Study for preservation of dentin using local gene deliveryof dentin-specific genes

利用牙本质特异性基因的局部基因递送来保存牙本质的研究

基本信息

批准号：
11557143
负责人：
TAKASHIBA Shogo
金额：
$ 5.57万
依托单位：
OKAYAMA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

项目摘要

Genes related to dentin regeneration were isolated by susbtractive hybridization from rat molar dental pulp after dentin cavity preparation. There were 16 induced and 7 reduced genes found after size selection (more than 250 bp). Induced genes included cytochrome c, cathepsin B, 3 EST genes (unknown function), and 1 novel gene. Reduced genes included ribosomal protein, laminin-γ2, type 1 collagen-α2, and 2 novels genes. Although in situ hybridization analysis of these genes was performed in order to elucidate their expression sites, no clear results have not been obtained. However, Northem blot analysis revealed that 1 EST gene showed most different expression signals compared before and after cavity preparation. Full length cDNA of this gene was cloned and nucleotide sequence analysis and homology search were performed. It is 3.8-kb length and has 83% homology to human FIP-2 (Adinovirus 14..7 KDa - interacting protein) in some part. Furthermore, this gene has longer putative coding region than that of FIP-2, and posess coild-coild domain including zinc finger domain and leuicin zipper domain. On the other hand, local gene delivery to dental pulp was performed using plasmid vector and andenovirus-associate vector using reporter gene. No significant difference of transfection efficiency was found between these vectors because cells on only the surface of dental pulp were transfected. Primitive problems of this study are 1) selection of genes for transfection and 2) efficiency of local gene delivery. The latter problem would be solved by modification or development of vectors for high transfection efficiency and repeated use. The former problem would be solved by identification of genes as this study. However, additional problem will arise where each gene should be transfected.

牙本质腔制备后，通过从大鼠摩尔牙髓中的大鼠摩尔牙髓中分离出与牙本质再生有关的基因。选择大小后发现了16个诱导的基因（超过250 bp）。诱导的基因包括细胞色素C，组织蛋白酶B，3个EST基因（未知功能）和1个新基因。减少的基因包括核糖体蛋白，层粘连蛋白-γ2，1型胶原蛋白-α2和2个小说基因。尽管对这些基因进行原位杂交分析是为了阐明其表达位点，但尚未获得明确的结果。然而，北部印迹分析表明，1个基因在腔内制备前后显示出最不同的表达信号。克隆该基因的全长cDNA，并进行了核肽序列分析和同源性搜索。它的长度为3.8-kb，与人FIP-2（adinovirus 14..7 kDa-相互作用蛋白）具有83％的同源性。此外，该基因的假定编码区比FIP-2的编码区更长，以及包括锌指域和Leuicin Zipper结构域在内的Posess Coil-Coild域。另一方面，使用质粒载体和Andenovirus-sosocotiate载体使用报告基因进行局部基因递送到牙髓。这些载体之间没有发现转化效率的显着差异，因为仅翻译了牙髓表面上的细胞。这项研究的原始问题是1）转化基因的选择和2）局部基因递送的效率。以后的问题将通过修改或开发向量来解决高转化效率和重复使用。以前的问题将通过将基因鉴定为本研究来解决。但是，将会出现每个基因应翻译的其他问题。