Fig. 4: Bioprinting of functional living tissues with cell-location-driven features enabled by GRACE.
a, Diagram of experiment process depicting the scanning of the volume, feature-driven model generation around the torus and printing of the scaffold. b,c, 3D segmentations of the resultant structures printed around the extruded toruses, both targeted and random, also showing light-sheet micrographs of the scaffold cross-sections at two planes. Scale bars, 2 mm. d, Graph showing the amount of insulin stored and released by iβ-cells by means of the bioluminescent reporter NanoLuc (Nluc) for the GRACE print, as well as for the random and bulk controls (mean ± s.d., ANOVA, n = 6, **P < 0.01, ***P < 0.001, F = 55.74). e, A light-sheet section of the construct immediately following printing with the automatically aligned femur-cartilage model showing distinct osteal and chondral regions. Scale bar, 2 mm. f, The histological sections following 4 weeks of maturation highlight the presence of a mineralized compartment (von Kossa positive, brown/black staining) and of glycosaminoglycans-rich cartilaginous tissue (SafO, red staining) (n = 3). Scale bars, 500 μm (left panels); 200 μm (right panels). g, 3D model of FLight construct generated with GRACE after scanning. Printing was performed around a subset of each spheroid population to avoid overcrowding. h, Resultant light-sheet 3D reconstruction after printing and washing. i, Filamentous construct is visible surrounding the spheroid. Scale bar, 100 μm. j, Discrimination of spheroids is evident in the schlieren image (captured immediately after printing) with fluorescence overlay, with the spheroids precisely positioned through the central axis of each star or circle FLight structure. Scale bar, 2 mm.
