Comparison of 2,3,5,4'-tetrahydroxystilbene-2-O-b-D-glucoside-induced proliferation and differentiation of dental pulp stem cells in 2D and 3D culture systems-gene analysis

Yen Wu; Yao-Yu Chung; Yu-Tang Chin; Chi-Yu Lin; Po-Jan Kuo; Ting-Yi Chen; Tzu-Yu Lin; Hsien-Chung Chiu; Haw-Ming Huang; Jiiang-Huei Jeng; Sheng-Yang Lee

doi:10.1016/j.jds.2021.09.021

Comparison of 2,3,5,4'-tetrahydroxystilbene-2-O-b-D-glucoside-induced proliferation and differentiation of dental pulp stem cells in 2D and 3D culture systems-gene analysis

J Dent Sci. 2022 Jan;17(1):14-29. doi: 10.1016/j.jds.2021.09.021. Epub 2021 Sep 24.

Authors

Yen Wu^{1

2

3}, Yao-Yu Chung^{1

3}, Yu-Tang Chin^{1

3}, Chi-Yu Lin^{1

3}, Po-Jan Kuo⁴, Ting-Yi Chen^{1

2

3}, Tzu-Yu Lin^{1

2

3}, Hsien-Chung Chiu⁴, Haw-Ming Huang¹, Jiiang-Huei Jeng^{5

6}, Sheng-Yang Lee^{1

2

3}

Affiliations

¹ School of Dentistry, College of Oral Medicine, Taipei Medical University, Taipei, Taiwan.
² Department of Dentistry, Wan-Fang Medical Center, Taipei Medical University, Taipei, Taiwan.
³ Center for Tooth Bank and Dental Stem Cell Technology, Taipei Medical University, Taipei, Taiwan.
⁴ Department of Periodontology, School of Dentistry, National Defense Medical Center and Tri-Service General Hospital, Taipei, Taiwan.
⁵ School of Dentistry, College of Dental Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan.
⁶ Department of Dentistry, Kaohsiung Medical, University Hospital, Kaohsiung, Taiwan.

Abstract

Background/purpose: Culture environments play a critical role in stem cell expansion. This study aimed to evaluate the effects of 2,3,5,4'-tetrahydroxystilbene-2-O-b-D-glucoside (THSG) on the proliferation and differentiation of human dental pulp stem cells (DPSCs) in 2-dimensional (2D) and 3-dimensional (3D) culture systems.

Materials and methods: Human DPSCs were seeded in T25 flasks for 2D cultivation. For the 3D culture system, DPSCs were mixed with microcarriers and cultured in spinner flasks. Cells in both culture systems were treated with THSG, and cell proliferation was determined using a cell counter and a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide assay. In THSG-treated DPSCs, the genes associated with proliferation, adipogenesis, neurogenesis, osteogenesis, pluripotency, oncogenesis, and apoptosis were analyzed using real-time polymerase chain reactions.

Results: The spinner flask time-dependently improved cell numbers, cell viability, and expansion rates in THSG-treated DPSCs. In both the T25 and spinner flasks, the messenger RNA (mRNA) levels of proliferation, osteogenesis, and pluripotent-related genes had a significant maximum expression with 10 μM THSG treatment. However, 0.1 μM of THSG may be the most suitable condition for triggering neurogenesis and adipogenesis gene expression when DPSCs were cultured in spinner flasks. Furthermore, the number of oncogenes and apoptotic genes decreased considerably in the presence of THSG in both the T25 and spinner flasks.

Conclusion: The spinner flask bioreactor combined with THSG may upregulate proliferation and lineage-specific differentiation in DPSCs. Thus, the combination can be used to mass-produce and cultivate human DPSCs for regenerative dentistry.

Keywords: 2,3,5,4′-tetrahydroxystilbene-2-O-B-glucoside; Dental pulp stem cells; Expansion.