[Experimental study of odontoblast differentiation of stem cells from human exfoliated deciduous teeth induced by
Wenyan Huang1, Yuhang Huang, Yanbing He
1Department of Pediatric Dentistry, School and Hospital of Stomatology, Guangzhou Medical University; Guangdong Engineering Research Center of Oral Restoration and Reconstruction; Guangzhou Key Laboratory of Basic and Applied Research of Oral Regenerative Medicine. Guangzhou 510182, China.
Purpose:
To simulate the mineralizing fluid with lithium chloride (LiCl) as a dentinogenic environment and investigate its potential to induce stem cells from human exfoliated deciduous teeth (SHEDs) into dentinogenesis.
Methods:
Primary SHEDs were isolated, cultured, and identified, and P3-6 SHEDs were used for subsequent experiments. Osteogenic medium(OM) containing 0, 1.0, 2.5, 5.0, 10 mmol/L LiCl were prepared. Cell proliferation was assessed using CCK-8 assay at 1, 3, 5, and 7 days. Alkaline phosphatase(ALP) activity was examined by staining at 4, 7, and 10 days. Gene expression levels of DSPP, Runx2 and COL-1 were detected by RT-PCR, and protein levels of DSPP and Runx2 were analyzed by Western blot at 10 days. Alizarin red staining and semi-quantitative analysis were conducted at 14 days to evaluate mineralization capacity and select the optimal LiCl concentration.
Results:
CCK-8 assay showed that low concentrations(≤2.5 mmol/L) of LiCl had minimal effect on cell proliferation from 1 to 7 days, while high concentrations (＞2.5 mmol/L) inhibited cell proliferation. ALP activity was enhanced by low concentrations of LiCl at 4, 7, and 10 days, with the 2.5 mmol/L LiCl group showing the most significant effect, while high concentrations suppressed ALP activity. Western blot results revealed that low concentrations of LiCl promoted the protein expression of DSPP and Runx2 in SHEDs. RT-PCR demonstrated that low concentrations of LiCl upregulated the gene expression of DSPP and COL-1 in SHEDs. Qualitative and quantitative Alizarin red staining indicated that 1.0, 2.5, 5.0 mmol/L LiCl improved SHEDs mineralization capacity, while 10 mmol/L LiCl showed no significant difference in mineralized nodule formation compared to the control group.
Conclusions:
Low concentrations of lithium compounds can promote the dentinogenesis of SHEDs.
