Fig 1.
Micro CT analysis of hydrogels following 28 days in vivo implantation.
Tissue volume (A), percentage bone volume (B), bone surface to volume ratio (C), trabecular number (D), trabecular thickness (E) and trabecular separation (F) were all assessed between growth factor groups. Emboldened columns depict statistically significant intragroup differences between those with and without Stro-1+ cell incorporation. Asterisks depict statistical difference between the group above which the asterisk is positioned and all the other groups; if positioned centrally above both groups with and without Stro-1+ cell incorporation, statistical difference was observed for both compared across all groups. Error bars are S.D. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001.
Fig 2.
Hydrogel mineralisation between growth factor groups following 28 days in vivo implantation.
Groups included irradiated ALG/ECM (A), ALG/Col (B), ALG/ECM (C), ALG/ECM HSA (D), ALG/ECM HSA/VEGF (E), ALG/ECM HSA/TGF-β3 (F), ALG/ECM HSA/BMP-2 (G), ALG/ECM HSA/PTHrP (H) and ALG/ECM HSA/VitD3 (I). Images were taken at high (scale bar is 500 μm) and low magnification (box inserts–scale bar is 50 μm).
Fig 3.
Histological analysis of hydrogels stained with Alcian blue/Sirius red.
Hydrogels were subcutaneously implanted within immunodeficient for 28 days. Colour quantification was through the use of an optimised Image J macro (S2 Fig). Blue indicated proteoglycan deposition and residual hydrogel (A), purple indicated collagen deposition within the hydrogel (B) and red indicated tissue invasion (C). Emboldened columns depict statistically significant intragroup differences between those with and without Stro-1+ cell incorporation. Asterisks depict statistical difference between the group above which the asterisk is positioned and all the other groups; if positioned centrally above both groups with and without Stro-1+ cell incorporation, statistical difference was observed for both compared across all groups. Error bars are S.D. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001.
Fig 4.
Histological analysis of hydrogels stained with Von Kossa.
Hydrogels were subcutaneously implanted within immunodeficient mice for 28 days. Colour quantification was through the use of an optimised Image J macro (S2 Fig). Black indicates mineralised tissue (A) and pink indicates cell invasion (B). Emboldened columns depict statistically significant intragroup differences between those with and without Stro-1+ cell incorporation. Asterisks depict statistical difference between the group above which the asterisk is positioned and all the other groups; if positioned centrally above both groups with and without Stro-1+ cell incorporation, statistical difference was observed for both compared across all groups. Error bars are S.D. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001.
Fig 5.
Histological analysis of hydrogels stained with Goldner’s Trichrome.
Hydrogels were subcutaneously implanted within immunodeficient mice for 28 days. A square grid (200 μm) overlay was used to quantify erythrocytes (A) and assess vascularisation (B). Error bars are S.D. * P ≤ 0.05.