietic and stromal cells in a single construct. In summary, we present a bioink enabling bioprinting of modified HA without further additives, bioprinting of encapsulated cells and injection of cells into pre-printed structures. The material is based on a polymer naturally present in bone marrow, resembles the mechanical properties of bone marrow and is suitable for bioprinting of hematopoietic and stromal cells. Thus, this bioink is a promising platform for bioprinting biomimetic bone marrow or other soft tissue constructs for future fundamental and applied research.