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. 2022 Jul;17(3):1225-1231.
doi: 10.1016/j.jds.2021.12.003. Epub 2021 Dec 20.

The effect of AMP kinase activation on differentiation and maturation of osteoblast cultured on titanium plate

Phanthavong Vansana  1 , Kae Kakura  1 , Yusuke Taniguchi  1 , Kei Egashira  1 , Etsuko Matsuzaki  2   3 , Takashi Tsutsumi  3   4 , Hirofumi Kido  1
Affiliations

The effect of AMP kinase activation on differentiation and maturation of osteoblast cultured on titanium plate

Phanthavong Vansana et al. J Dent Sci. 2022 Jul.

Abstract

Background/purpose: 5' Adenosine monophosphate-activated protein kinase (AMPK) is known as an enzyme that maintains intracellular homeostasis and has various biological activity. The purpose of this study is evaluation effect of AMPK activation on implant prognosis.

Materials & methods: MC3T3-E1 osteoblast-like cells were cultured on titanium using a 24-well plate. The experimental group was divided into the following 3 groups: (1) the normal culture group (control group), (2) the osteogenic induction group, and (3) the osteogenic induction + AMPK activation group. The cell counts were measured; real-time PCR was used to assess the expression of ALP and Osterix as osteogenic related genes at Day 0,7,14 and 21 after experiments. Additionally, ALP activity and calcification were assessed.

Results: The results of the real-time PCR assessments revealed that the expression of ALP, which is a marker for the initial stages of calcification, was significantly increased by AMPK activation compared to the normal culture or osteogenic induction. A significant increase was also observed in the expression of Osterix, which is a marker for the later stages of calcification. Because significant increases were observed in ALP activity and calcification potential, this suggested that AMPK activation could elicit an increase in osteoblast calcification potential.

Conclusion: AMPK activation promotes implant peripheral osteoblast differentiation and maturation and enhances calcification. Our results suggest that AMPK activation may help to maintain implant stability.

Keywords: 5′ adenosine monophosphate-activated protein kinase; Acadesine/AICA riboside; Dental implant; Osteoblast.

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Conflict of interest statement

The authors declare no conflict of interest in this study.

Figures

Figure 1
Figure 1
Cell proliferation assay of MC3T3-E1 cells cultured on titanium plate. Counting cells was performed with Cell Counting Kit-8 measuring 405 um absorbance at days 7, 14, and 21 after experiments in control group, OG group and OG + AICAR group. (∗p < 0.05, compared to the control group).
Figure 2
Figure 2
Evaluation of mRNA expression levels of ALP; Alkaline phosphatase and Osterix of MC3T3-E1 cells cultured on titanium plate was performed by real-time PCR at day 7, 14, and 21 after experiments in control group, OG group and OG + AICAR group. (A) ALP. (B) Osterix. (∗p < 0.05, compared to the control group).
Figure 3
Figure 3
Evaluation of ALP activity of MC3T3-E1 cells cultured on titanium plate was performed using ALP Assay kit measuring 405um absorbance. Measurement was performed at day 7, 14, and 21 after experiments in control group, OG group and OG + AICAR group. (∗p < 0.05, compared to the control group).
Figure 4
Figure 4
Mineralization of MC3T3-E1 cells cultured on titanium plate was evaluated using Alizarin red staining kit. (A) Representative image of MC3T3-E1 cells cultured on titanium plate stained by Alizarin red at day 7, 14, and 21 after experiments in control group, OG group and OG + AICAR group. Original magnification × 40.The height and width of titanium plate placed in center is 10 mm × 10 mm as mentioned above. (B) Evaluation of mineralization was performed measuring 405um absorbance at day 7, 14, and 21 after experiments in control group, OG group and OG + AICAR group. (∗p < 0.05, compared to the control group).
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