Dental pulp and dentin tissue engineering and regeneration: advancement and challenge

Abstract

Hard tissue is difficult to repair especially dental structures. Tooth enamel is incapable of self-repairing whereas dentin and cementum can regenerate with limited capacity. Enamel and dentin are commonly under the attack by caries. Extensive forms of caries destroy enamel and dentin and can lead to dental pulp infection. Entire pulp amputation followed by the pulp space disinfection and filling with an artificial rubber-like material is employed to treat the infection -- commonly known as root canal or endodontic therapy. Regeneration of dentin relies on having vital pulps; however, regeneration of pulp tissue has been difficult as the tissue is encased in dentin without collateral blood supply except from the root apical end. With the advent of modern tissue engineering concept and the discovery of dental stem cells, regeneration of pulp and dentin has been tested. This article will review the recent endeavor on pulp and dentin tissue engineering and regeneration. The prospective outcomes of current advancements and challenges in this line of research are discussed.

Fig. 1

Tooth structure loss resulting from caries, root canal treatment and restoration. (A) Tooth is decayed (c) and causes irreversible pulpitis that requires root canal therapy. p, pulp. (B) Tooth decay is completely removed and the root canal space enlarged (cleaned and shaped) and filled with gutta percha (gp). (C) A post is placed in a canal and core material placed to fill the coronal space. The natural crown is then prepared/cut into a specific configuation and an artificial crown is manufactured for insertion. (D) the artificial crown is cemented onto the tooth as a final restoration and the tooth is back to its function. (E) If without good and more rigorous care, tooth decay can recur. Tooth may also undergo fracture (dashed line) due to loss of structure which weakens its capacity to withhold mechanical stress. In this condition, the tooth is not salvageable and to be extracted.

Fig. 2

Micrographes of human dental pulp. (Top) Light micrograph of the pulpodentin complex from the pulp horn region of a decalcified human tooth following staining with hematoxylin and eosin. The pulpodentin complex consists of a highly differentiated tissue with a consistent morphological pattern that includes dentin and dentinal tubules (D), the odontoblast layer (O), the cell free zone (zone of Weil; CF), and the cell rich zone (CR). Other components of the pulp include Schwann cells (S) that enwrap nerve fibers, blood vessels (BV), and fibroblasts (F). (Bottom) Confocal micrograph of the pulpodentin complex from the pulp horn region of the same sample as seen above shows immunofluorescence for N52 (green; identifies nerve fibers) and von Willebrand factor (red; identifies endothelial cells associated with blood vessels), whereas cellular nuclei are stained with ToPro-3 (blue). The nerves fibers form an extensive plexus just below the odontoblasts. Some nerve fibers pass through the odontoblastic layer, where they enter and continue within dentinal tubules for about 100 microns (white arrow) before terminating. Both images courtesy of Dr. Michael Henry, University of Texas Health Science Center at San Antonio).

Fig. 3

Source of dental stem cells and use for dental tissue regeneration. Note DPSCs are from pulp of permanent teeth, SHED from exfoliated primary teeth.

Fig 4

Lost portion of pulp is replaced by periapical tissues. A dog tooth was accessed and the pulp tissue partially removed and infected. The root canal was then disinfected and the space filled with blood clot. (A) Histologic view of the healed pulp tissue 3 months after the disinfection. The pulp tissue on the left side healed and right side of the pulp tissue was lost and the space filled in by periodontal tissue including soft connective soft tissue and intracanal cementum (IC). Dashed line separates the healed pulp tissue (left) and the ingrown periapical connective tissue (right). (B) Higher magnification view of the odontoblast layer (od) from the left boxed region in (A). (C) Magnified view of the right boxed region in (A). Pulp space is filled with soft connective tissue. The hard tissue IC extends from the dentinal wall toward the opposite of the canal forming a bridge. Scale bars: 500 μm (A); 50 μm (B); 200 μm (C). (66).

Fig. 5

Partial pulp regeneration in dogs after autologous transplantation of CD31-/CD146- side population cells. (A) Regenerated pulp tissue in the cavity on the amputated sites (arrows). Note the tubular dentin and/or osteodentin only along the dentinal wall. (B) Tubular dentin formation along the dentinal wall in the cavity. Image originated from boxed area in (A). (C) Osteodentin formation at the top of the cavity under cement. (Adapted after permission (81)).

Fig. 6

De novo regeneration of human dental pulp/dentin. Illustration at upper left depicts SCID mouse subcutaneous study model for pulp/dentin regeneration. The canal space of human tooth root fragments (~6–7 mm long) was enlarged to ~2.5 mm in diameter. One end of the canal opening was sealed with MTA cement. (A–I) Histological analysis of in vivo pulp/dentin regeneration using SCAP. A root fragment was prepared and the canal space inserted with SCAP/PLG and transplanted into a SCID mouse for 3 months. The sample was harvested and processed for H&E staining. D, original dentin; rD, regenerated dentin-like tissue; rP, regenerated pulp-like tissue. Blue arrow in (A) indicates the blood supply entrance; green arrows in (B&C) indicate continuous layer of uniformed thickness of rD; yellow arrows in (E&F) indicate the region of well-aligned odontoblast-like cells with polarized cell bodies; green arrows in (G&H) indicate junctions between D and rD. Scale bars: (A) 1 mm; (B& C) 500 μm. (D) 100 μm; (E & F) 20 μm; (G–I) 50 μm. (Adapted from (85) with permission)

Fig. 7

Histological analysis of in vivo pulp/dentin regeneration using DPSCs. Samples were prepared using the same procedures as described in Fig. 6, except that the sample was harvested from the SCID mouse 4 months post-implantation. D, original dentin; rD, regenerated dentin-like tissue; rP, regenerated pulp-like tissue. Green arrows in (A) indicate rD; blue arrows in (A) indicate the entrance of blood supply; blue arrow in (B&C) indicate the thin layer of rD under MTA cement; blue arrows in (F&G) indicate the junction of D and rD; black arrow in (G) indicates well-aligned odontoblast-like cells. Scale bars: (A) 1 mm; (B) 200 μm; (C–E) 100 μm; (F&G) 50 μm. (Adapted from (85) with permission)

Fig. 8

Induced pluripotent stem (iPS) cells derived from human dental stem cells. (A, E, I) ES cell-like colonies. SHED, SCA, and DPSCs were reprogrammed into ES-cell like colonies with the 4 factors (Lin28, Nanog, Oct4 and Sox2). These iPS cells form teratomas in SCID mice containing tissues of all three germ layers. (B, F, J) Mainly primitive neural tissues, neural rosettes and retinal epithelium (ectoderm); (C, G, K) mainly cartilage (mesoderm); D, H, L) mainly glandular tissue or respiratory epithelium (endoderm). Scale bars: (A, E, I) 500 μm; (B, F–H, L), 200 μm; (C, J, K), 50 μm. *, in (K) indicates the space of a cavity inside the teratoma. (Adapted from (97) with permission)