钙对成釉细胞系ALC细胞生物学特性的影响

doi:10.13591/j.cnki.kqyx.2023.01.007

摘要/Abstract

摘要： 目的观察钙对成釉细胞系ALC细胞增殖、凋亡及细胞周期等生物学特性的影响。方法培养ALC细胞系,在高糖DMEM培养基中配制0、2.0、2.5、3.0、3.5 mmol/L CaCl₂溶液,利用倒置显微镜检测在两种培养时间(24、48 h)下ALC细胞形态是否改变。通过CCK-8法分析钙离子对ALC细胞增殖的作用,运用Hoechst染色观察钙离子对ALC细胞凋亡的影响;通过PI染色FCM法分析钙离子对ALC细胞生长周期的干预。利用Western blot检测钙离子对ALC细胞中周期蛋白Cyclin A、Cyclin B、Cyclin D表达的调节。结果 0 mmol/L CaCl₂组,ALC细胞形态呈卵圆或多边形,细胞间连接紧密呈铺路石样生长。其他不同浓度的钙干预24、48 h后,ALC细胞形态未见明显变化;CCK-8法结果显示钙干预24、48 h后,随着钙浓度增加,ALC细胞的存活率呈微弱降低趋势,但此趋势无统计学意义;Hoechst染色结果显示不同浓度的钙干预24、48 h后,ALC细胞的凋亡数目随时间延长可见增加,但不同钙离子浓度组间差异并不显著;PI染色FCM检测显示随着钙离子浓度的升高,ALC细胞的细胞周期中S期逐渐增加,G₁期、G₂期呈下降趋势;Western blot结果表明钙干预24、48 h后,ALC细胞中的周期蛋白Cyclin A、Cyclin B表达呈下降趋势,Cyclin D表达呈上升趋势。结论在本研究条件下,钙对ALC细胞增殖、凋亡未见明显影响,但改变了细胞周期。通过实验研究结果发现本实验中的钙离子浓度对成釉细胞无明显毒副作用,可用于进一步探讨钙对氟牙症治疗的可能机制及作用。

关键词: ALC细胞, 钙, 增殖, 凋亡

Abstract: Objective To observe the effect of calcium on biological characteristics (proliferation, apoptosis and cell cycle) of ALC ameloblasts. Methods ALC cell lines were cultured in vitro in DMEM medium with high glucose at different concentrations (0, 2.0, 2.5, 3.0 and 3.5 mmol/L CaCl₂ aqueous solution) for 24 h and 48 h, respectively. Changes of ALC cells under two kinds of incubation time were observed with an inverted microscope. CCK-8 method was used to analyze the effect of calcium ion on ALC cell proliferation. Hoechst staining was used to observe the effect of calcium ion on ALC cell apoptosis. PI staining and FCM method were used to analyze the effect of calcium ions on the growth cycle of ALC cells. Western blot was used to detect the effect of calcium ions on the expression of Cyclin A, Cyclin B and Cyclin D in ALC cells. Results In the 0 mmol/L CaCl₂ group, ALC cells were oval or polygonal in shape, and the cells were closely connected and grew like paving stones. In other concentration groups, the morphology of ALC cells did not change significantly after calcium intervention for 24 h and 48 h. Results of CCK-8 method showed that the survival rate of ALC cells slightly decreased with increasing calcium ions concentration after calcium intervention for 24 h and 48 h. However, there was no significant differences in this trend. Results of Hoechst staining showed that the number of ALC cell apoptosis did not increase significantly after different concentrations of calcium intervention for 24 h and 48 h. With the increase of calcium ion concentration, results of PI staining and FCM method showed that the cell cycle of ALC cells gradually increased in S phase and decreased in G₁ and G₂ phase gradually. Western blot results showed that the expression of Cyclin A and Cyclin B in ALC cells decreased and the expression of Cyclin D increased after different concentrations of calcium intervention for 24 h and 48 h. Conclusion In this study, calcium has no significant effect on the proliferation and apoptosis of ALC cells. Calcium, however, has an effect on the ALC cell cycle. Results of this study show that calcium ions has no obvious toxic or side effects on the ameloblasts, which could be used to explore the possible mechanism and effect of calcium on dental fluorosis.

Key words: ALC cells, calcium, proliferation, apoptosis

中图分类号:

R329.28

高震,侯瑞凯,宋索成,阮建平. 钙对成釉细胞系ALC细胞生物学特性的影响[J]. 口腔医学, 2023, 43(1): 39-45.

GAO Zhen,HOU Ruikai,SONG Suocheng,RUAN Jianping. Effect of calcium on biological properties of the ameloblast ALC[J]. Stomatology, 2023, 43(1): 39-45.

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