In recent years, there has been increasing interest in the treatment of bone defects using undifferentiated mesenchymal stem cells (MSCs) in vivo. Recently, dental pulp has been proposed as a promising source of pluripotent mesenchymal stem cells (MSCs), which can be used in various clinical applications. Dentin is the hard tissue that makes up teeth, and has the same composition and strength as bone. However, unlike bone, dentin is usually not remodeled under physiological conditions. Here, we generated odontoblast-like cells from mouse dental pulp stem cells and combined them with honeycomb tricalcium phosphate (TCP) with a 300 μm hole to create bone-like tissue under the skin of mice. The bone-like hard tissue produced in this study was different from bone tissue, i.e., was not resorbed by osteoclasts and was less easily absorbed than the bone tissue. It has been suggested that hard tissue-forming cells induced from dental pulp do not have the ability to induce osteoclast differentiation. Therefore, the newly created bone-like hard tissue has high potential for absorption-resistant hard tissue repair and regeneration procedures.

The ideal scaffold for cartilage regeneration is expected to provide adequate mechanical strength, controlled degradability, adhesion, and integration with the surrounding native tissue. As it does this, it mimics natural ECMs functions, which allow for nutrient diffusion and promote cell survival and differentiation. Injectable hydrogels based on tyramine (TA)-functionalized hyaluronic acid (HA) and dextran (Dex) are a promising approach for cartilage regeneration. The properties of the hydrogels used in this study were adjusted by varying polymer concentrations and ratios. To investigate the changes in properties and their effects on cellular behavior and cartilage matrix formation, different ratios of HA- and dextran-based hybrid hydrogels at both 5 and 10% w/v were prepared using a designed mold to control generation. The results indicated that the incorporation of chondrocytes in the hydrogels decreased their mechanical properties. However, rheological and compression analysis indicated that 5% w/v hydrogels laden with cells exhibit a significant increase in mechanical properties after 21 days when the constructs are cultured in a chondrogenic differentiation medium. Moreover, compared to the 10% w/v hydrogels, the 5% w/v hybrid hydrogels increased the deposition of the cartilage matrix, especially in constructs with a higher Dex-TA content. These results indicated that 5% w/v hybrid hydrogels with 25% HA-TA and 75% Dex-TA have a high potential as injectable scaffolds for cartilage tissue regeneration.

Recently, dental pulp has been attracting attention as a promising source of multipotent mesenchymal stem cells (MSCs) for various clinical applications of regeneration fields. To date, we have succeeded in establishing rat dental pulp-derived cells showing the characteristics of odontoblasts under in vitro conditions. We named them Tooth matrix-forming, GFP rat-derived Cells (TGC). However, though TGC form massive dentin-like hard tissues under in vivo conditions, this does not lead to the induction of polar odontoblasts. Focusing on the importance of the geometrical structure of an artificial biomaterial to induce cell differentiation and hard tissue formation, we previously have succeeded in developing a new biomaterial, honeycomb tricalcium phosphate (TCP) scaffold with through-holes of various diameters. In this study, to induce polar odontoblasts, TGC were induced to form odontoblasts using honeycomb TCP that had various hole diameters (75, 300, and 500 μm) as a scaffold. The results showed that honeycomb TCP with 300-μm hole diameters (300TCP) differentiated TGC into polar odontoblasts that were DSP positive. Therefore, our study indicates that 300TCP is an appropriate artificial biomaterial for dentin regeneration.

β-Cyclodextrin (β-CyD) is cyclic oligosaccharide of a glucopyranose, containing a relatively hydrophobic central cavity and hydrophilic outer surface. However, the usefulness of β-CyD is limited owing to its low aqueous solubility whereas we found that its apparent high solubility was evident in some injectable solvents including 2-pyrrolidone (PYR), N-methyl pyrrolidone (NMP) and dimethyl sulfoxide (DMSO). Therefore, in the present study, the physicochemical properties of the 30-60% w/w β-CyD in PYR, NMP and DMSO were investigated such as viscosity, water resistant, matrix formation rate and syringeability. The higher the concentration of β-CyD resulted in the increased viscosity and the higher force and energy of syringeability. β-CyD in PYR gave the highest viscosity which contributed to the lowest syringeability while β-CyD in DMSO exhibited the highest syringeability. The β-CyD in DMSO and NMP exhibited the higher rate of matrix formation. β-CyD in PYR showed the highest water resistant for phase separation while β-CyD in NMP gave the faster de-mixing rate compared to that from PYR. The difference in physicochemical properties of β-CyD dried ppts studied by scanning electron microscope (SEM), differential scanning calorimetry (DSC), X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FT-IR) and thermogravimetric analysis (TGA) revealed that there was partial complexation of β-CyD with respective solvents. Both solution and precipitate characteristic properties will be useful for using β-CyD in further investigation as matrix material dissolved in the injectable vehicles as the in situ forming gel for periodontitis treatment.

OBJECTIVE: The objective of the study was to examine the effect of irisin on human periodontal ligament cells (hPDLCs) growth, migration and osteogenic behaviour in vitro.
METHODS: Primary hPDLCs and human osteoblasts (hOBs), used as positive controls, were cultured with irisin (10 and 100 ng/ml), and effect on cell proliferation was evaluated with 5-bromo-2`-deoxyuridine incorporation at 1, 2, and 3 days, and on migration capacity was investigated by scratch assay at 2, 6, and 24 h. Osteogenic behaviour was assessed with alkaline phosphatase activity, immunoassay at 3, 7, 14, and 21 days, and confocal laser scanning microscopy at 21 days. Mineralization was examined by Alizarin red staining at 21 days. Data were compared group wise using ANOVA tests.
RESULTS: Irisin induced increased proliferation of primary hPDLCs and hOBs at all time points compared to untreated controls. This was confirmed by scratch assay where irisin enhanced migration of both hPDLCs and hOBs after 6 and 24 h compared to controls. Irisin treatment promoted osteogenic behaviour of both cell types by enhancement of extracellular matrix formation. In hPDLCs irisin increased expression of type I collagen, secretion of osteoblastogenesis related proteins osteocalcin and leptin, and calcium deposition/mineralization compared to controls at 21 days. In addition, to enhance calcium deposition/mineralization in hOBs, irisin increased expression of periostin, and secretion of osteoblastogenesis related proteins osteopontin, alkaline phosphatase and osteocalcin, as compared to controls at 21 days.
CONCLUSIONS: Primary hPDLCs responded to irisin treatment with enhanced cell growth, migration, and matrix formation in vitro.

BACKGROUND: Cartilaginous endplate (CEP), a thin layer of hyaline cartilage located between the vertebral endplate and nucleus pulposus, transports the nutrient into the disc. The objective of this study was to evaluate the influence of T140 (polyphemusin II-derived peptide) on the CEP cell growth, apoptosis, and the matrix formation via the stromal cell-derived factor-1 (SDF-1)/cysteine X cysteine (CXC) receptor-4 (CXCR4) signaling pathway.
METHODS: Sprague-Dawley rats were euthanized by cervical dislocation and dissected for the isolation and the appraisal of CEP cells that were extracted from the endplate in rat intervertebral discs and were then added with different concentrations of reagents (SDF-1 and T140). The effect of T140 on CEP cell proliferation and apoptosis were analyzed. The messenger RNA (mRNA) and protein expressions of CXCR4, prominin-1, proteoglycans, type II collagen, B-cell lymphoma-2 (Bcl-2), and Bcl-2 associated X protein were analyzed by reverse transcription quantitative polymerase chain reaction and Western blot analysis.
RESULTS: T140 promoted the proliferation of CEP cells and inhibited the apoptosis of CEP cells. Additionally, T140 suppressed the mRNA and protein expression of CXCR4, prominin-1, and Bcl-2 associated X protein, and increased the mRNA and protein expression of proteoglycans, type II collagen, and Bcl-2.
CONCLUSIONS: T140 promotes the proliferation and matrix formation and inhibits the apoptosis of CEP cells by blocking the SDF-1/CXCR4 signaling pathway in vitro, which provides a certain therapeutic effect on the degeneration of intervertebral discs.

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 β-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.