Inactivation of PI3K/Akt promotes the odontoblastic differentiation and suppresses the stemness with autophagic flux in dental pulp cells

Sam Young Park; Heui Seung Cho; Kyung Hwun Chung; Bin Na Lee; Sun Hun Kim; Won Jae Kim; Ji Yeon Jung

doi:10.1016/j.jds.2021.05.013

Inactivation of PI3K/Akt promotes the odontoblastic differentiation and suppresses the stemness with autophagic flux in dental pulp cells

J Dent Sci. 2022 Jan;17(1):145-154. doi: 10.1016/j.jds.2021.05.013. Epub 2021 Jun 16.

Authors

Sam Young Park¹, Heui Seung Cho¹, Kyung Hwun Chung², Bin Na Lee³, Sun Hun Kim⁴, Won Jae Kim¹, Ji Yeon Jung¹

Affiliations

¹ Department of Oral Physiology, Dental Science Research Institute, School of Dentistry, Chonnam National University, Gwangju, Republic of Korea.
² Dental Research Institute, School of Dentistry, Seoul National University, Seoul, Republic of Korea.
³ Department of Operative Dentistry, Dental Science Research Institute, School of Dentistry, Chonnam National University, Gwangju, Republic of Korea.
⁴ Department of Oral Anatomy, Dental Science Research Institute, School of Dentistry, Chonnam National University, Gwangju, Republic of Korea.

Abstract

Background/purpose: Autophagy is involved in controlling differentiation of various cell types. The present study aimed to investigate the mechanism related to autophagy in regulating odontogenic differentiation of dental pulp cells.

Materials and methods: Human dental pulp cells (HDPCs) were cultured in differentiation inductive medium (DM) and odontoblastic differentiation and mineralization were evaluated by alkaline phosphatase (ALP) staining and Alizarin red S staining, respectively. Tooth cavity preparation was made on the mesial surface of lower first molars in rat. The expression of autophagy-related signal molecules was detected using Western blot analysis and Immunohistochemistry.

Results: HDPCs cultured in DM showed increased autophagic flux and declined phosphorylation of phosphoinositide 3-kinases (PI3K), protein kinase B (Akt), and mTOR. Dentin matrix protein-1 (DMP-1) and dentin sialoprotein (DSP), markers of odontoblastic differentiation, were upregulated and autophagic activation showing increased LC3-II and decreased p62 levels was observed during odontogenic differentiation of HDPCs. However, PI3K blocker 3-methyladenine (3MA), lentiviral shLC3 and Akt activator SC79 attenuated the expression of LC3II as well as DMP-1, ALP activity and mineralization enhanced in HDPCs under DM condition. In addition, 3MA, shLC3 and SC79 recovered the expression of pluripotency factor CD146, Oct4 and Nanog downregulated in DM condition. In rat tooth cavity preparation model, the expression of LC3B and DMP-1 was elevated near odontoblast-dentin layer during reparative dentin formation, whereas 3MA significantly reduced the expression of LC3B and DMP-1.

Conclusion: These findings indicated autophagy promotes the odontogenic differentiation of dental pulp cells modulating stemness via PI3K/Akt inactivation and the repair of pulp.

Keywords: Akt; Autophagy; Dental pulp cells; Odontogenic differentiation; Stemness.