Plasma Membrane Ca2+–ATPase in Rat and Human Odontoblasts Mediates Dentin Mineralization
Abstract
:1. Introduction
2. Materials and Methods
2.1. Ethical Approval
2.2. Odontoblast Cell Culture
2.3. Acute Isolation of Rat Odontoblasts from Dental Pulp Slices
2.4. RT-PCR
2.5. Fluorescence Measurement of [Ca2+]i
2.6. Mineralization Assay
2.7. Solutions and Reagents
2.8. Immunostaining
2.9. Statistics and Offline Analysis
3. Results
3.1. Measurement of PMCA1–4 mRNA Levels in Human Odontoblasts
3.2. Extrusion of Ca2+ by PMCA Following Hypotonic or High-pH Stimulation in Acutely Isolated Rat Odontoblasts and HOB Cells
3.3. PMCA Mediates Mineralization by Odontoblasts
3.4. Effects of PMCA Inhibitor on Detected mRNA Levels of Non-Collagenous Extracellular Matrix Proteins as Odontoblast Markers
3.5. Immunofluoresence Analysis of PMCA1 in HOB Cells
4. Discussion
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Name | 5′-Sequence-3′ | GenBank Number |
---|---|---|
β-actin | Forward TGGCACCCAGCACAATGAA Reverse CTAAGTCATAGTCCGCCTAGAAGCA | NM_001101.5 |
PMCA1 | Forward ACCATATGCTAGAATGCCCACCTC Reverse CTGGTGAAATCTGGGCCCTAAC | NM_001001323.2 |
PMCA2 | Forward AGAGCTTCCGCATGTACAGCAA Reverse CAAGCCATGGGCTCAATCAC | NM_001001331.4 |
PMCA3 | Forward CGTAACGTCTATGACAGCATCTCCA Reverse TCCATGATCAAGTTCACCCACAA | NM_001001344.2 |
PMCA4 | Forward TGGCATGGTTAAATCTGAATGG Reverse CTGCTTCAATTGTAAGGCAAAGG | NM_001001396.2 |
DMP-1 | Forward TCCAGTCTCACAGCAGCTCA Reverse TCTCCGTGGAGTTGCTATCTTC | NM_004407.4 |
DSPP | Forward TGATAGCAGTGACAGCACATCTGAC Reverse GTTGTTACCGTTACCAGACTTGCTC | NM_014208.3 |
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Kimura, M.; Mochizuki, H.; Satou, R.; Iwasaki, M.; Kokubu, E.; Kono, K.; Nomura, S.; Sakurai, T.; Kuroda, H.; Shibukawa, Y. Plasma Membrane Ca2+–ATPase in Rat and Human Odontoblasts Mediates Dentin Mineralization. Biomolecules 2021, 11, 1010. https://doi.org/10.3390/biom11071010
Kimura M, Mochizuki H, Satou R, Iwasaki M, Kokubu E, Kono K, Nomura S, Sakurai T, Kuroda H, Shibukawa Y. Plasma Membrane Ca2+–ATPase in Rat and Human Odontoblasts Mediates Dentin Mineralization. Biomolecules. 2021; 11(7):1010. https://doi.org/10.3390/biom11071010
Chicago/Turabian StyleKimura, Maki, Hiroyuki Mochizuki, Ryouichi Satou, Miyu Iwasaki, Eitoyo Kokubu, Kyosuke Kono, Sachie Nomura, Takeshi Sakurai, Hidetaka Kuroda, and Yoshiyuki Shibukawa. 2021. "Plasma Membrane Ca2+–ATPase in Rat and Human Odontoblasts Mediates Dentin Mineralization" Biomolecules 11, no. 7: 1010. https://doi.org/10.3390/biom11071010
APA StyleKimura, M., Mochizuki, H., Satou, R., Iwasaki, M., Kokubu, E., Kono, K., Nomura, S., Sakurai, T., Kuroda, H., & Shibukawa, Y. (2021). Plasma Membrane Ca2+–ATPase in Rat and Human Odontoblasts Mediates Dentin Mineralization. Biomolecules, 11(7), 1010. https://doi.org/10.3390/biom11071010