Biomechanical characterization of a fibrinogen-blood hydrogel for human dental pulp regeneration.

In dental practice, Regenerative Endodontic Treatment (RET) is applied as an alternative to classical endodontic treatments of immature necrotic teeth. This procedure, also known as dental pulp revitalization, relies on the formation of a blood clot inside the root canal leading to the formation of a reparative vascularized tissue similar to dental pulp, which would provide vitality to the affected tooth. Despite the benefit of this technique, it lacks reproducibility due to the fast degradation and poor mechanical properties of blood clots. This work presents a method for constructing a fibrinogen-blood hydrogel that mimics the viscoelastic properties of human dental pulp while preserving the biological properties of blood for application in RET. By varying the blood and fibrinogen concentrations, gels with different biomechanical and biological properties were obtained. Rheology and atomic force microscopy (AFM) were combined to study the viscoelastic properties. AFM was used to evaluate the elasticity of human dental pulp. The degradation and swelling rates were assessed by measuring weight changes. The biomimetic properties of the gels were demonstrated by studying the cell survival and proliferation of dental pulp cells (DPCs) for 14 days. The formation of an extracellular matrix (ECM) was assessed by multiphoton microscopy (MPM). The angiogenic potential was evaluated by an ex vivo aortic ring assay, in which the endothelial cells were observed by histological staining after migration. The results show that the Fbg-blood gel prepared with 9 mg ml-1 fibrinogen and 50% blood of the Fbg solution volume has similar elasticity to human dental pulp and adequate degradation and swelling rates. It also allows cell survival and ECM secretion and enhances endothelial cell migration and formation of neovessel-like structures.

In vitro, ex vivo, and in vivo models for dental pulp regeneration.

Based on the concept of tissue engineering (Cells-Scaffold-Bioactive molecules), regenerative endodontics appeared as a new notion for dental endodontic treatment. Its approaches aim to preserve dental pulp vitality (pulp capping) or to regenerate a vascularized pulp-like tissue inside necrotic root canals by cell homing. To improve the methods of tissue engineering for pulp regeneration, numerous studies using in vitro, ex vivo, and in vivo models have been performed. This review explores the evolution of laboratory models used in such studies and classifies them according to different criteria. It starts from the initial two-dimensional in vitro models that allowed characterization of stem cell behavior, through 3D culture matrices combined with dental tissue and finally arrives at the more challenging ex vivo and in vivo models. The travel which follows the elaboration of such models reveals the difficulty in establishing reproducible laboratory models for dental pulp regeneration. The development of well-established protocols and new laboratory ex vivo and in vivo models in the field of pulp regeneration would lead to consistent results, reduction of animal experimentation, and facilitation of the translation to clinical practice.

Porous Tantalum VS. Titanium Implants: Enhanced Mineralized Matrix Formation after Stem Cells Proliferation and Differentiation.

Titanium dental implants are used routinely, with surgical procedure, to replace missing teeth. Even though they lead to satisfactory results, novel developments with implant materials can still improve implant treatment outcomes. The aim of this study was to investigate the efficiency of porous tantalum (Ta) dental implants for osseointegration, in comparison to classical titanium (Ti). Mesenchymal stem cells from the dental pulp (DPSC) were incubated on Ta, smooth titanium (STi), and rough titanium (RTi) to assess their adhesion, proliferation, osteodifferentiation, and mineralized matrix production. Cell proliferation was measured at 4 h, 24 h, 48 h with MTT test. Early osteogenic differentiation was followed after 4, 8, 12 days by alkaline phosphatase (ALP) quantification. Cells organization and matrix microstructure were studied with scanning electron microscopy (SEM) and energy dispersive X-ray spectroscopy (EDX). Collagen production and matrix mineralization were evaluated by immunostaining and histological staining. MTT test showed significantly higher proliferation of DPSC on Ta at 24 h and 48 h. However, APL quantification after 8 and 12 days was significantly lower for Ta, revealing a delayed differentiation, where cells were proliferating the more. After 3 weeks, collagen immunostaining showed an efficient production of collagen on all samples. However, Red Alizarin staining clearly revealed a higher calcification on Ta. The overall results tend to demonstrate that DPSC differentiation is delayed on Ta surface, due to a longer proliferation period until cells cover the 3D porous Ta structure. However, after 3 weeks, a more abundant mineralized matrix is produced on and inside Ta implants. Cell populations on porous Ta proliferate greater and faster, leading to the production of more calcium phosphate deposits than cells on roughened and smooth titanium surfaces, revealing a potential enhanced capacity for osseointegration.

Pulp Regeneration Concepts for Nonvital Teeth: From Tissue Engineering to Clinical Approaches.

Following the basis of tissue engineering (Cells-Scaffold-Bioactive molecules), regenerative endodontic has emerged as a new concept of dental treatment. Clinical procedures have been proposed by endodontic practitioners willing to promote regenerative therapy. Preserving pulp vitality was a first approach. Later procedures aimed to regenerate a vascularized pulp in necrotic root canals. However, there is still no protocol allowing an effective regeneration of necrotic pulp tissue either in immature or mature teeth. This review explores in vitro and preclinical concepts developed during the last decade, especially the potential use of stem cells, bioactive molecules, and scaffolds, and makes a comparison with the goals achieved so far in clinical practice. Regeneration of pulp-like tissue has been shown in various experimental conditions. However, the appropriate techniques are currently in a developmental stage. The ideal combination of scaffolds and growth factors to obtain a complete regeneration of the pulp-dentin complex is still unknown. The use of stem cells, especially from pulp origin, sounds promising for pulp regeneration therapy, but it has not been applied so far for clinical endodontics, in case of necrotic teeth. The gap observed between the hope raised from in vitro experiments and the reality of endodontic treatments suggests that clinical success may be achieved without external stem cell application. Therefore, procedures using the concept of cell homing, through evoked bleeding that permit to recreate a living tissue that mimics the original pulp has been proposed. Perspectives for pulp tissue engineering in the near future include a better control of clinical parameters and pragmatic approach of the experimental results (autologous stem cells from cell homing, controlled release of growth factors). In the coming years, this therapeutic strategy will probably become a clinical reality, even for mature necrotic teeth.