The Expression Pattern Of Runx2 And Osx In The Process Of Reparative Dentine Formation

Backgrounds and objectives:Many factors,including caries and trauma,cancauses dental pulp inflammatory injury,at the same time can cause dentition defect.Nevertheless,when the teeth suffer from caries or the mild stimulus,the dental pulp cell will produce a series of changes,including migration,proliferation and differentiate into odontoblasts,then the odontoblasts will mineralize into mature,and then form reparative dentin.Obvious the more we understand the mechanism of tooth repairation after damage,the more we have a chance to preserve tooth.Both Runx2(Runt-related transcription factor 2)and Osx(Osterix)are the key transcription factors of osteogenesis.Recent study showed that the development of tooth formation also requires the two transcription factors.DSPP(Dentin Sialoph-osphoprotein),which transcription is regulated by Runx2,is a key mark protein in odontoblasts maturing.Many study about osteogenesis found that Osx is the downstream of Runx2.Osx gene promoter regions exist some specific binding sites for Runx2 to combine.Runx2 regulate the expression of Osx by the combination of the specific binding sites.However,we have little know about the expression pattern of Runx2 and Osx in reparative dentine formation.While Osx can regulate the expression of DSPPin tooth development.Therefore,we establish the rat reparative dentine formation model with different time after injury,to investigate the expression patterns of Runx2 and Osx and the relationship of Runx2?Osx and reparative dentine.Methods:1.We establish the rat reparative dentine formation model.60 rats about 200 g body quality were randomly divided into normal control group and injury model group(n = 10).In damage model group,they pulps was open using fast drilling machine ball in bilateral mandibular first molar regularly in rats.Don't do any processing after opening.The injury model groups by the exposure time of the medullary cavity 0 h,3 h,12 h,3 d and 7 d.The control group do not open pulp.Then four experimental animals are randomly selected from each group,are perfusion to death.We extract the rat mandibular first molar and the bone around and made into slices.hematoxylin-eosin(H&E)staining method are used to determine whether the reparative dentin model established.2.IF(Immunofluorescence)technique was used to investigate the expression pattern of Runx2?Osx and DSPP protein and m RNA in the progress of reparative dentin formation in the mandibular first molar of rats,The control group and 0h,3h,12 h,3d and 7d groupare stained with IF method separately.Meanwhile we use con focal laser-scanning microscopy to observe the changes of the Runx2?Osx and DSPP protein.The surplus control group and 0h,3h,12 h,3d and 7d group rats were killed and then extract the rat mandibular first molar.We compare the m RNA of Runx2?Osx and DSPP among each group by R-T PCR.Result:1.Dental pulp cells can migrate after the dental pulp injury.There is apparent inflammatory reaction and clearly visible reparative dentine formation in the seventh day at the bottom of the dental pulp injury.2.R-T PCR:The expression of Runx2 m RNA is the highest at 12 h after injury and is significantly higher than those in control group(P<0.01).The expression of Runx2 m RNA in 3d after injury is lower than those in 12 h.Until 7d after injury,the expression of Runx2 m RNA is lower than those in 12 h and 3d after injury(P<0.01),which is significantly higher than those in control group(P<0.01).The expression of Osx and DSPP m RNA has no significant differences with 0h after injury and control group(P>0.05).The expression of Osx and DSPP m RNA quantity increases gradually since 3h after injury,and significantly higher than the control group,the comparison between groups have significant differences(P<0.01);IF: Reparative dentine formation was detected in 7d after injury.The expression of Runx2?Osx and DSPP were Mainly at the bottom of dental pulp injury.The expression of Runx2 was strongest at 3d after injury(P<0.01)and gradually declined,which looked like "parabola".The expression of Osx and DSPP were both showed a trend of increase gradually.The expression of Osx was strongest at 7d after injury(P<0.01).While The expression of DSPP was strongest at 3d after injury(P<0.01).Conclusion:1.The rats after birth can occur stress reaction after the external stimuli with dental pulp tissue.The dental pulp of rats has the ability of forming reparative dentin.2.There is different expression patterns of Runx2,Osx and DSPP in dental pulp after injury in different periods and different parts of the tooth.