Abstract:

Objective: This study aimed to evaluate the role of long intergenic non-coding RNA-EPS (lincRNA-EPS) in enamel development using lincRNA-EPS knockout(KO) mice model. Methods: The genotypes of lincRNA-EPS KO mice were identified by polymerase chain reaction(PCR). The mandibular molars of 8-week-old KO mice and wildtype (WT) mice was analyzed by micro-CT. The morphology and microstructure of enamel were detected by scanning electron microscope (SEM). The degree of enamel mineralization was explored by X-ray and energy dispersive X-ray spectroscopy (EDX). Integrated mandibular first molar germs of 0.5 day post-natal mice were isolated and used for real-time quantitative polymerase chain reaction(RT-qPCR) to detect the relative mRNA expression of AMELX, ENAM, AMBN, AMTN. Results: Micro-CT imaging showed that compared with WT mice, lingual side of mandibular molar exhibited large enamel defects in lincRNA-EPS KO mice, and lingual cusps were worn into pits. SEM analysis showed that the enamel from lincRNA-EPS KO mice displayed structural disorganization. Moreover, there were gaps between the hydroxyapatite crystals in the rods. Compared with WT mice, the relative mRNA expression of enamel matrix protein significantly decreased in KO mice(P<0.05). Conclusion: Deficiency of lincRNA-EPS may lead to alternation of the gene expression of enamel matrix protein, which resulted in defects in the structure of enamel in KO mice. Our findings support an essential role of lincRNA-EPS in the process of amelogenesis.

Key words: lincRNA-EPS, amelogenesis, knockout mouse, tooth phenotypes

摘要：

目的： 利用基因间长链非编码RNA-EPS（long intergenic non-coding RNA- EPS，lincRNA-EPS）基因敲除小鼠模型研究lincRNA-EPS对牙釉质发育的影响。方法： 应用聚合酶链式反应（polymerase chain reaction，PCR）分析方法鉴定lincRNA-EPS基因敲除小鼠的基因型；微型CT（micro-CT）检测8周龄lincRNA-EPS基因敲除小鼠和野生型小鼠下颌磨牙区的形态；扫描电子显微镜（scanning electron microscope，SEM）观察下颌磨牙区釉质的微观结构；X线及X射线能谱分析（energy dispersive X-ray spectroscopy，EDX）检测釉质矿化程度的改变；体视显微镜下分离PN0.5小鼠下颌第一磨牙牙胚；实时定量聚合酶链式反应（real-time quantitative polymerase chain reaction，RT-qPCR）技术检测其釉质发育相关基因AMELX、ENAM、AMBN、AMTN的mRNA表达量。结果： 与野生型小鼠相比，micro-CT图像显示lincRNA-EPS基因敲除小鼠下颌磨牙舌侧出现大面积釉质缺损，舌侧牙尖磨耗成凹坑状；SEM结果显示，lincRNA-EPS基因敲除小鼠釉质结构紊乱，釉柱内羟基磷灰石晶体间存在间隙；lincRNA-EPS基因敲除小鼠下颌磨牙牙胚中，AMELX、ENAM、AMBN、AMTN的mRNA表达量较野生型小鼠显著降低，差异存在统计学意义（P<0.05）。结论： LincRNA-EPS的缺失导致釉质发育相关基因表达下调，并引起牙釉质结构的紊乱，最终导致了釉质结构的缺损，证明lincRNA-EPS在牙釉质发育过程中具有重要作用。

关键词: 长链非编码RNA-EPS, 釉质发育, 基因敲除小鼠, 牙齿表型

CLC Number: 

• R782