Establishment of odontoblastic cells, which indicate odontoblast features both in vivo and in vitro.

Tooth tissue engineering offers very attractive perspectives for elaboration of regenerative treatments, which enables to cure tooth loss and restore quality of life of the patients. To elaborate such treatment, isolation and culture of dental pulp cell must be achieved as a key element. In this article, we report the establishment of a stable cell line from GFP transgenic rat dental pulp, named TGC (Tooth Matrix-forming, GFP Rat-derived Cell). TGCs have exhibited odontoblastic feature both in vitro and in vivo. In vitro, TGC exposed to osteogenic medium demonstrated collagen fiber synthesis with matrix vesicle and mineralization and formed a sheet-like substrate on the cell culture dish. Increased ALP activity and elevated transcription level of various genes involved in calcification and dentin formation were also observed. In vivo, transplanted TGC in SCID mice with ß-TCP particles formed dentin-like and pulp-like structure with lining odontoblast. Notably, even after up to 80 passages, TGCs retain their morphological features and differentiation ability. To our knowledge, this is the first report of a dental pulp-derived cell with such stable odontoblastic characteristics. TGC could be a very useful model for further study on dental pulp cell.

Immunohistochemical study of hard tissue formation in the rat pulp cavity after tooth replantation.

While mineralized tissue is formed in the pulp cavity after tooth replantation or transplantation, little is known of this hard tissue formation. Therefore, we conducted histological and immunohistochemical evaluations of hard tissue formed in the pulp of rat maxillary molars after tooth replantation. At 5 days after replantation, degenerated odontoblasts were lining the pulp cavity. At 14 days, dentin- or bone-like tissue was present in the pulp cavity. Immunoreactivity for osteopontin (OPN) and bone sialoprotein (BSP) was strong in the bone-like tissue, but weak in the dentin-like tissue. Conversely, dentin sialoprotein (DSP) was localized in the dentin-like tissue, but not in the bone-like tissue. Cells positive for BMP4, Smad4, Runx2, and Osterix were found around the blood vessels of the root apex at 5 days. At 14 days, these cells were also localized around the bone-like tissue. Cells expressing alpha-smooth muscle actin (SMA) were seen around the newly formed bone-like tissue, whereas no such cells were found around the newly formed dentin-like tissue. In an experiment involving the transplantation of a green fluorescent protein (GFP)-transgenic rat tooth into a wild-type rat tooth socket, GFP-positive cells were detected on the surface of the bone-like tissue and over all dentin-like tissue. These results indicate that the original pulp cells had the ability to differentiate into osteoblast-like cells as well as into odontoblast-like cells.

The Effects of Platelet-Derived Growth Factor-BB on Human Dental Pulp Stem Cells Mediated Dentin-Pulp Complex Regeneration.

Dentin-pulp complex regeneration is a promising alternative treatment for the irreversible pulpitis caused by tooth trauma or dental caries. This process mainly relies on the recruitment of endogenous or the transplanted dental pulp stem cells (DPSCs) to guide dentin-pulp tissue formation. Platelet-derived growth factor (PDGF), a well-known potent mitogenic, angiogenic, and chemoattractive agent, has been widely used in tissue regeneration. However, the mechanisms underlying the therapeutic effects of PDGF on dentin-pulp complex regeneration are still unclear. In this study, we tested the effect of PDGF-BB on dentin-pulp tissue regeneration by establishing PDGF-BB gene-modified human dental pulp stem cells (hDPSCs) using a lentivirus. Our results showed that PDGF-BB can significantly enhance hDPSC proliferation and odontoblastic differentiation. Furthermore, PDGF-BB and vascular endothelial growth factor (VEGF) secreted by hDPSCs enhanced angiogenesis. The chemoattractive effect of PDGF-BB on hDPSCs was also confirmed using a Transwell chemotactic migration model. We further determined that PDGF-BB facilitates hDPSCs migration via the activation of the phosphatidylinositol 3 kinase (PI3K)/Akt signaling pathway. In vivo, CM-DiI-labeled hDPSCs were injected subcutaneously into mice, and our results showed that more labeled cells were recruited to the sites implanted with calcium phosphate cement scaffolds containing PDGF-BB gene-modified hDPSCs. Finally, the tissue-engineered complexes were implanted subcutaneously in mice for 12 weeks, the Lenti-PDGF group generated more dentin-like mineralized tissue which showed positive staining for the DSPP protein, similar to tooth dentin tissue, and was surrounded by highly vascularized dental pulp-like connective tissue. Taken together, our data demonstrated that the PDGF-BB possesses a powerful function in prompting stem cell-based dentin-pulp tissue regeneration. Stem Cells Translational Medicine 2017;6:2126-2134.

Xenografts of recombined bovine odontogenic tissues and cultured cells to hypothymic mice.

It was demonstrated in this study that recombined tissues of bovine dental papilla and reduced enamel epithelium, when transplanted to the subcapsular kidney site of hypothymic mice, elaborated tissues resembling osteodentin and dentin matrices with which latter tissues were closely associated cells resembling odontoblasts. In xenografts of cultured cell populations of these tissues, osteodentin-like matrices occurred in the absence of dentin-like matrices.

Odontoblast-like cell differentiation and dentin formation induced with TGF-ß1.

OBJECTIVE: To investigate the inductive potential of scaffold material combing with transforming growth factor-ß1 (TGF-ß1), and to induce odontoblast differentiation and dentin formation from dental pulp cells both in vitro and in vivo. METHODS: Primarily cultured dental pulp cells were used for MTT, ALP activity assay and Alizarin red staining in the presence of TGF-ß1. Pelleted cells were put on the filters combining with or not with TGF-ß1 and cultured in vitro or in vivo. The in vitro and in vivo cell response and tissue formation were analysed with Haematoxylin-Eosin (HE), transmission electron microscopy (TEM) and immunohistochemical staining. RESULTS: TGF-ß1 increased the mineralization and ALP activity of dental pulp cells as revealed by Alizarin red staining and ALP activity assay. After in vitro culture for 7 days, cells polarized in the TGF-ß1 group and expressed dentin sialoprotein (DSP), osteopontin (OPN) and type I collagen (Col I). After in vivo transplantation for 7 days, columnar odontoblast formed on the surface of filter in experimental group, and tubular dentin expressing DSP formed after 3 months transplantation. CONCLUSION: It was concluded that TGF-ß1 combining with transfilter could induce odontoblast differentiation and dentin formation. Our results implied that suitable substrate for the progenitors of odontoblast to anchor on and inductive signals to initiate the differentiation of odontoblast should be taken into consideration when designing scaffold material for inducing dentin tissue engineering.

Bionic restorative system: its potential value in caries therapy.

Dental defect caused by dental caries is usually restored by fillings, inlays or onlays at the present day. Although the therapeutic effects of these methods have already been confirmed, complications occasionally set in, such as pulp injury, fracture and secondary caries. Bionic dental organic center possesses similar functions of the natural dental organic center. So we put forward a hypothesis that bionic organic center can be transplanted onto the conditioned pulpal walls of the prepared cavity and a specific filling material, which the cavity will be filled with, provides oxygen, nutrition and raw materials for it to regenerate the lost odontal tissue in vivo. The regenerated odontal tissue which has similar properties of the healthy odontal tissue will restore the defect and it will be combined with the residual odontal tissue tightly, not only in physical structure but also in function. Then the teeth suffering from dental caries could live and function like healthy ones.

Side population cells isolated from porcine dental pulp tissue with self-renewal and multipotency for dentinogenesis, chondrogenesis, adipogenesis, and neurogenesis.

Dental pulp has the potential to form dentin as a regenerative response to caries. This regeneration is mediated by stem/progenitor cells. Thus, stem cell therapy might be of potential utility in induction of reparative dentin. We isolated side population (SP) cells from dental pulp based on the exclusion of the DNA binding dye Hoechst 33342 by flow cytometry and compared its self-renewal capacities and multipotency with non-SP cells and primary pulp cells. The cumulative cell number of the SP cells was greater than the non-SP cells and primary pulp cells. Bmi1 was continuously expressed in SP cells, suggesting longer proliferative lifespan and self-renewal capacity of SP cells. Next, the maintenance of the multilineage differentiation potential of pulp SP cells was investigated. Expression of type II collagen and aggrecan confirmed chondrogenic conversion (30%) of SP cells. SP cells expressed peroxisome proliferator-activated receptor gamma and adaptor protein 2, showing adipogenic conversion. Expression of mRNA and proteins of neurofilament and neuromodulin confirmed neurogenic conversion (90%). These results demonstrate that pulp SP cells maintain multilineage differentiation potential. We further examined whether bone morphogenetic protein 2 (BMP2) could induce differentiation of pulp SP cells into odontoblasts. BMP2 stimulated the expression of dentin sialophosphoprotein (Dspp) and enamelysin in three-dimensional pellet cultures. Autogenous transplantation of the Bmp2-supplemented SP cells on the amputated pulp stimulated the reparative dentin formation. Thus, adult pulp contains SP cells, which are enriched for stem cell properties and useful for cell therapy with BMP2 for dentin regeneration.

La aplicación de la ingeniería tisular a la regeneración de la pulpa y la dentina en endodoncia / The application of tissue engineering to regeneration of pulp and dentin in endodontics

La caries, la pulpitis y la periodontitis incrementan los costes de la atención sanitaria y por pérdida de productividad económica de los afectados. El resultado final es la pérdida prematura de dientes y por lo tanto la disminución de la calidad de vida. Los avances en el tratamiento de la pulpa vital con células madre / progenitoras puede dar impulso a la regeneración del complejo pulpo-dentinario sin la eliminación de toda la pulpa. La ingeniería tisular es la ciencia del diseño y la fabricación de tejidos nuevos para reemplazar partes perdidas debido a enfermedades, entre ellas el cáncer y los traumatismos. Los tres ingredientes clave de la ingeniería tisular son las señales para la morfogénesis, las células madre para responder a los morfógenos y el soporte o matriz extracelular. En estudios preclínicos se han desarrollado tratamientos celulares y genéticos para muchos tejidos y órganos, como el hueso, el corazón, el hígado y el riñón, como método de liberar factores de crecimiento, citocinas o morfógenos con células madre / progenitoras en un soporte en las zonas de lesión tisular para acelerar y/o inducir una regeneración biológica natural. El tejido pulpar contiene células madre / progenitoras que tienen el potencial de diferenciarse en odontoblastos en respuesta a proteinas morfogenéticas óseas (BMPs). Hay dos estrategias para regenerar la dentina. La primera es el tratamiento in vivo, en que las proteínas BPM o los genes BMP se aplican directamente a la pulpa expuesta o amputada. La segunda es el tratamiento ex vivo y consiste en el aislamiento de células madre / progenitoras del tejido pulpar, la diferenciación en odontoblastos con genes BMP recombinantes o BMP y finalmente el trasplante autólogo para regenerar la dentina. Esta revisión está enfocada al progreso reciente en este área y discuta las barreras y los retos para la utilidad clínica en endodoncia

Caries, pulpitis, and apical periodontitis in crease health care costs and attendant loss of economic productivity. They ultimately result in premature tooth loss and therefore diminishing the quality of life. Advances in vital pulp therapy ith pulp stem/progenitor cells might give impetus to regenerate dentin-pulp complex without the removal of the whole pulpo Tissue engineering is the science of design and manufacture of new tissues to replace lost parts because of diseases including cancer and trauma. The three key ingredients for tissue engineering are signals for morphogenesis, stem cells for responding to morphogens and the scaffold of extracellular matrix. In preclinical studies cell/ therapy and gene therapy have been developed for many tissues and organs such as bone, heart, liver, and kidney as a menas of delivering growth factors, cytokines, or morphogens with stem/progenitor cells in a scaffold to the si/es of tissue injury to accelerate and/or induce a natural biological regeneration. The pulp tissue contains stem/progenitor cells that potential/y differentiate into odontoblasts in response to bone morphogenetic proteins (BMPs). There are two strategies to renerate dentin. First, is in vivo therapy, where BMP proteins or BMP genes are directly applied to the exposed or amputated pulpo Second is ex vivo therapy and consists of isolation os stem/progenitor cells from pulp tissue, differentiation into odontoblasts with recombinant BMPs or BMP genes and final/y transplanted autogenously to regenerate dentin. This review is focused on the recent progress in this area and discusses the barriers and challenges for clinical utility in endodontics