The Expression And Its Role Of BTBD7 Gene On The Molar Root Development

Tooth development is a continuous and complex process, tooth germ is the ideal model of tooth development. Tooth development including crown development, root development and periodontal tissue development and root development is an indispensable part of tooth development. At present, the research on root development focused on the initial stages of tooth root development, the mid-root development especially the formation of furcation is very little. The Hertwig’s Epithelial Root Sheath is generally regarded as the morphological hallmark root development started. And the developing apical complex is a whole functional complex, its organization structure and development of teeth are inseparable. Study found Btbd7 gene expressed protein is very important to the development of critical organs such as lung and salivary glands. Its main function is to contain cadherin, thereby disperse the cells form a branch. By operating the researchers found that if make "Btbd7" genes unable to play a role, so the number of the salivary gland branch structure will be reduced to less than half the normal number. E-cadherin is a kind of calcium dependent transmembrane glycoprotein, important molecules for epithelial cell adhesion, mediating cell recognition and connection, involved in cell differentiation and organ formation, inhibit cell migration. Btbd7 proteins control the salivary glands and epithelial cells of the lung growth branch, branch of the cells and organs to form characteristic structure needed to "crack". By inhibiting adhesion between cell and release the cells, and make them can according to the branch in mammalian organs grow the necessary direction of migration.So we speculate that in the process of root development, whether there is any BTBD7 gene expression, if the expression, it influence on the growth of root number how? There are no reports of on this subject. In order to determine Btbd7 genes have expression in the process of tooth development, if there is expression, the main expression in what place, we used immunohistochemical method to detect the first Btbd7 gene expression in complex root end position. Make sure its in the epithelial root sheath area have clearly expressed, and molar teeth in the formation of root furcation express more obvious. In order to determine the further effect, we extract the eight days of birth were epithelial root sheath cells, by RT-PCR and Western blot detection incisor and molar epithelial root sheath cells Btbd7 gene expression, in order to make clear Btbd7 genes with multiple root tooth formation correlation.Objective:Research on single tooth and tooth tooth development process Btbd7 gene expression changes in epithelial root sheath, to observe Btbd7 gene in the process of tooth development more tooth root formation, the influence of its role in the process.Methods:1. Select 5,8,11,15,19 days of Wistar rat pups, take off the neck to death after anatomy of the upper and lower jaws, take its incisive and first molar tooth germ. Made paraffin section, HE staining, observed in different periods of root development.Immunohistochemical detection of incisor and odontoprisis Btbd7 gene expression of space and time in the embryo and differences.2. Pick up 8 days 3 rat pups, only to take off the neck to death after stripping out from top to bottom in front teeth and jaw of first permanent molar tooth germ. From the tooth crown hard edge cut off part of the neck, I collagenase digestion and trypsin digestion, primary cell culture. Then use differential digestion method for purification of epithelial root sheath cells. Finally using the CK14 Vimentin and cell identification.3. By RT-PCR method for detecting BTBD7 gene in molars and incisors epithelial root sheath cells mRNA expression changes.4. Using Western blot method to detect BTBD7 gene in molars and incisors epithelial root sheath cells protein expression changesResults:1. HE staining showed that the first five days after the pups were born, molars development nearing completion, Cusp section begins calcification, neck ring structure formed in the tooth neck region, but this time there is no obvious epithelial root sheath structure, root development has not yet started; After the pups born on the 8th day, molars development is substantially complete, the root begins to develop, inner enamel epithelium and outer enamel epithelium hyperplasia in the collar, visible structure of the epithelial root sheath; After the pups born in the first 11 days, molar root zone HERS continue to grow, forming epithelial barrier, and partly broken; 15 days after the pups were born, molars developed, root development about two-thirds complete, visible bifurcation structure; After the pups born in the first 19 days, molar root form the branch and continue to grow. And in this phase, incisor crown development is substantially complete, but does not continue to develop the root, in contrast, it gradually formed enamel. Immunohistochemical staining showed that in the first five days after the pups born, Btbd7 gene has no obvious expression at the molar collar; After the pups born in the first 8 days, Btbd7 gene mainly expressed in epithelial root sheath region, in incisors the Btbd7 gene in the epithelial root sheath was weakly positive, in molar was strongly expressed; The first 11 days after the pups were born, Btbd7 gene mainly expressed in molar fracture epithelial root sheath cells, and the expression was not significant; After the pups born in the first 15 days, the Btbd7 gene expression in epithelial root sheath cells molar pups was not significant. After the pups born in the first 19 days, Btbd7 gene has no obvious expression, only has a small amount of expression in the epithelial root sheath rupture.2. Epithelial root sheath cells grew well, gather round or polygonal cells in flake, close connection between cells, the pebbles sample form.CK14 positive, tan can be seen in the cell.3. RT-PCR results showed that the molar Btbd7 gene mRNA expression in epithelial root sheath cells than incisor Btbd7 gene mRNA expression in epithelial root sheath cells significantly higher 。4. Western blot the results showed that the molar Btbd7 gene protein expression in epithelial root sheath cells obviously than incisor Btbd7 epithelial root sheath cells gene protein expression significantly betterConclusion:Btbd7 genes in molar root development process involved in the formation of root bifurcation, and form more correlated with root tooth root development.