Sipert Carla R, Morandini Ana C, Dionísio Thiago J, Trachtenberg Alexander J, Kuo Winston P, Santos Carlos F
Department of Biological Sciences, Bauru School of Dentistry, University of São Paulo.
J Oral Sci. 2014 Jun;56(2):157-64. doi: 10.2334/josnusd.56.157.
MicroRNAs (miRNAs) are small non-coding RNAs showing a tissue-specific expression pattern, and whose function is to suppress protein synthesis. In this study, we hypothesized that expression of miRNAs would differ among fibroblasts from dental pulp (DPF), gingiva (GF) and periodontal ligament (PLF) in vitro. Once established by an explant technique, DPF, GF and PLF were collected for RNA isolation and subjected to a miRNA microarray. Next, cells were stimulated with E. coli lipopolysaccharide (LPS) for 24 h and then collected for RNA isolation. Expression of miR-146a and miR-155 was investigated by qPCR. Microarray screening revealed several miRNAs that showed specifically high expression in at least one of the fibroblast subtypes. These molecules are potentially involved in the regulation of extracellular matrix turnover and production of inflammatory mediators. Microarray analysis showed that both miR-146a and miR-155 were among the miRNAs expressed exclusively in GF. qPCR demonstrated significant upregulation of miR-146a only in GF after LPS stimulation, whereas basal expression of miR-155 was higher in GF than in the other cell subtypes. LPS downregulated the expression of miR-155 only in GF. Our results suggest that the expression and regulation of miR-146a and miR-155 are more pronounced in GF than in DPF and PLF.
微小RNA（miRNA）是一类小的非编码RNA，呈现组织特异性表达模式，其功能是抑制蛋白质合成。在本研究中，我们假设体外培养的牙髓成纤维细胞（DPF）、牙龈成纤维细胞（GF）和牙周膜成纤维细胞（PLF）中miRNA的表达会有所不同。通过组织块培养技术建立细胞系后，收集DPF、GF和PLF用于RNA提取，并进行miRNA微阵列分析。接下来，用大肠杆菌脂多糖（LPS）刺激细胞24小时，然后收集细胞用于RNA提取。通过qPCR研究miR-146a和miR-155的表达。微阵列筛选发现了几种在至少一种成纤维细胞亚型中特异性高表达的miRNA。这些分子可能参与细胞外基质周转和炎症介质产生的调节。微阵列分析表明，miR-146a和miR-155均为仅在GF中表达的miRNA。qPCR显示，LPS刺激后仅GF中的miR-146a显著上调，而miR-155的基础表达在GF中高于其他细胞亚型。LPS仅在GF中下调miR-155的表达。我们的结果表明，miR-146a和miR-155的表达及调控在GF中比在DPF和PLF中更为明显。
