| Objective:Bone resorption under pressure and bone deposition under strain are the biological basis of orthodontic treatment.Root resorption occurs when bone remodeling is abnormal,which is one of the common complications of orthodontic treatment.The main manifestations are as follows:shorter root in CBCT,root surface depression in X-ray and even teeth losing and falling out at clinic.Root absorption and repairing mostly rely on cementoblasts,which can be activated to be predentin,and finally turns into mature cementum after mineralization.It will be an effective way to promote cementoblast's function in order to repair root absorption during orthodontic tooth movement.Ursolic Acid(UA)is a kind of pentacyclic triterpene compound,which is widely distributed in plants,such as Sage,Oleander,Ligustrum Lucidum,Loquat Leaf and so on.Recently,some studies have focused on the role of UA on bone formation and absorption.Scholar L has verified that UA both inhibits bone mineral density loss and promotes bone formation in bone resorption and ossification.Osteoblasts and cementoblasts have many similarities in biological characteristics,but the role of UA on cementoblasts is still unknown.It is hopeful to provide experimental basis for orthodontic tooth restoration by studying the effects of UA on differentiation of cementoblast cell line OCCM-30.Methods:The osteoblasts in logarithmic growth phase were selected.The cells were cultured in Dulbecco's Modified Eagle medium(DMEM)at various concentrations of UA(0.625,1.25,2.5 ?mol/L)and the cells in control group did not receive any treatment.1.The inhibitory rate was detected by MTT at different time points(24,48 and 72 h).2.Cell differentiation was checked by ALP(alkaline phosphatase)activity assay.3.The mRNA expression of OPN in 24 hours was tested by real-time quantitative polymerase chain reaction(PCR).4.The protein expression levels in three and five days of osteogenic marker,osteopontin(OPN)were tested by Western Blotting.Results:1.Cementoblasts were treated with 0.625,1.25 and 2.5?mol/L UA,compared with control group,the ursolic acid concentrations we chose showed no effect on cell proliferation(P>0.05).2.There was no significant difference in ALP activity between the experimental group and the control group when the cementoblasts were treated with 2.5?mol/L UA for one or three days(P>0.05).After treating cementoblasts with 2.5?mol/L UA for 5 days and 7 days,we can observe that the ALP activity in the experimental group was higher than that in the control group,with statistical difference(P<0.05).3.The results of real-time PCR showed that when the concentration of UA was 0.625 ?mol/L,the OPN mRNA level of the experimental group reached its peak at 12 hours,which was about 6 times of that in the control group(P<0.05).When the concentration of UA was 1.25?mol/L,OPN mRNA level in the experimental group was higher than it was in control group at 3 hours,and reached its peak at 6 hours,which was 3.5 times higher than that in the control group(P<0.05).When the concentration of UA was 2.5 ?mol/L,the OPN mRNA level in the experimental group was higher than it was in control group at 3 hours,and reached its peak at 6 hours,which was about 8 times of that in the control group(P<0.05).4.Western blotting results showed that the expression of OPN protein in cementoblasts increased significantly after 3 and 5 days UA treatment,which is dose-dependent and time-dependent.After 5 days of 2.5?mol/L UA treatment,the expression of OPN protein in the experimental group was 7 times higher than that in the control group(P<0.01).Conclusion:Ursolic acid promotes the differentiation of cementoblast.Appropriate concentrations of UA can promote differentiation of cementoblasts and up-regulate the expression levels of OPN,but shows no effect in proliferation. |
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