Process- and bio-inspired hydrogels for 3D bioprinting of soft free-standing neural and glial tissues

Abstract

A bio-inspired hydrogel for 3D bioprinting of soft free-standing neural tissues is presented. The novel filler-free bioinks were designed by combining natural polymers for extracellular matrix biomimicry with synthetic polymers to endow desirable rheological properties for 3D bioprinting. Crosslinking of thiolated Pluronic F-127 with dopamine-conjugated (DC) gelatin and DC hyaluronic acid through a thiol-catechol reaction resulted in thermally gelling bioinks with Herschel-Bulkley fluid rheological behavior. Microextrusion 3D bioprinting was used to fabricate free-standing cell-laden tissue constructs. The bioinks exhibited flattened parabolic velocity profiles with tunable low shear regions. Two pathways were investigated for curing the bioink: chelation and photocuring. The storage modulus of the cured bioinks ranged from 6.7 to 11.7 kPa. The iron (III) chelation chemistry produced crosslinked neural tissues of relatively lower storage modulus than the photocuring approach. In vitro cell viability studies using the 3D bioprinted neural tissues showed that the cured bioink was biocompatible based on minimal cytotoxic response observed over seven days in culture relative to control studies using alginate hydrogels. Rodent Schwann cell-, rodent neuronal cell-, and human glioma cell-laden tissue constructs were printed and cultured over seven days and exhibited comparable viability relative to alginate bioink controls. The ability to fabricate soft, free-standing 3D neural tissues with low modulus has implications in the biofabrication of microphysiological neural systems for disease modeling as well as neural tissues and innervated tissues for regenerative medicine.

Engineered 3D Polymer and Hydrogel Microenvironments for Cell Culture Applications

“…Modifications of hyaluronic acid can help to improve the viscoelastic and rheological properties during extrusion as well as the stability after extrusion-based fabrication. For example hyaluronic acid has been modified with hydrazide and aldehyde groups allowing shear thinning and self-healing of the bioink due to formation of physical crosslinks mediated by hydrazine bonds [207]; crosslinked with Pluronic F-127 and dopamine conjugated gelatin to introduce Herschel-Bulkley rheological properties and improve thermal gelling [208]; and combined with methacrylated collagen [209] for improving the printing process.…”

“…Hyaluronic acid and modified hyaluronic acid show good cyto-and bio-compatibility [207,210], as it is naturally a part of the extra-cellular matrix. Extruded hyaluronic acid structures show excellent bioactivity, with hyaluronic acid based hydrogel scaffolds for improving cell viability [208], promotion of stromal cell elongation with applications in building liver models for drug screening [209], retinal cells culturing [216], immobilization of peptides for mesenchymal stem cell culturing with high angiogenic and osteogenic activity [217], deposition of regenerative scaffolds directly during surgery [218], and supporting human adipose progenitor cell and stromal cell adhesion and proliferation [211]. Hyaluronic acid combined with alginate and fibrin has been bioprinted for peripheral nerve tissue regeneration, showing good Schwann cell elongation [219].…”

“…Gelatin combined with other hydrogels has been used to optimize printability, improving shape retention during printing. For example, gelatin-alginate [240,[319][320][321], Laponite [322], nanoclays [323], and synthetic polymers such as Pluronic F-127 [208] and PCL [324], have all been studied. Alginate dialdehyde-gelatin scaffolds were printed in the presence of a cross-linker for reaching feature sizes of~500 µm [245], while alginate-GelMA interpenetrating networks via UV crosslinking of GelMA followed by Ca crosslinking of alginate was reported [321].…”

“…Hyaluronic acid (4.3.1) [207][208][209]; Inclusion of GelMA, feature size 500 µm [210]; Cryogel E: 2-2.5 kPa [211]; Post-curing via UV, E: 1.3-10.6 kPa [213] Feature size 300 µm (SLA) [214]; Compressive E: 780 kPa [215] Cartilage tissue engineering and human adipose stem cells [215]; stromal cell elongation and drug screening [209]; retinal cell culturing [216]; hMSCs [217]; human adipose progenitor and stromal cells [211]; Schwann cells [219] Chitosan (4.3.2) [222,223]; Feature size 50 µm [224] [225,226]; Feature size 50 µm (SLA), E: 160-680 kPa [173]; Feature size 400 nm (TPP) [228]; Inclusion of HA [229] Anti-bacterial [230,231]; wound dressings [232]; skin tissue engineering [231]; bone tissue engineering [229]; pluripotent stem cells for neural tissue engineering [233]; articular cartilage tissue engineering [234]; skin constructs [235] Alginate (4.3.3)…”

“…Inclusion of alginate [240,[319][320][321], Laponite [322], nanoclays [323], Pluronic F-127 [208], PCL [324]; Feature size 500 µm [245] [ [313][314][315][316]; Feature size 300 µm (SLA), Compressive E: 0.5-18 MPa [328] Mouse planta dermis [319]; dental pulp stem cells [320]; hMSCs and amniotic epithelial cells [240]; chondrocytes [214,327] Silk ( Mouse articular chondrocytes [342]; human fibroblasts [338]; porcine chondrocytes [340]; hMSCs [344,345]; human mesenchymal progenitor cells [346]…”