Generation Of TAK1 Transgenic Mouse Models And Related Studies

AIM:Establish constitutively active TAK1 (caTAK1) and dominant negative TAK1 (dnTAK1) transgenic mice, and study the effects of TAK1 overexpression on tooth enamel forms using these transgenic animal models. To further search the target gene of TAK1 during enamel development, we detect the relative expression level of enamel matrix protein in TAK1 transgenic mice. We study the effects of TAK1 on enamel development from histological level and mRNA expression level, thereby laying the foundation for the study of the mechanism of TAK1 in enamel formation. METHODS:Full length DNA of Amelx promoter and TAK1 were cloned. The human caTAK1 and dnTAK1 gene were constructed according to the theory of deletion mutation. The transgenic plasmids were constructed by inserting the caTAKl or dnTAKl gene in the downstream of Amelx promoter. The transgenic mice were produced by microinjection method, then the genotype was detected by PCR and the expression of caTAK1 or dnTAKl were detected by RT-PCR. The mineralization and morphology of enamel were observed by stereomicroscope, Micro X-ray and Micro CT. Finally, we detect Amelx, Ambn, Mmp-20, Klk4 and Alp gene mRNA expression changes between FVB wild-type and TAK1 transgenic mice using real-time quantitative PCR. RESULTS:The transgenic FVB mice of caTAKl or dnTAKl gene were established. The expression of human TAK1 gene in mouse jaw of TAKl transgenic mice was specific. Stereo microscope analysis showed that the caTAK1 transgene mice have poor mineralization of enamel while the enamel of the dnTAK1 transgene mice was similar with FVB mice. In the first month after birth, there was no significant difference in dental enamel morphology between caTAKl or dnTAKl transgenic mice and wild-type FVB mice. While sixth month after birth, the wear degree in tooth of caTAK1 transgenic mice was very serious compared with wild-type mice. Micro X-ray and Micro CT analysis showed that the TAK1 transgenic mice at six months had have severe wear in molar occlusal surface compared the FVB wild-type mice. Real-time PCR data showed that TAK1 upregulates the expression of enamel matrix protein such as Amelx, Ambn, Mmp-20, Klk4 and Alp gene expression level in caTAK1 transgenic mice, and significantly increase Amelx and Ambn expression level. CONCLUSION:TAK1 significantly upregulates the expression level of Amelx and Ambn, which leads to the increase of enamel matrix protein, thereby affecting the mineralization of enamel and resulting in a low of enamel mineralization, easier to be wear.