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Comparison of Biological Characteristics of Stem Cells from the Apical Papilla and Dental Pulp Stem Cells

Author: MaLi
Tutor: YangZuoShan；WangYan
School: Shandong University
Course: Clinical Stomatology
Keywords: Tumor necrosis factorα Stem cells from the apical papilla dental pulp stem cells Cell culture MTT assay Mineralization
CLC: R781
Type: Master's thesis
Year: 2010
Downloads: 177
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Abstract

Objective:Gronthos et al. isolated human dental pulp stem cells (DPSCs) with high proliferative capacity, self-renewal property, and multilineage differentiation potential. DPSCs are able to differentiate into odontoblast-like cells under appropriate conditions in vitro. More importantly, DPSCs can differentiate into odontoblast-like cells and form dentin/pulp-like complex when implanted into the subcutaneous space of immunocompromised mice. Recently, a new population of mesenchymal stem cells residing in the apical papilla of permanent immature teeth has been discovered and was termed stem cells from the apical papilla (SCAP).These stem cells appear to be the source of odontoblasts that are responsible for the formation of root dentin. The aim of this study was to culture human cells from the apical papilla (APC) in vitro, stimulation APC differentiation, and study the proliferation effects and the differentiation effects of TNF-αon these cells, and explore the possibility of their dentinogenesis under inflammation; Meanwhile to compare the dental pulp stem cells (DPCs) with them.Methods:The apical papilla, as well as the pulp tissues were obtained from healthy third permanent molars under the age 18,were digested in collagenase I and Dispase, and cultured and subcultured at confluence. Experiments were carried out with cells from third to fifth passages.1. Cells from third passage were plated into 6-well plate at an initial concentration of 1 X 104 cells/mL. STRO-1 was examined by immunofluorescent technique.2. The fourth cells were plated into 96-well plate at an initial concentration of 1 X 104 cells/mL. After incubated for 4 days, the cells were measured proliferation by MTT assay.3. The fourth cells were plated into 24-well plate at an initial concentration of 1 X 103 cells/mL. After incubated for 24 hours, mineralized medium(15%FBS, lOmmol/Lβ-glycerophosphate,10nmol/L dexamethasone,50mg/L vitamin C andα-MEM) were added into the test group, andα-MEM was added into the medium of the control. After incubated for 28 days, the mineralized modules formation was detected with Alizarin Red staining for calcium. DSP and BSP were examined by immunohistochemistry stains.4. Cells were plated into 6-well plate at an initial concentration of 1×104 cells/mL. After incubated for 24 hours, multiple concentrations of TNF-a (5,10,50ng/mL) were added into the group, andα-MEM with 0.5%FBS was added into the medium of the control. The cells were then incubated with drugs for 4 days to measure proliferation by MTT assay.5. The fourth cells were plated into 24-well plate at an initial concentration of 1 X 103 cells/mL. After incubated for 24 hours, 10ng/mL TNF-a with mineralized medium(15%FBS, 10mmol/Lβ-glycerophosphate,10nmol/L dexamethasone,50mg/L vitamin C andα-MEM) were added into the test group. After incubated for 28 days, the mineralized modules formation was detected with Alizarin Red staining for calcium. DSP、BSP were examined by immunohistochemistry stains. Results:1. APCs presented elongate shape, polygon or spindle, respectively. They exhibited fibroblastic characteristics. SCAP have stronger proliferation activity than DPSCs. STRO-1 was positive strong in APCs and DPCs.2. SCAP have stronger proliferation activity than DPSCs by MTT assay.3. SCAP and DPSCs formed white calcified modules by induced with mineralized conditional medium. The calcified nodules were stained by Alizarin Red. DSP and BSP were positive stronger in induced cells.4. At the concentration of lOng/mL and 50ng/mL, TNF-a enhanced significantly the proliferating activity of the two kinds of the cells(P<0.05).5. SCAP and DPSCs formed in white calcified modules by induced with mineralized conditional medium and lOng/mL TNF-a. The calcified nodules were stained by Alizarin Red. DSP and BSP were positive stronger in induced cells.Conclusion:APCs and DPCs isolated in vitro exhibited fibroblastic characteristics and could differentiate into odontoblast-like cells. STRO-1 was positive strong in APCs and DPCs. SCAP have stronger proliferation activity than DPSCs. The two kinds of cells have both stronger proliferation and differention activity when especially under inflammation. The discovery would possibly be beneficial to continuous development of the root of young permanent tooth with dental pulp necrosis and apical periodontitis after appropriate treatment.

Comparison of Biological Characteristics of Stem Cells from the Apical Papilla and Dental Pulp Stem Cells

Abstract

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