The effect of growth factors and extracellular matrices on periodontal ligament cell differentiation during periodontal tissue regeneration.

探讨生长因子和细胞外基质在牙周再生治疗中的作用。1.首次开展了碱性成纤维细胞生长因子的研究，并建立了抗人碱性成纤维细胞生长因子的单抗。其中一种抗体BF-24用于免疫组织化学分析，结果显示碱性成纤维细胞生长因子在牙周膜和牙周上皮细胞中密集分布。为探讨碱性成纤维细胞生长因子在组织再生和创面愈合中的作用，制备了犬人工三壁骨内缺损区。BF-24在术后1周瞬时检测到碱性成纤维细胞生长因子的显著表达。2、在碱性成纤维细胞生长因子(BFGFf)存在下建立了小鼠单克隆性PDL细胞MPDL-22。MPDL-22在bFGF2存在下可长期增殖，但一旦bFGF2被剥夺，则会导致细胞成熟并开始钙化。骨基质蛋白如骨钙素、骨桥蛋白、牙骨质蛋白、Ⅲ型胶原和韧带蛋白、XII型胶原在MPDL-22成熟后表达增加，提示这些蛋白的表达可能是牙周韧带组织分化的标志。3、在人HPDL中，碱性成纤维细胞生长因子刺激后，层粘连蛋白mRNA量增加，而I型胶原mRNA量减少，纤维连接蛋白和II型胶原mRNA量无明显变化。4、对牙骨质酸性提取物进行分级，以鉴定未知的韧带生长因子。经过几步生化纯化，mW10-20 kDa蛋白具有促HPDL有丝分裂活性，多肽片段序列分析表明其与胰岛素样生长因子-I部分同源。

The roles of growth factors and extracellular matrices were investigated for the improvement of periodntal regeneration therapy. 1., We first forcused this study on basic fibroblast growth factor (bFGF) and established the anti-human bFGF monoclonal antibodies (mAbs). One of those Abs, BF-24 was utilyzed for immunohistochemical analysisand showed the dense distribution of bFGF especially in periodontal ligament (PDL) and epithelium. In order to examine the involvement of bFGF during the tissue regeneration ana wound healing, artificial three wall intrabony defect was prepared in beagle dogs. BF-24 was able to detect the prominent expression of bFGF one week after surgery transiently. 2, Mouse monoclonal PDL cell, MPDL-22 was established in the presence of bFGF.MPDL-22 revealed long term proliferation in the presence of bFGF,however, once bFGF was deprived, it lead the to maturation and started calcification. The mRNA of bone matix protein, such as osteocalcin, osteopontin, and cementum protein, type III collagen and ligamentum protein, typeXII collagen was increased after the maturation of MPDL-22, suggesting that the expression of these protein may be a differentiation marker for perodontal ligament tissue. 3, In the case of human HPDL,the stimulation by bFGF increased the expression of laminin mRNA.In contrast, the amount of type I collagen mRNA was decreased and no significant alteration was observed in fibronectin and type II collagen mRNA expression. 4, The acid extract of cementum was fractionated in order to identify unknown ligament growth factor. Several step of biochemical purification resulted that m.w.10-20kDa protein contained the mitogenic activity for HPDL and the sequence analysis of peptide fragment revealed its partial homology to Insulin like growth factor-I.