Figure 3.

Implantation of cell-laden and acellular Matrigel beads to intertransects of spinal cords. (a) Schematic of the production of cell-laden Matrigel beads by using the CTM technique. (b) Collected Matrigel beads in DMEM/F12 medium. The oil residue was removed by gentle pipetting. (c, d) Bright-field (c) and fluorescence (d) images of the collected monodisperse cell-laden Matrigel beads (~500 μm in diameter). Each bead was encapsulated with EGFP-labeled NPCs. (e) Schematic of injection of cell-laden Matrigel beads in (d) to the intertransects of a spinal cord. (f) The cell-laden Matrigel beads were injected into the space where a 4-5 mm spinal cord tissue in length had been cut and removed. (g) The harvested spinal cord tissue at 8 weeks postimplantation. The dotted circle marks the boundary of implanted beads. (h–m) Confocal sagittal imaging of the immunostained spinal cord tissue. DAPI shows the nuclei. Neurofilament (NF, green, labeled by anti-NF antibody) indicates the axons (h–k). The enlarged area in (h) shows the EGFP-labeled NPCs (pseudocolor in grayscale) and axons (green) (i). Zoom-in view shows the nonoverlapped but physically proximate EGFP-NPCs (indicated by yellow arrows) and axons (j, k). White dot lines in (i) indicate the interfaces between the grafted and host tissue. (l, m) The fluorescence distribution of GFAP, the marker for astrocytes, and DAPI. White dot lines represent the boundaries of the glial scar. (n–u) Schematic (n) and follow-up characterization (o–u) of injection of acellular Matrigel beads to the intertransects of a spinal cord. (v–BB) Schematic (v) and follow-up characterization (w–BB) of implantation of molded acellular Matrigel trunk to the intertransects of a spinal cord. Scale bars, 20 μm (s, t), 100 μm (c, d, i, j, k, l, m, r, u, z, AA, BB), and 1 mm (h, q, y).