Skip to main content

Strategies for differentiation of hiPSCs into dental epithelial cell lineage

Abstract

Different stem cell–based strategies, especially induced pluripotent stem cells (iPSCs), have been exploited to regenerate teeth or restore biological and physiological functions after tooth loss. Further research is needed to establish an optimized protocol to effectively differentiate human iPSCs (hiPSCs) into dental epithelial cells (DECs). In this study, various factors were precisely modulated to facilitate differentiation of hiPSCs into DECs, which are essential for the regeneration of functional teeth. Embryoid bodies (EBs) were formed from hiPSCs as embryo-like aggregates, retinoic acid (RA) was used as an early ectodermal inducer, and bone morphogenic protein 4 (BMP4) activity was manipulated. The characteristics of DECs were enhanced and preserved after culture in keratinocyte serum-free medium (K-SFM). The yielded cell population exhibited noticeable DEC characteristics, consistent with the expression of epithelial cell and ameloblast markers. DECs demonstrated odontogenic abilities by exerting an inductive effect on human dental pulp stem cells (hDPSCs) and forming a tooth-like structure with the mouse tooth mesenchyme. Overall, our differentiation protocol provides a practical approach for applying hiPSCs for tooth regeneration.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3

References

  1. Almushayt A, Narayanan K, Zaki AE, George A (2006) Dentin matrix protein 1 induces cytodifferentiation of dental pulp stem cells into odontoblasts. Gene Ther 13:611–620

    CAS  Article  Google Scholar 

  2. Arakaki M, Ishikawa M, Nakamura T, Iwamoto T, Yamada A, Fukumoto E, Saito M, Otsu K, Harada H, Yamada Y, Fukumoto S (2012) Role of epithelial-stem cell interactions during dental cell differentiation. J Biol Chem 287:10590–10601

    CAS  Article  Google Scholar 

  3. Balic A (2019) Concise review: cellular and molecular mechanisms regulation of tooth initiation. Stem Cells 37:26–32

    Article  Google Scholar 

  4. Baranova J, Buchner D, Gotz W, Schulze M, Tobiasch E (2020) Tooth formation: are the hardest tissues of human body hard to regenerate? Int J Mol Sci 21:4031

  5. Cai J, Zhang Y, Liu P, Chen S, Wu X, Sun Y, Li A, Huang K, Luo R, Wang L, Liu Y, Zhou T, Wei S, Pan G, Pei D (2013) Generation of tooth-like structures from integration-free human urine induced pluripotent stem cells. Cell Regen 2:6

    Article  Google Scholar 

  6. Gronthos S, Mankani M, Brahim J, Robey PG, Shi S (2000) Postnatal human dental pulp stem cells (DPSCs) in vitro and in vivo. Proc Natl Acad Sci U S A 97:13625–13630

    CAS  Article  Google Scholar 

  7. Guo NN, Liu LP, Zhang YX, Cai YT, Guo Y, Zheng YW, Li YM (2019) Early prediction of the differentiation potential during the formation of human iPSC-derived embryoid bodies. Biochem Biophys Res Commun 516:673–679

    CAS  Article  Google Scholar 

  8. Harvey NT, Hughes JN, Lonic A, Yap C, Long C, Rathjen PD, Rathjen J (2010) Response to BMP4 signalling during ES cell differentiation defines intermediates of the ectoderm lineage. J Cell Science 123:1796–1804

    CAS  Article  Google Scholar 

  9. Honda MJ, Fong H, Iwatsuki S, Sumita Y, Sarikaya M (2008) Tooth-forming potential in embryonic and postnatal tooth bud cells. Med Mol Morphol 41:183–192

    Article  Google Scholar 

  10. Ikeda E, Morita R, Nakao K, Ishida K, Nakamura T, Takano-Yamamoto T, Ogawa M, Mizuno M, Kasugai S, Tsuji T (2009) Fully functional bioengineered tooth replacement as an organ replacement therapy. Proc Natl Acad Sci U S A 106:13475–13480

    CAS  Article  Google Scholar 

  11. Jia S, Zhou J, Gao Y, Baek JA, Martin JF, Lan Y, Jiang R (2013) Roles of Bmp4 during tooth morphogenesis and sequential tooth formation. Development 140:423–432

    CAS  Article  Google Scholar 

  12. Kamocki K, Nor JE, Bottino MC (2015) Dental pulp stem cell responses to novel antibiotic-containing scaffolds for regenerative endodontics. Int Endod J 48:1147–1156

    CAS  Article  Google Scholar 

  13. Kawano E, Toriumi T, Iguchi S, Suzuki D, Sato S, Honda M (2017) Induction of neural crest cells from human dental pulp-derived induced pluripotent stem cells. Biomed Res 38(2):135–147

    CAS  Article  Google Scholar 

  14. Kim EJ, Yoon KS, Arakaki M, Otsu K, Fukumoto S, Harada H, Green DW, Lee JM, Jung HS (2019) Effective differentiation of induced pluripotent stem cells into dental cells. Dev Dyn 248:129–139

    Article  Google Scholar 

  15. Kim GH, Yang J, Jeon DH, Kim JH, Chae GY, Jang M, Lee G (2020) Differentiation and establishment of dental epithelial-like stem cells derived from human ESCs and iPSCs. Int J Mol Sci 21:4384

  16. Lee JH, Seo SJ (2016) Biomedical application of dental tissue-derived induced pluripotent stem cells. Stem Cells Int 2016:9762465

    PubMed  PubMed Central  Google Scholar 

  17. Li Q, Zhang S, Sui Y, Fu X, Li Y, Wei S (2019) Sequential stimulation with different concentrations of BMP4 promotes the differentiation of human embryonic stem cells into dental epithelium with potential for tooth formation. Stem Cell Res Ther 10:276

    Article  Google Scholar 

  18. Lin H, Xu L, Liu H, Sun Q, Chen Z, Yuan G, Chen Z (2011) KLF4 promotes the odontoblastic differentiation of human dental pulp cells. J Endod 37:948–954

    Article  Google Scholar 

  19. Moshy E, Radwan IA, Rady D, Abbass MMS, El-Rashidy AA, Sadek KM, Dorfer CE, Fawzy El-Sayed KM (2020) Dental stem cell-derived secretome/conditioned medium: the future for regenerative therapeutic applications. Stem Cells Int 2020:7593402

    Article  Google Scholar 

  20. Otsu K, Kishigami R, Oikawa-Sasaki A, Fukumoto S, Yamada A, Fujiwara N, Ishizeki K, Harada H (2012) Differentiation of induced pluripotent stem cells into dental mesenchymal cells. Stem Cells Dev 21:1156–1164

    CAS  Article  Google Scholar 

  21. Ross SA, McCaffery PJ, Drager UC, De Luca LM (2000) Retinoids in embryonal development. Physiol Rev 80:1021–1054

    CAS  Article  Google Scholar 

  22. Shinmura Y, Tsuchiya S, Hata K, Honda MJ (2008) Quiescent epithelial cell rests of Malassez can differentiate into ameloblast-like cells. J Cell Physiol 217:728–738

    CAS  Article  Google Scholar 

  23. Smith EE, Angstadt S, Monteiro N, Zhang W, Khademhosseini A, Yelic PC (2018) Bioengineered tooth buds exhibit features of natural tooth buds. J Dent Res 97(10):1144–1151

    CAS  Article  Google Scholar 

  24. St Amand TR, Zhang Y, Semina EV, Zhao X, Hu Y, Nguyen L, Murray JC, Chen Y (2000) Antagonistic signals between BMP4 and FGF8 define the expression of Pitx1 and Pitx2 in mouse tooth-forming anlage. Dev Biol 217:323–332

    CAS  Article  Google Scholar 

  25. Thesleff I (2003) Epithelial-mesenchymal signalling regulating tooth morphogenesis. J Cell Sci 116:1647–1648

    CAS  Article  Google Scholar 

  26. Wu Z, Wang F, Fan Z, Wu T, He J, Wang J, Zhang C, Wang S (2019) Whole-tooth regeneration by allogeneic cell reassociation in pig jawbone. Tissue Eng Part A 25:1202–1212

    CAS  Article  Google Scholar 

  27. Yang R, Zheng Y, Burrows M, Liu S, Wei Z, Nace A, Guo W, Kumar S, Cotsarelis G, Xu X (2014) Generation of folliculogenic human epithelial stem cells from induced pluripotent stem cells. Nat Commun 5:3071

    Article  Google Scholar 

  28. Zhang C, Lee HJ, Shrivastava A, Wang R, McQuiston TJ, Challberg SS, Pollok BA, Wang T (2018) Long-term in vitro expansion of epithelial stem cells enabled by pharmacological inhibition of PAK1-ROCK-myosin II and TGF-beta signaling. Cell Rep 25:598–610

  29. Zhang Y, Li Y, Shi R, Zhang S, Liu H, Zheng Y, Li Y, Cai J, Pei D, Wei S (2017) Generation of tooth-periodontium complex structures using high-odontogenic potential dental epithelium derived from mouse embryonic stem cells. Stem Cell Res Ther 8:141

    Article  Google Scholar 

Download references

Funding

This research was supported by the Bio & Medical Technology Development Program of the National Research Foundation (NRF) and funded by the Korean government (MSIP&MOHW) (No. 2017M3A9E4048172). This research was financially supported by grants from the National Research Foundation of Korea (NRF) Grant funded by the Korean Government (MSIP) (NRF-2019R1A2C3005294).

Author information

Affiliations

Authors

Corresponding author

Correspondence to Han-Sung Jung.

Ethics declarations

Ethics approval

All experiments were performed according to the guidelines of the Yonsei University College of Dentistry, Intramural Animal Care and Use Committee (2018–0183).

Conflict of interest

The authors declare no competing interests.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Kim, EJ., Mai, H.N., Lee, DJ. et al. Strategies for differentiation of hiPSCs into dental epithelial cell lineage. Cell Tissue Res (2021). https://doi.org/10.1007/s00441-021-03512-w

Download citation

Keywords

  • Tooth development
  • Tooth regeneration
  • hiPSCs
  • Dental epithelial cell
  • BMP4