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目的筛选人牙髓细胞(human dental pulp cells,HDPCs)在未分化状态及在矿化诱导液中差异表达的miRNA并预测其靶基因。方法体外分离培养临床就诊患者来源的人牙髓细胞并鉴定后,分别在普通培养基和矿化诱导培养基中培养,用矿化诱导液诱导细胞向成牙本质细胞方向分化,对细胞结节形成情况进行观察并用Vonkossa染色确定HDPCs分化情况。取培养21 d的细胞用miRNA芯片检测差异表达的miRNA,并通过miRGen数据库预测其靶基因,通过GO和KEGG Pathway进行靶基因功能富积分析。结果矿化诱导的HDPCs较未诱导的HDPCs,有72种miRNA发生1.5倍以上表达上调,61种miRNA发生1.5倍以上表达下调。通过miRNA靶标预测工具分析发现,带有靶标基因的miRNA有35个,35个miRNA的靶标基因总和1 327个,miRNA和靶标基因关系对总共2158个。结论成功筛选出HDPCs miRNAs表达谱,预测到部分靶基因。
Objective To screen miRNAs differentially expressed in undifferentiated human dental pulp cells (HDPCs) and their mineralizing inducing fluids and to predict their target genes. Methods Human dental pulp cells derived from clinical patients were isolated and cultured in vitro. The cells were cultured in normal medium and mineralization induction medium respectively. The mineralized inducing medium was used to induce cells to differentiate into odontoblasts. Formation was observed and Vonkossa staining was used to determine HDPCs differentiation. Differentially expressed miRNAs were detected by miRNA microarray for 21 days and their target genes were predicted by miRGen database. The gene enrichment analysis of target genes was performed by GO and KEGG Pathway. Results Compared with non-induced HDPCs, 72 miRNAs were upregulated in mineralization-induced HDPCs more than 1.5-fold and 61 miRNAs were more than 1.5-fold down-regulated. According to the miRNA target prediction tool, there were 35 miRNAs with target genes, 1 327 miRNA target genes and 3558 miRNAs with a total of 2158 miRNA and target genes. Conclusion The expression profiles of miRNAs in HDPCs were successfully screened and some target genes were predicted.