Effects of low level of calcium on the biological behavior of rat primary ameloblasts cultured in vitro / 西安交通大学学报(医学版)

ABSTRACT

【Objective】 To explore the effect of low levels of calcium on the biological characteristics of ameloblasts. 【Methods】 Rat primary ameloblasts were cultured in standard DMEM medium. After five days they were identified by RT-PCR and immunohistochemistry. Then 0, 0.6 and 1.2 mmoL/L Ca2+ and 100 mL/L fetal bovine serum were added into DMEM medium without calcium. After 48 hours, the cell morphology was observed by inverted microscope. The proliferation and apoptosis of cells were separately examined by MTT and AnnexinV-PI. Real-time quantitative PCR was used to detect the expression levels of amelogenin and KLK4 mRNA. 【Results】 After Five days in standard DMEM medium, the cells were shaped like the paving pattern. RT-PCR showed that both amelogenin and KLK4 were expressed in the cells. Immunohistochemical staining showed that most cells had positive staining for amelogenin. After 48 hours of calcium intervention, some cells in 1.2 mmoL/L Ca2+ group had higher nuclear density and poor light transmittance, and more high columnar cells could be observed in 1.2 mmoL/L Ca2+ group than those in 0 and 0.6 mmoL/L Ca2+ groups. With the decrease in calcium concentration in the medium, MTT showed that the proliferation activity of ameloblasts reduced (P<0.01). Annexin V-PI showed that the percentage of apoptotic cells decreased, and there was a significant difference between 1.2 mmoL/L and 0 mmoL/L Ca2+ groups (P<0.05). Real time-PCR showed that the expressions of amelogenin and KLK4 mRNA reduced (P<0.01). 【Conclusion】 Low-level calcium may inhibit the differentiation of ameloblasts, thereby affecting the formation of enamel mineralization.