Regeneration of Tooth Roots With Borrowed Stem Cells in Pigs


Because a recent post about tooth-making stem cells in alligators generated so much interest, I found another recent paper that reports the regeneration of the tooth root structure in pigs. This is a proof-of-concept paper that demonstrated the feasibility of such a procedure.

The journal is Stem Cells and Development and the research team is from the Beijing Key Laboratory of Tooth Regeneration and Function Reconstruction in Beijing, China. The corresponding author is Songlin Wang from the Molecular Laboratory for Gene Therapy and Tooth Regeneration.

Tooth loss represents a growing problem in an aging population. Dental implants provide one solution, but without a good jaw bone into which these implants can be attached, implants have little chance of staying put. Regenerating a tooth root that can support a natural or artificial crown is the most important part of the tooth in maintaining tooth function.

In previous work, Wang and his collaborator Songtao Shi from UCLA have shown that stem cells from root apical papilla and periodontal ligament stem cells from exfoliated teeth can coat bioengineered surfaces and form tooth structures that can support artificial crowns in miniature pigs (see Sonoyama et al., PLoS One 1:e79-e92). However, aged patients sometimes have bone marrow stem cells that do not grow well in culture and respond poorly to bioengineering protocols. Therefore, Wang and his crew sought to demonstrate that mesenchymal stem cells from donor animals (allogeneic stem cells) could provide the same kind of benefit.

The two stem cell populations used in this paper was dental pulp stem cells (DPSCs) and periodontal ligament stem cells (PDLSCs). The DPSCs were cultured from exfoliated minipig teeth and grown in culture for two or three passages. The culture medium used, as far as I can tell, was the same one used the Gronthos in his PNAS paper that reported the isolation and characterization of DPSCs. That medium was a modified Eagle’s medium supplemented with 20% Fetal Calf Serum and 100 μM L-ascorbic acid 2-phosphate, 2 mM L-glutamine, 100 units/ml penicillin, and 100 μg/ml streptomycin. Gronthos then grew his cells at 37°C in 5% CO2 (see S. Gronthos, et al PNAS 97(25): 13625–13630). After 2-3 passages, the DPSCs were seeded on a hydroxyapatite tricalcium phosphate scaffold and grown in a bioreactor for 5-7 days

PDLSCs were grown in culture with approximately the same cocktail as the DPSCs and then plated on 60 mm dishes with vinylene carbonate (Vc). Vc induces the PDLSCs to grow s sheets that could be used to wrap the hydroxyapatite tricalcium phosphate structures that had been seeded with DPSCs.

These wrapped structures were implanted into the gums of minipigs and then used to secure dental implants.

Tooth Root Regeneration

After 6 months, the implants were assessed as was the integrity and strength of the surrounding tissue.

Gross, radiographic, and histological analysis of the bio-root 6 months after transplantation. (A, C) Gross view of the general shape of HA/TCP and the bio-root 6 months after transplantation (ellipse). (B, D) X-rays revealed that HA/TCP formed tissues without an obvious dental structure (ellipses), but the HA/TCP/DPSC/PDLSC sheet implant formed a hard root structure (ellipses). (E, F) No obvious boundary was observed between newly regenerated tissue and bone in the microcomputed tomography scan image of the HA/TCP group. (G, H) A hard root structure (arrows) was present and a clear PDL space found between the implant and surrounding bony tissue (triangle arrows). (I–K) HE staining showed some bone formation and HA/TCP remaining in the HA/TCP group (I), and PDL-like tissues were generated parallel to the dentin-like matrix structure in the autologous group (J) and allogeneic group (K). (L) Semiquantitative analysis showed that mineralized tissue regeneration capacity of autologous or allogeneic groups was significantly higher compared with the HA/TCP group. Percentage of mineralized tissues at 6 months after crown restoration was significantly higher than that before crown restoration in both autologous and allogeneic groups. No significant difference of regenerated mineralized tissue percentages was found between autologous and allogeneic groups. Scale bar: (I–K) 200 μm. B, bone; HA/TCP, hydroxyapatite/tricalcium phosphate; PDL, periodontal ligament; MT, mineralized tissue. *P<0.01 compared with autologous or allogeneic groups; #P<0.01 compared with autologous or allogeneic groups after crown restoration.
Gross, radiographic, and histological analysis of the bio-root 6 months after transplantation. (A, C) Gross view of the general shape of HA/TCP and the bio-root 6 months after transplantation (ellipse). (B, D) X-rays revealed that HA/TCP formed tissues without an obvious dental structure (ellipses), but the HA/TCP/DPSC/PDLSC sheet implant formed a hard root structure (ellipses). (E, F) No obvious boundary was observed between newly regenerated tissue and bone in the microcomputed tomography scan image of the HA/TCP group. (G, H) A hard root structure (arrows) was present and a clear PDL space found between the implant and surrounding bony tissue (triangle arrows). (I–K) HE staining showed some bone formation and HA/TCP remaining in the HA/TCP group (I), and PDL-like tissues were generated parallel to the dentin-like matrix structure in the autologous group (J) and allogeneic group (K). (L) Semiquantitative analysis showed that mineralized tissue regeneration capacity of autologous or allogeneic groups was significantly higher compared with the HA/TCP group. Percentage of mineralized tissues at 6 months after crown restoration was significantly higher than that before crown restoration in both autologous and allogeneic groups. No significant difference of regenerated mineralized tissue percentages was found between autologous and allogeneic groups. Scale bar: (I–K) 200 μm. B, bone; HA/TCP, hydroxyapatite/tricalcium phosphate; PDL, periodontal ligament; MT, mineralized tissue. *P

As you can see in panel E and F, control implants that had no cells and only hydroxyapatite calcium triphosphate showed no tooth-like structures, but in G and F, the structures with cells showed a consistent tooth structure with a periodontal ligament (see broad arrow).  In panels J and K, there is obvious bone formation with periodontal ligament in the autologous and allogeneic stem cell transplants.

Cross sections of the implants also showed that not only did these structures look like real tooth root structures, but they contained structures proteins indicative of real tooth root structures.  Dentin sialophosphoprotein (mercifully abbreviated to DSPP) is present in the cell-seeded implants, but in on the hydroxyapatite calcium triphosphate-only implants.

Clinical assessment of implants failed to detect any gingivitis or periodontal disease associated with the implants.

This experiment shows that stem cell-seeded scaffolds can regenerate tooth root structures.  Since this worked in minipigs and not simply rodents, these results strongly suggest that such a strategy could work in humans.  Clinical trials anyone?

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mburatov

Professor of Biochemistry at Spring Arbor University (SAU) in Spring Arbor, MI. Have been at SAU since 1999. Author of The Stem Cell Epistles. Before that I was a postdoctoral research fellow at the University of Pennsylvania in Philadelphia, PA (1997-1999), and Sussex University, Falmer, UK (1994-1997). I studied Cell and Developmental Biology at UC Irvine (PhD 1994), and Microbiology at UC Davis (MA 1986, BS 1984).

2 thoughts on “Regeneration of Tooth Roots With Borrowed Stem Cells in Pigs”

  1. I responded immediately to a news item in the daily paper (News Journal, Daytona, Beach, FL) dated September 27, 2002 which indicated “TEETH RESEARCH MAY REPLACE DENTURES”
    At that time, I was eager to be a trial subject. It is now August 25, 2013, and I face implants. Might I qualify?? My name is Charlotte Venetianer, PO Box 351054, Palm Coast, Florida, (386) 445-5824,
    cvenetianer@cfl.rr.com.

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