Human Recombinant Lactoferrin Promotes Differentiation and Calcification on MC3T3-E1 Cells
Abstract
:1. Introduction
2. Materials and Methods
2.1. Cell Culture and Materials
2.2. Cell Viability Assay
2.3. Calcification Assay
2.4. ALP Activity Assay
2.5. Immunocytochemical Staining
2.6. RNA Isolation and Real-Time Quantitative PCR (RT-qPCR)
2.7. Statistical Analysis
3. Results
3.1. Effect of LF on Osteoblast Proliferation in Two Types of Media
3.2. Evaluation of Calcification Using Alizarin Red Staining
3.3. Number of ALP-Positive Cells
3.4. Immunocytochemical Staining of OPN Expression
3.5. Osteocalcin, Runx2, and Osterix Expression with RT-qPCR
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Abbreviation | Source | Purity |
---|---|---|
LF-I | Bovine | >95% |
LF-II | Human recombinant | >95% |
Mouse | Universal Probe Library | Forward Primer | Reverse Primer |
---|---|---|---|
Osteocalcin | #32 | 5′-AGACTCCGGCGCTACCTT-3′ | 5′-CTCGTCACAAGCAGGGTTAAG-3′ |
Runx2 | #34 | 5′-GCCCAGGCGTATTTCAGA-3′ | 5′-TGCCTGGCTCTTCTTACTGAG-3’ |
Osterix | #106 | 5′-CTCCTGCAGGCAGTCCTC-3′ | 5′-GGGAAGGGTGGGTAGTCATT-3′ |
β-actin | #64 | 5′-CTAAGGCCAACCGTGAAAAG-3′ | 5′-ACCAGAGGCATACAGGGACA-3′ |
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Nagashima, D.; Ishibashi, Y.; Kawaguchi, S.; Furukawa, M.; Toho, M.; Ohno, M.; Nitto, T.; Izumo, N. Human Recombinant Lactoferrin Promotes Differentiation and Calcification on MC3T3-E1 Cells. Pharmaceutics 2023, 15, 60. https://doi.org/10.3390/pharmaceutics15010060
Nagashima D, Ishibashi Y, Kawaguchi S, Furukawa M, Toho M, Ohno M, Nitto T, Izumo N. Human Recombinant Lactoferrin Promotes Differentiation and Calcification on MC3T3-E1 Cells. Pharmaceutics. 2023; 15(1):60. https://doi.org/10.3390/pharmaceutics15010060
Chicago/Turabian StyleNagashima, Daichi, Yukiko Ishibashi, Sachiko Kawaguchi, Megumi Furukawa, Masahiro Toho, Megumi Ohno, Takeaki Nitto, and Nobuo Izumo. 2023. "Human Recombinant Lactoferrin Promotes Differentiation and Calcification on MC3T3-E1 Cells" Pharmaceutics 15, no. 1: 60. https://doi.org/10.3390/pharmaceutics15010060
APA StyleNagashima, D., Ishibashi, Y., Kawaguchi, S., Furukawa, M., Toho, M., Ohno, M., Nitto, T., & Izumo, N. (2023). Human Recombinant Lactoferrin Promotes Differentiation and Calcification on MC3T3-E1 Cells. Pharmaceutics, 15(1), 60. https://doi.org/10.3390/pharmaceutics15010060