Characterization of Living Dental Pulp Cells in Direct Contact with Mineral Trioxide Aggregate
Abstract
1. Introduction
2. Materials and Methods
2.1. Gene Transfection of Discosoma Species Red Fluorescent Protein (DsRed) into a Porcine Dental-Pulp Cell Line (PPU7)
2.2. Alkaline Phosphatase (ALP) Activity Assay of the PPU7 Cell Line
2.3. Changes in the Number of DsRed-PPU7 Cells on the Mineral Trioxide Aggregate (MTA) Disk
2.4. Correlation of Fluorescence Intensity and Cell-Proliferation Rate of DsRed-PPU7 Cells
2.5. Characterization of DsRed-PPU7 Cells on MTA Disks
2.6. Quantitative Polymerase Chain Reaction (qPCR) Analysis
2.7. Scanning Electron Microscopy (SEM) and Electron-Probe Microanalysis (EPMA)
2.8. Statistical Analysis
3. Results
3.1. Comparison of PPU7 and DsRed-PPU7 Cells
3.2. Changes in the Number of DsRed-PPU7 Cells on the MTA Disk
3.3. Correlation between Fluorescence Intensity and Cell-Proliferation Rate of DsRed-PPU7 Cells
3.4. DsRed-PPU7 Cells on the MTA Disk
3.5. Effect of BMP and TGF-β on DsRed-PPU7 Cells on the MTA Disk
3.6. Direction of Differentiation of DsRed-PPU7 Cells on the MTA Disk
3.7. Observation of Mineralized Precipitates on the MTA Disk
4. Discussion
5. Conclusions
Author Contributions
Funding
Acknowledgments
Conflicts of Interest
Abbreviations
MTS: | 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphentl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt). |
LDN | 4-[6-[4-(1-piperazinyl)phenyl]pyrazolo[1,5-a]pyrimidin-3-yl]-quinolinehydrochloride. |
RNA | ribonucleic acid |
mRNA | messenger ribonucleic acid |
PBS | phosphate buffer saline |
NaOH | sodium hydroxide |
MgCl2 | magnesium chloride. |
HCl | hydrochloric acid |
CO2 | carbon dioxide |
Ca | calcium |
P | phosphorus |
Si | silicon |
Smad | homologs of Caenorhabditis elegans (Sma) and Drosophila (Mad) proteins |
Appendix A
Appendix A.1. Fluorescence Microscope Observation of DeRed-PPU7 Cells on MTA Disk
Appendix A.2. Experimental Animal Model
Appendix A.3. Preparation of DsRed-PPU7 Cells
Appendix A.4. Splice Variants of Dentin Sialophosphoprotein
Gene | Sequence (5′→3′) | Size (bp) | qPCR Protocol (45 Cycles) | ||
---|---|---|---|---|---|
Dspp-v1 | F | CCCAGAAACCCAATCAGAGA | 300 | Denaturation Annealing Extension | 95 °C, 10 s 60 °C, 10 s 72 °C, 15 s |
R | TATGTGTTTTGCTGGGTCCA | ||||
Dspp-v2 | F | CCCAGAAACCCAATCAGAGA | 149 | ||
R | GGGAAGGAAGGGGAGAATTT | ||||
Mmp20 | F | ATGCAGCTTACGAAGTGGCT | 95 | ||
R | GGGAGGACCTTGCATTTGGA | ||||
Col1a1 | F | TGCGACGAAATCAAGAACTG | 204 | ||
R | TCCAGGAAGTCCAGGTTGTC | ||||
Gapdh | F | CCATCACCATCTTCCAGGAG | 346 | ||
R | ACAGTCTTCTGGGTGGCAGT |
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Hattori-Sanuki, T.; Karakida, T.; Chiba-Ohkuma, R.; Miake, Y.; Yamamoto, R.; Yamakoshi, Y.; Hosoya, N. Characterization of Living Dental Pulp Cells in Direct Contact with Mineral Trioxide Aggregate. Cells 2020, 9, 2336. https://doi.org/10.3390/cells9102336
Hattori-Sanuki T, Karakida T, Chiba-Ohkuma R, Miake Y, Yamamoto R, Yamakoshi Y, Hosoya N. Characterization of Living Dental Pulp Cells in Direct Contact with Mineral Trioxide Aggregate. Cells. 2020; 9(10):2336. https://doi.org/10.3390/cells9102336
Chicago/Turabian StyleHattori-Sanuki, Tamaki, Takeo Karakida, Risako Chiba-Ohkuma, Yasuo Miake, Ryuji Yamamoto, Yasuo Yamakoshi, and Noriyasu Hosoya. 2020. "Characterization of Living Dental Pulp Cells in Direct Contact with Mineral Trioxide Aggregate" Cells 9, no. 10: 2336. https://doi.org/10.3390/cells9102336
APA StyleHattori-Sanuki, T., Karakida, T., Chiba-Ohkuma, R., Miake, Y., Yamamoto, R., Yamakoshi, Y., & Hosoya, N. (2020). Characterization of Living Dental Pulp Cells in Direct Contact with Mineral Trioxide Aggregate. Cells, 9(10), 2336. https://doi.org/10.3390/cells9102336