Pulp Regeneration by 3-dimensional Dental Pulp Stem Cell Constructs

Abstract

Dental pulp regeneration therapy for the pulpless tooth has attracted recent attention, and clinical trial studies are underway with the tissue engineering approach. However, there remain many concerns, including the extended period for regenerating the dental pulp. In addition, the use of scaffolds increases the risk of inflammation and infection. To establish a basic technology for novel dental pulp regenerative therapy that allows transplant of pulp-like tissue, we attempted to fabricate scaffold-free 3-dimensional (3D) cell constructs composed of dental pulp stem cells (DPSCs). Furthermore, we assessed viability of these 3D DPSC constructs for dental pulp regeneration through in vitro and in vivo studies. For the in vitro study, we obtained 3D DPSC constructs by shaping sheet-like aggregates of DPSCs with a thermoresponsive hydrogel. DPSCs within constructs remained viable even after prolonged culture; furthermore, 3D DPSC constructs possessed a self-organization ability necessary to serve as a transplant tissue. For the in vivo study, we filled the human tooth root canal with DPSC constructs and implanted it subcutaneously into immunodeficient mice. We found that pulp-like tissues with rich blood vessels were formed within the human root canal 6 wk after implantation. Histologic analyses revealed that transplanted DPSCs differentiated into odontoblast-like mineralizing cells at sites in contact with dentin; furthermore, human CD31-positive endothelial cells were found at the center of regenerated tissue. Thus, the self-organizing ability of 3D DPSC constructs was active within the pulpless root canal in vivo. In addition, blood vessel-rich pulp-like tissues can be formed with DPSCs without requiring scaffolds or growth factors. The technology established in this study allows us to prepare DPSC constructs with variable sizes and shapes; therefore, transplantation of DPSC constructs shows promise for regeneration of pulpal tissue in the pulpless tooth.

Revascularization and vital pulp therapy in immature molars with necrotic pulp and irreversible pulpitis: A case report with two‐year follow‐up

“…Herein, a biodegradable scaffold serves as a temporary matrix for regenerating cells to attach, proliferate, and differentiate to mature and functional cells which produce pulp and dentin. 18,19 It is now well-established that mesenchymal stem cells which migrate from different sources such as apical papilla, periodontal ligament, and bone marrow play a significant role in pulp and dentin regeneration. 20 In the present case, migrating stem cells are suggested to be responsible for pulp regeneration, dentin production, and root development.…”

“…In the revascularization procedure, the clinician uses the approach of tissue engineering for the regeneration of pulp tissue and maturation of root apex. Herein, a biodegradable scaffold serves as a temporary matrix for regenerating cells to attach, proliferate, and differentiate to mature and functional cells which produce pulp and dentin . It is now well‐established that mesenchymal stem cells which migrate from different sources such as apical papilla, periodontal ligament, and bone marrow play a significant role in pulp and dentin regeneration .…”

Fabricating large‐scale three‐dimensional constructs with living cells by processing with syringe needles

“…There are several methods of fabricating 3D cell constructs, such as applying a low attachment cell culture plate, thermos‐responsive polymers, and the layer‐by‐layer technique . These methods represent easy ways of obtaining cell constructs; however, it is difficult to control the morphology of constructs obtained via these methods.…”

“…13 It has also been reported that cell interactions and drug sensitivities could be more accurately evaluated for anticancer drug development using cell constructs. 14 There are several methods of fabricating 3D cell constructs, such as applying a low attachment cell culture plate, 15 thermosresponsive polymers, 16 and the layer-by-layer technique. 17 These methods represent easy ways of obtaining cell constructs; however, it is difficult to control the morphology of constructs obtained via these methods.…”

Contraction dynamics of dental pulp cell rod microtissues

“…Scaffold-free schemes using microtissues of various cell types in complex shapes such as spheroids, rods, toroids [5][6][7], and sheets not only facilitate a more in vivo-like environment for the transplanted cells but also avoid issues sprouting from inflammation and slow degradation of the scaffolds [8]. In order to mimic the rod-like shape of the root canal, attempts have been made to fabricate the most suitable cell-seeding system including sheets and spheroids [4,9]. The rationale behind our research was to check the feasibility of molding dental pulp cells into rodshaped microtissues and to investigate the potential signaling pathways responsible for regulating their contraction.…”