Loss of Stat3 in Osterix+ cells impairs dental hard tissues development.

BACKGROUND: Mutations in the signal transducers and activators of transcription 3 (STAT3) gene result in hyper-IgE syndrome(HIES), a rare immunodeficiency that causes abnormalities in immune system, bones and teeth. However, the role of Stat3 in development of dental hard tissues was yet to investigate. METHODS: In this study, a transgenic mouse of conditional knockout of Stat3 in dental mesenchymal cells (Osx-Cre; Stat3fl/fl, Stat3 CKO) was made. The differences of postnatal tooth development between control and Stat3 CKO mice were compared by histology, µCT and scanning electron microscopy. RESULT: Compared with the control, Stat3 CKO mice were presented with remarkable abnormal tooth phenotypes characterized by short root and thin dentin in molars and incisors. The enamel defects were also found on mandibular incisors. showed that Ki67-positive cells significantly decreased in dental mesenchymal of Stat3 CKO mice. In addition, ß-catenin signaling was reduced in Hertwig's epithelial root sheath (HERS) and odontoblasts of Stat3 CKO mice. CONCLUSIONS: Our results suggested that Stat3 played an important role in dental hard tissues development, and Stat3 may regulate dentin and tooth root development through the ß-catenin signaling pathway.

A Simplified and Systematic Method to Isolate, Culture, and Characterize Multiple Types of Human Dental Stem Cells from a Single Tooth.

This chapter describes a simplified method that allows the systematic isolation of multiple types of dental stem cells such as dental pulp stem cells (DPSC), periodontal ligament stem cells (PDLSC), and stem cells of the apical papilla (SCAP) from a single tooth. Of specific interest is the modified laboratory approach to harvest/retrieve the dental pulp tissue by minimizing trauma to DPSC by continuous irrigation, reduction of frictional heat from the bur rotation, and reduction of the bur contact time with the dentin. Also, the use of a chisel and a mallet will maximize the number of live DPSC for culture. Steps demonstrating the potential for multiple cell differentiation lineages of each type of dental stem cell into either osteocytes, adipocytes, or chondrocytes are described. Flow cytometry, with a detailed strategy for cell gating and analysis, is described to verify characteristic markers of human mesenchymal multipotent stromal cells (MSC) from DPSC, PDLSC, or SCAP for subsequent experiments in cell therapy and in tissue engineering. Overall, this method can be adapted to any laboratory with a general setup for cell culture experiments.

Three-Dimensional Printed Scaffolds with Multipotent Mesenchymal Stromal Cells for Rabbit Mandibular Reconstruction and Engineering.

Multipotent mesenchymal stromal cells (MSC) derived from both the bone marrow and adipose tissue possess the ability to differentiate into multiple cell lineages, regulate the immune function by secreting numerous bioactive paracrine factors, and hold great potential in cell therapy and tissue engineering. When combined with three-dimensional (3D) scaffolds, MSC can be used for bone defect reconstruction and engineering. This protocol describes the isolation of bone marrow mesenchymal stromal cells (BMMSC) and adipose-tissue derived stem cells (ADSC) from rabbits for subsequent seeding on tissue-engineered 3D-printed scaffolds and transplantation into a rabbit-model with the goal of repairing large osseous mandibular defects (one quarter of the lower jaw is removed surgically). Steps to demonstrate the three cell differentiation lineage potentials of BMMSC and ADSC into osteocytes, adipocytes, and chondrocytes are described. A modified cell seeding method using syringes on scaffold is detailed. Creating a large mandibular bone defect, the rapid prototyping method to print a customized 3D-scaffold, the scaffold implantation procedure in rabbits, and microcomputed tomography (micro-CT) analysis are also described.