Fig 1.
Schematic representation for the methacrylation reaction of HA and photograph of cross-linked MeHA gel.
A. HA reacted with methacrylic anhydride in an aqueous environment in presence of DMF, yielding the photo-crosslinkable MeHA. B. Hydrogel disc prepared from 3% MeHA (DS = 5%). Scale bar represents 1 mm.
Table 1.
MeHA hydrogel characteristics before and after application of UV irradiation.
Fig 2.
MeHA hydrogel swelling and degradation as a function of gel concentration.
A. Increase of hydrogel wet weight (swelling) during 3 weeks incubation at 37°C, which was most pronounced at lower MeHA concentrations. B. Decrease of hydrogel dry weight during incubation. C. Decrease of hydrogel dry weight followed in time in the presence of hyaluronidase. Gel degradation was slower with increasing MeHA concentration. Data presented as mean ± SD, n = 5 for all measurements.
Fig 3.
MSC survival and morphology in MeHA hydrogels.
A. Survival of human MSCs after 1 and 21 days of MeHA hydrogel encapsulation. N/A: The 1% (w/v) hydrogels disintegrated before day 21. B. Cell morphology in the MeHA hydrogels after 21 days of culturing. Representative pictures are shown. Scale bars = 100 μm. * Represents p<0.05.
Fig 4.
Osteogenic differentiation of MSCs in MeHA hydrogels.
A. Calcium mineralization per mg of hydrogel scaffold after 21 days of culturing in αMEM (control, white bars) or αMEM supplemented with BMP-2 (black bars). Experiments performed in duplicate and replicated with 3 MSC donors. Data shown as mean ± SD. N/A: The 1% (w/v) hydrogels without BMP-2 disintegrated before the day 21 time point. * Represents p<0.05. B. Whole-mount Alizarin red staining of the MeHA hydrogels after 21 days of incubation in the presence of BMP-2. At lower MeHA concentration (1–1.5% (w/v)) gels, only some cells stained red. When the gel’s MeHA polymer concentration was increased, larger areas around the cells stained red, indicating calcium deposition into the surrounding matrix. Most intense staining is seen in the 2.5% (w/v) gels. Representative pictures are shown. Scale bars = 100 μm.
Fig 5.
Porous cubes (upper rows) and non-porous human L3 vertebrae shapes (lower rows) were 3D bioprinted, using the designs shown in the left column, and subsequently UV irradiated. After crosslinking, scaffolds were lifted using a spatula (horizontal bar) to test handling. Scale bars = 500 μm.
Fig 6.
Optimal MeHA hydrogel selection.
Combining MSC survival (yellow), osteogenic differentiation (green) and 3D printability (dotted grey line) data, reveals the boundary conditions for selection of optimal MeHA gel composition (dotted red line) for bone tissue engineering.