MiR-135b调控BMPs/Smads信号通路在牙髓细胞成牙本质向分化中的作用及机制研究
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方法：采用集成的比较性miRNA微阵列分析来确定hDPC的成牙本质细胞样分化中的差异miRNA表达。通过定量实时逆转录聚合酶链反应(qRT-PCR)和原位杂交(ISH)测量microRNA-135b(miR-135b)的丰度。生物信息学分析与萤光素酶分析相结合，用于鉴定与miR-135b相互作用的靶标。进行了miR-135b的过表达以研究hDPC在成牙本质细胞样分化中的作用和机制。通过单因素方差分析(方差分析)或学生t检验进行统计分析。
结果：鉴定了36种在hDPC的成牙本质细胞样分化中差异表达的microRNA。在hDPCs分化过程中,MiR-135b表达明显下调(P<0.05)。此外,miR-135b能够与Smad5和Smad4的3'-UTR结合并抑制这两个基因的表达(P<0.05)。此外,miR-135b的过表达抑制了hDPC的成牙本质细胞样分化，并减弱了Smad5和Smad4的表达(P<0.05)。
结论：这些观察结果表明miR-135b在介导成牙本质细胞样hDPC分化中具有潜在作用，并且抑制miR-135b可能是促进牙本质组织工程的有前途的治疗方法。
Methodology:Integrated comparative miRNA microarray profiling was used to determine the differential miRNAs expression in odontoblast-like differentiation of hDPCs.The abundance of microRNA-135b(miR-135b)was measured by quantitative real-time reverse transcriptase polymerase chain reaction(qRT-PCR)and in situ hybridization(ISH).Bioinformatic analyses combined with luciferase assays were utilized to identify the targets interacting with miR-135b.Overexpression of miR-135b was performed to investigate the role and mechanism in odontoblast-like differentiation of hDPCs.Statistical analysis was performed by one-way analysis of variance(anova)or Student's t-test.
Results:Thirty-six differentially expressed microRNAs in odontoblast-like differentiation of hDPCs were identified.MiR-135b expression was significantly downregulated during hDPCs differentiation(P<0.05).In addition,miR-135b was able to bind to the 3'-UTR of the Smad5 and Smad4 and repressed these two genes expression(P<0.05).Furthermore,overexpression of miR-135b suppressed odontoblast-like differentiation of hDPCs and attenuated the expression of Smad5 and Smad4(P<0.05).
Conclusions:These observations indicated a potential role of miR-135b in mediating odontoblast-like differentiation of hDPCs and inhibition of miR-135b might be a promising therapeutic way to facilitate dentine tissue engineering.