Subcutaneous DPSC transplants in immunocompromised mice (A-C) and characterization of DPSC-mediated dentinogenesis in vivo (D-F). (A) Four wks after transplantation, DPSCs differentiated into odontoblasts (open arrows) responsible for new dentin (D) formation on the surface of the HA/TCP (HA). (B,C) At 8 and 16 wks post-transplantation, respectively. (D) Newly formed reparative dentin-like structure (ND) attached to the surfaces of human dentin in DPSC/dentin transplants. BV, blood vessels; CT, connective tissue; dentinogenic cells (black arrowheads). DPSCs formed reparative dentin-like structure containing entrapped cells (open arrowheads). (E) In DPSC/dentin transplants, dentinogenic cells (open arrows) and trapped cells (open arrowheads) within the newly formed reparative dentin-like structure (ND) were immunoreactive to human DSP antibody, as was the pre-existing dentin (black arrows). (F) Staining of human-specific anti-mitochondria antibody, showing the human origin of DPSCs (open arrows). Bar, 40 µm in A-C, 20 µm in D-F (adapted from Batouli et al., 2003).

In vitro neurogenesis of SHED (A-D), transplanted SHED into immunocompromised mice (E-H) and into the mouse brain (I-K). (A,B) Toluidine blue staining of the altered morphology of SHED after neurogenic induction. (C,D) Immunopositive staining for MAP2 and Tau on dendrites and axons (arrows), respectively. (E,F) Eight wks after transplantation into the subcutaneous space, SHED differentiate into odontoblasts (open arrows) and form dentin-like structure (D) on the surfaces of HA. The same field is shown for human-specific alu in situ hybridization, indicating the human origin of odontoblasts (open arrows in F). (G) Immunohistochemical staining of DSPP on the regenerated dentin (black arrows). (H) Newly generated bone (B) by host cells in the same SHED transplant shows no reactivity to the DSPP antibody. (I-K) Neurogenically induced SHED injected into the dentate gyrus of the hippocampus of immunocompromised mice for 10 days. (I) NFM (red) and (J) human-specific anti-mitochondrial antibody (green) and (K) merged images showing co-localization of the two (adapted from Miura et al., 2003, with permission).

The anatomy of the human apical papilla (A-C) and dentinogenesis of SCAP in immunocompromised mice (D-F). (A) An extracted human third molar depicting root attached to the root apical papilla (open arrows) at the developmental stage. (B) Hematoxylin and eosin staining of human developing root (R) depicting epithelial diaphragm (open arrows) and apical cell-rich zone (open arrowheads). (C) Harvested root apical papilla for stem cell isolation. (D) Eight wks after transplantation, SCAP differentiated into odontoblasts (arrows) that formed dentin (D) on the surfaces of a HA carrier. (E) SCAP differentiated into odontoblasts (arrows) are positive for anti-human specific mitochondria antibody staining. (F) Immunohistochemical staining of SCAP-generated dentin (D) showing positive anti-DSP antibody staining (arrows) (adapted from Sonoyama et al., 2006, 2008, with permission).

Generation of cementum-like structure and collagen fibers by PDLSCs in immunocompromised mice (A,B) and PDLSCs in periodontal tissue repair in immunocompromised rats (C-E). (A,B) Transplanted PDLSCs formed cementum-like structures (C) that connected to newly formed collagen fibers (dashed lines), similar to the structure of Sharpey’s fibers, and generated a substantial amount of collagen fibers (arrows in B). (C-E) Staining of human-specific anti-mitochondria antibody showing human PDLSCs located in the PDL compartment (arrowheads in C), involved in the attachment of PDL to the tooth surface (arrows in D), and participating in the repair of alveolar bone (arrows in E) and PDL (arrowhead in E) (adapted from Seo et al., 2004, with permission).

Swine SCAP/PDLSC-mediated bio-root engineering. (A) Extracted minipig lower incisor and root-shaped HA/TCP carrier loaded with SCAP. (B) Gelfoam containing PDLSCs (open arrow) to cover the HA/SCAP (black arrow) and implanted into the lower incisor socket (open arrowhead). (C) HA/SCAP-Gelfoam/PDLSCs were implanted into a newly extracted incisor socket. A post channel was pre-created inside the root-shaped HA carrier (arrow). (D) Three months after implantation, the bio-root was exposed and a porcelain crown inserted. (E) Four wks after fixation, the porcelain crown was retained after normal tooth use. (F) After 3 months’ implantation, the HA/SCAP-Gelfoam/PDLSC implant formed a hard root structure (open arrows) in the mandibular incisor area, as shown by CT scan image. A clear PDL space was found between the implant and surrounding bony tissue (arrowhead). (G, H) H&E staining showed that implanted HA/SCAP-Gelfoam/PDLSC contains newly regenerated dentin (D) and PDL tissue (PDL) on the outside of the implant. (I) Compressive strength measurement showed that newly formed bio-roots have compressive strength much higher than that of the original HA/TCP carrier (*P = 0.0002), but lower than that in natural swine root dentin (*P = 0.003) (NR, natural minipig root; BR, newly formed bio-root; HA, original HA carrier) (adapted from Sonoyama et al., 2006, with permission).