Investigation of rheology, printability, and biocompatibility of N,O-carboxymethyl chitosan and agarose bioinks for 3D bioprinting of neuron cells

A Guide to Polysaccharide-Based Hydrogel Bioinks for 3D Bioprinting Applications

“…Among the various seaweed-derived polysaccharides, alginic acid (and particularly its salt form, sodium alginate) [ 55 , 56 , 57 , 58 , 59 , 60 , 61 , 62 , 63 , 64 , 65 , 66 ], carrageenan [ 67 , 68 , 69 , 70 , 71 , 72 , 73 , 74 ], and agarose [ 75 , 76 , 77 , 78 , 79 ] have been widely used as polymeric matrices (either solely or in combination with other polysaccharides or proteins) for the development of hydrogel-based bioinks for 3D bioprinting, as outlined in Table 1 .…”

“…However, the use of agarose hydrogels for bioprinting is still very limited, mainly because of their biological inertness, resulting in poor cell viability in long-term cultures [ 91 ]. In fact, to the best of our knowledge, only two studies have been reported so far [ 75 , 79 ]. The first, by López-Marcial et al [ 75 ], compared the mechanical and rheological properties, including yield stress, storage modulus, and shear thinning, of single agarose and of agarose–alginate hydrogels with those of Pluronic hydrogels, commonly used in bioinks design, to assess their suitability for extrusion bioprinting of cartilage constructs.…”

“…However, the combination of agarose with alginate in a 3:2 ratio improved print fidelity and demonstrated excellent cell viability for auricular cartilage cells, which was maintained over a 28-day culture period post-bioprinting (>70% cell viability at the end of 28 days) [ 75 ]. More recently, Butler et al [ 79 ] explored the combination of agarose with N , O -carboxymethyl chitosan (NOCC) in different ratios (80:20, 60:40, 40:60, and 20:80) for the development of bioinks laden with neuron cells (neuro2A) ( Figure 2 C). The rheological properties and printability by extrusion of these bioinks were mainly influenced by the NOCC content, with the agarose-NOCC 40:60 and agarose-NOCC 20:80 bioink formulations presenting the highest storage modulus (20–25 Pa) and printability, evaluated by the assessment of the printability number.…”

“…The rheological properties and printability by extrusion of these bioinks were mainly influenced by the NOCC content, with the agarose-NOCC 40:60 and agarose-NOCC 20:80 bioink formulations presenting the highest storage modulus (20–25 Pa) and printability, evaluated by the assessment of the printability number. However, comparing these two formulations, the post-bioprinting cell viability of neuro2A for 14 days was higher (100%) for the scaffolds produced with the bioink with the highest content of agarose (agarose-NOCC 40:60), which the authors considered to be the best blend in terms of a compromise between mechanical performance and cell viability [ 79 ]. This study opens the possibility for future developments in the field of agarose-based bioinks by exploiting the combination of agarose with bioactive compounds or polymers [ 94 , 95 ], as extensively explored for alginate-based bioinks and previously highlighted in Section 2.1.1 .…”

Biomaterials-based strategies for in vitro neural models

“…92 It is also an excellent structural component for bioprinting of neurons within rheologically-tunable constructs. 93 Blends of agarose with the electroactive molecule aniline pentamer have also shown promise to support cell viability and attachment. 94 In this context, a particularly interesting use of agarose has been in the photothermal microfabrication of controlled neuronal networks in which an infrared laser is used to etch pathways into the hydrogels to create conduits to spatially direct and tune axonogenesis and synaptogenesis.…”