Bioorthogonally Cross-Linked Hyaluronan-Laminin Hydrogels for 3D Neuronal Cell Culture and Biofabrication

Abstract

Laminins (LNs) are key components in the extracellular matrix of neuronal tissues in the developing brain and neural stem cell niches. LN-presenting hydrogels can provide a biologically relevant matrix for the 3D culture of neurons towards development of advanced tissue models and cell-based therapies for the treatment of neurological disorders. Biologically derived hydrogels are rich in fragmented LN and are poorly defined concerning composition, which hampers clinical translation. Engineered hydrogels require elaborate and often cytotoxic chemistries for cross-linking and LN conjugation and provide limited possibilities to tailor the properties of the materials. Here we show a modular hydrogel system for neural 3D cell culture, based on hyaluronan (HA) and poly(ethylene glycol) (PEG), that is cross-linked and functionalized with human recombinant LN 521 using bioorthogonal copper-free click chemistry. Encapsulated human neuroblastoma cells demonstrate high viability and grow into spheroids. Neuroepithelial stem cells (lt-NES) cultured in the hydrogels can undergo spontaneous differentiation to neural fate and demonstrate significantly higher viability than cells cultured without LN. The hydrogels further support the structural integrity of 3D bioprinted structures and maintain high viability of syringe extruded lt-NES, which can facilitate the development of advanced neuronal tissue and disease models and translation of stem cell-based therapies.

Astrocyte 3D culture and bioprinting using peptide functionalized hyaluronan hydrogels

“…However, peptide functionalization had limited effect on promoting proliferation. These results align with our previous observations for the culture and differentiation of neural cells [ 26 ]. In addition to the culture of primary astrocytes, we explored the often-used astrocytic model of the glioblastoma cell line U87.…”

“…Due to the loss of some BCN groups, the peptide functionalized hydrogels showed a minor but not statistically significant decrease in the stiffness of the hydrogels ( Figure 1(a,b) ). This is similar to observations in prior work where functionalization of the hydrogels had limited effects on the viscoelastic properties [ 26 ]

Figure 1. a) i.

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