Study on the mechanism for the periodontal ligament tissue regeneration using human periodontal ligament tissues derived from complex odontoma

正畸牙齿的运动依赖于牙周韧带的主动重塑和再生。为了分析这一机制，我们利用齿瘤PDL，试图找到齿瘤PDL的特异性基因。切除源自前磨牙复合耳瘤的人PDL组织，将其从正畸原因中取出，置于柔性底板上。1个月后，异质细胞汇合，克隆成功。每个无性系在6-8代收集并储存。分析各克隆的细胞形态和生长分化能力。从牙瘤PDL和正常牙PDL的异质原代培养细胞中提取总RNA。为了研究控制牙齿发育的Msx-1、Msx-2在牙瘤PDL和正常牙齿PDL中的表达差异，我们利用特异性引物将总RNA逆转录为cDNA，然后进行定量real-tune PCR。结果，所有rna都表达Msx-1和Msx-2 mrna，尽管表达水平因细胞而异。细胞形态也各不相同。并采用另一复杂牙瘤的牙周韧带组织，分析了Msx-1和Msx-2的表达趋势。这些结果表明，牙瘤的形成过程至少涉及Msx-1和Msx-2，但也涉及其他转录因子。我们正在分析cDNA阵列法和差异显示PCR法，以确定在耳瘤PDL中特异性表达的基因。我们还在准备克隆细胞的永生化和克隆特异性基因的鉴定。

Orthodontic tooth movement depends on the active periodontal ligament remodeling and regeneration. To analyze this mechanism, we utilized odontoma PDL and tried to find the genes specific in odontoma PDL. Human PDL tissue derived from a premolar complex odotoma was excised, which was extracted from orthodontic reasons, and placed onto flexible bottom plate. One month later, the heterogeneous cells reached confluence and they were cloned. Each clone was collected at 6-8 passages and stocked. Cell morphology and growth and differentiation ability of each clone is analyzing.Total RNA was extracted from the heterogeneous primary culture cells from the odontoma PDL and from normal tooth PDL. To investigate the difference in the expression of Msx-1, Msx-2, which has been reported to control the tooth development, in odontoma PDL and normal tooth PDL, total RNA was reverse transcribed into cDNA, followed by quantitative real-tune PCR, using specific primers. As a result, all RNAs expressed the Msx-1 and Msx-2 mRNAs, though the expression level varied with the cells. The morphology of the cells also differed from each other.Furthermore, another periodontal ligament tissues from another complex odontoma were used and analyzed the expression of Msx-1 and Msx-2 and the same tendency was observed. These results suggested that the process for the formation of odontoma involves at least Msx-1 and Msx-2, but also other transcriptional factors. We are analyzing cDNA array method and differential display PCR method, to identity the genes that is expressed specific in odotoma PDL. We are also preparing to immortalize the cloned cells, and to identify the clone specific genes.