Department of Stomatology, Guangzhou Women and Children's Medical Center, Guangzhou, China.
International Medical Center, Guangdong Second Provincial General Hospital, Guangzhou, China.
Int Endod J. 2019 Apr;52(4):491-503. doi: 10.1111/iej.13032. Epub 2018 Dec 8.
To profile miRNA expression between inflamed and healthy human dental pulp tissues and to investigate how the upregulation of miR-223-3p in the inflamed pulp tissue regulates odontoblast differentiation and regeneration.
Microarray analysis was used to identify differences in miRNA expression patterns between healthy and inflamed pulp tissue. The results were validated using quantitative real-time PCR. To determine the effect of miR-223-3p on odontoblast differentiation, miR-223-3p was overexpressed in human dental pulp stem cells (DPSCs), which were cultured in mineralizing induction medium (to induce odontoblast differentiation). To identify the target genes of miR-223-3p, an SABiosciences Human Osteogenesis PCR Array, combined with bioinformatics, was used. Furthermore, a dual-luciferase reporter assay and a small interfering RNA (siRNA) experiment were used to confirm the relationship between miR-223-3p and its target gene. Statistical analysis was performed using the Student's t-test or one-way analysis of variance (anova); P < 0.05 was considered statistically significant.
Seventy-nine miRNAs were significantly differentially expressed (fold change >2.0; P < 0.05) between the two tissues. In particular, miR-223-3p was markedly upregulated in inflamed dental pulp. Overexpression of miR-223-3p in DPSCs significantly increased the protein levels of dentine sialophosphoprotein (DSPP) and dentine matrix protein 1 (DMP-1) (P < 0.05). However, the SMAD family member 3 (SMAD3) protein level was significantly lower than in control DPSCs (P < 0.05). Bioinformatics and the dual-luciferase assay reporter assay indicated that Smad3 was a potential target of miR-223-3p. Knockdown of Smad3 in DPSCs subjected to mineralization induction resulted in detection of DSPP and DMP-1 earlier than in control DPSCs, and it increased the protein level of alkaline phosphatase (ALP), thereby promoting odontoblast differentiation.
miR-223-3p is implicated in the regulation of odontoblast differentiation, which may be involved in the process of pulpitis repair.
分析人牙髓组织中炎症和健康组织之间 miRNA 表达谱的差异，并探讨在牙髓炎症组织中 miR-223-3p 的上调如何调节成牙本质细胞分化和再生。
使用微阵列分析鉴定健康和炎症牙髓组织之间 miRNA 表达模式的差异。使用定量实时 PCR 验证结果。为了确定 miR-223-3p 对成牙本质细胞分化的影响，在牙髓干细胞（DPSCs）中过表达 miR-223-3p，然后在矿化诱导培养基中培养（诱导成牙本质细胞分化）。为了鉴定 miR-223-3p 的靶基因，使用 SABiosciences 人类成骨 PCR 阵列结合生物信息学进行分析。此外，还使用双荧光素酶报告基因检测和小干扰 RNA（siRNA）实验来验证 miR-223-3p 与其靶基因之间的关系。使用 Student's t 检验或单因素方差分析（anova）进行统计分析；P<0.05 被认为具有统计学意义。
两种组织之间有 79 个 miRNA 显著差异表达（倍数变化>2.0；P<0.05）。特别是，miR-223-3p 在炎症性牙髓中明显上调。在 DPSCs 中过表达 miR-223-3p 显著增加了牙本质涎磷蛋白（DSPP）和牙本质基质蛋白 1（DMP-1）的蛋白水平（P<0.05）。然而，SMAD 家族成员 3（SMAD3）蛋白水平明显低于对照组 DPSCs（P<0.05）。生物信息学和双荧光素酶报告基因检测表明，Smad3 是 miR-223-3p 的潜在靶基因。在矿化诱导的 DPSCs 中敲低 Smad3 会比对照组 DPSCs 更早地检测到 DSPP 和 DMP-1，并增加碱性磷酸酶（ALP）的蛋白水平，从而促进成牙本质细胞分化。
miR-223-3p 参与了成牙本质细胞分化的调节，这可能涉及到牙髓炎修复过程。
