Page 92 - IJB-9-1

International Journal of Bioprinting Progress in bioprinting of bone

encapsulated in a Laponite -alginate-methylcellulose hTERT-overexpressing MSCs, fetal MSCs from the

hydrogel to create bioprint scaffolds, showing 90% cell umbilical cord MSCs, BMSCs, and ADSCs, among which
viability on day 7 of culture with decreased ALP expression ADSCs showed the greatest osteogenic differentiation
and increased calcium mineral deposition over 21 days potential indicated by the ALP assay, which were then
of culture [113] . MSC-laden scaffolds were implanted in chosen for bioprinting with the F/G/H/Gl hydrogel. Chiesa
athymic BALB/c mice, where extensive mineralization et al. [118] fabricated a vascularized bone model using gelatin-
was observed after 4 weeks of implantation, while bone nanohydroxyapatite (Gel-nHAp), hMSCs, and HUVECs.
volume and bone density increased significantly from 2 Gel-nHAp was first extruded followed by hMSCs being
to 8 weeks. Furthermore, subcutaneous implantation of seeded on scaffolds and osteogenically differentiated for
BMP-2-containing bioprinted scaffolds in mice exhibited 2 weeks. Next, lentiviral-GFP transfected HUVECs were
increased level of glycosaminoglycans (GAGs) deposition placed into the macropores of 3D bioprinted scaffolds.
and mineralization. The assembly of a complex capillary-like network was
Alginate is readily processable using cross-linking observed, and vascular lumen formation and osteogenic
mechanisms such as ionic interaction, cell cross-linking, differentiation were confirmed by immunostaining and
photopolymerization, and Schiff-base reaction [114] . The gene expression.
prevalent method for alginate gelation is to combine Although gelatin is easily accessible and has good
the alginate with divalent cations, such as Ca , Mg , biocompatibility, its low viscosity, low yield stress,
2+
2+
Ferrous (Fe ), Barium (Ba ), or Sr . In the cell cross- and relatively long cross-linking time during or after
2+
2+
2+
linking mechanism, the ligands (e.g., RGD) were grafted bioprinting lead to poor shape retention properties,
onto alginate for cell adhesion [111] . As cells were loaded resulting in difficulty in creating 3D reliable structures
into the RGD-modified alginate, the receptors on the cell with an interconnected pore network [118] . To overcome this
surface can bind to ligands of the modified alginate. In obstacle, non-modified gelatin can be bioprinted below
addition, some researchers exploited photopolymerization the melting temperature (usually below 4°C) to increase
to solidify methacrylated alginate by employing UV the viscosity for extrusion [119] or with the aid of a sacrificial
irradiation [115] . Alginate was also prepared in the form of frame to ensure sufficient time for cross-linking with cross-
microspheres to deliver growth factors, proteins, and drugs linkers, such as genipin and transglutaminase [118] .
in tissue engineering [114] . Inspired by the advantage of the
alginate microsphere, Wu et al. embedded the alginate 3.1.3. Alginate/Gelatin-based composite bioinks
microspheres within the cell aggregates to generate porous To integrate the benefits of alginate (fast cross-linking)
tissue strands with high cell density [116] . The incorporation and gelatin (good biocompatibility), alginate/gelatin-
of alginate microspheres facilitated the permeation of based composite bioinks have drawn a lot of attention.
oxygen and nutrition, promoting cell viability within the As reported by Neufurth et al. , a polyphosphate Ca salt
[75]
cell aggregates, which offers new insight into scaffold-free overlay (polyP·Ca -complex) was applied to the bioprinted
2+
biofabrication. alginate/gelatine/SaOS-2 cell scaffold to modulate the
biological result of the construct. PolyP·Ca -complex was
2+
3.1.2. Gelatin-based composite bioinks surprisingly found to significantly increase the ability of
As mentioned in section 2.2., bioinks whose main cells to proliferate in the underlining hydrogel. The hardness
composite is gelatin have also been reported by several and mineralization of cell-laden alginate/gelatin hydrogels
studies. Das et al. used silk in a bioprinted bone study, significantly increased when the overlaid polymer was
[56]
where silk fibroin-gelatin (SF-G) bioink was embedded present. A follow-up study by Wang et al. [120] investigated
with hTMSC, and in situ cytocompatible gelation occurred the effect of bioglasses including polyP·Ca -complex,
2+
(enzymatic cross-linking using mushroom tyrosinase silica, and biosilica on the mineralization of SaOS-2 cells
and physical cross-linking using sonication). SF-G bioink embedded in a gelatine/alginate hydrogel. Bioglass particles
caused no harm to the cells, and a higher chondrogenic and significantly enhanced the mineralization ability of the
adipogenic potential was observed in the tyrosinase cross- entrapped cells, as evidenced by staining with alizarin red
linked groups, whereas a higher osteogenic differentiation S, while element analysis of mineral nodules formed by
was observed in the sonication cross-linked groups. Using the SaOS-2 cells indicated a gathering of minerals. Wüst
[51]
six hydrogel blends of fibrin, gelatin, HA, and glycerol et al. took advantage of the thermal gelation of gelatin
(F/G/H/Gl), Wehrle et al. [117] identified the best hydrogel and the irreversible cross-linking of alginate to develop
blend which showed high cell viability (>96%) and cell a two-step process. Cell viability was 85% after 3 days of
proliferation. The osteogenic differentiation process culture after bioprinting when MSCs were mixed into the
was carried out in the hydrogels with immortalized hydrogel precursor. Furthermore, a two-phased structure

Volume 9 Issue 1 (2023) 84 https://doi.org/10.18063/ijb.v9i1.628