circRNA_079813调控ROR2和ERK1/2-MAPK信号通路对牙髓损伤大鼠成骨分化的影响

Effect of circRNA_079813 promoting osteogenic differentiation via regurating ROR2 and ERK1/2-MAPK signaling pathway in dental pulp injury rats

目的:探讨circRNA_079813对牙髓损伤模型大鼠成骨分化的作用及其机制。方法:将70只SD大鼠随机分为7组,即假手术组、牙髓损伤组、pcDNA3.1空载体(pcDNA-null)+牙髓损伤组、过表达circRNA_079813(pcDNA-circ)+牙髓损伤组、pcDNA-circ+siRNA-NC+牙髓损伤组、pcDNA-circ+siRNA-ROR2+牙髓损伤组、pcDNA-circ+PD98059(ERK1/2抑制剂)+牙髓损伤组,每组10只。术后3 d采用苏木精—伊红(HE)染色观察牙髓组织病理变化。检测各组碱性磷酸酶(ALP)活性,RTqPCR法检测circRNA_079813表达,western blotting法检测ROR2、磷酸化(p-)ERK1/2、骨涎蛋白(BSP)、骨钙蛋白(OCN)、ALP、牙本质涎磷蛋白(DSPP)和牙本质基质蛋白-1(DMP-1)蛋白表达。结果:与假手术组比较,牙髓损伤组牙髓组织有明显炎症浸润和损伤特征,circRNA_079813表达下调(P<0.05)。与pcDNA-null+牙髓损伤组比较,pcDNA-circ+牙髓损伤组circRNA_079813、ROR2、p-ERK1/2、BSP、ALP、OCN、DSPP、DMP-1表达水平及ALP活性均升高(均P<0.05)。沉默ROR2表达后,与pcDNA-circ+siRNA-NC+牙髓损伤组比较,pcDNA-circ+siRNA-ROR2+牙髓损伤组ROR2、p-ERK1/2、BSP、ALP、OCN、DSPP、DMP-1表达及ALP活性均下降(均P<0.05)。与pcDNA-circ+牙髓损伤组比较,pcDNA-circ+PD98059+牙髓损伤组pERK1/2、ALP、BSP、OCN、DSPP、DMP-1表达及ALP活性均下降(均P<0.05)。结论:circRNA_079813通过促进ROR2表达激活牙髓损伤大鼠ERK1/2-MAPK信号通路促进成骨分化。

Objective:To explore the role and mechanism of circular RNA(circRNA)079813 in osteogenic differentiation in a dental pulp injury model in rats.Methods:A total of 70 SD rats were randomly divided into 7 groups:sham operation group,dental pulp injury group,pcDNA3.1 empty vector(pcDNA-null)+dental pulp injury group,overexpression of circRNA_079813(pcDNA-circ)+dental pulp injury group,pcDNA-circ+siRNANC+dental pulp injury group,pcDNA-circ+siRNA-ROR2+dental pulp injury group,and pcDNA-circ+PD98059(ERK1/2 inhibitor)+dental pulp injury group,with 10 in each group.Hematoxylin-eosin(HE)staining was used to observe the pathological changes in dental pulp tissue 3 days after surgery.Alkaline phosphatase(ALP)activity was detected in each group,and circRNA_079813 expression was detected by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR).The expression of ROR2,phosphorylated ERK1/2(p-ERK1/2),bone sialoprotein(BSP),osteocalcin(OCN),ALP,dentin sialophosphoprotein(DSPP)and dentin matrix protein-1(DMP-1)were detected by western blotting.Results:Compared with the sham operation group,the dental pulp tissue in the dental pulp injury group showed significant inflammatory infiltration and damage characteristics,and the expression of circRNA_079813 was down-regulated(P<0.05).Compared with the pcDNA-null+dental pulp injury group,the expression levels of circRNA_079813,ROR2,p-ERK1/2,BSP,ALP,OCN,DSPP and DMP-1 and the activity of ALP were up-regulated in the pcDNA-circ+dental pulp injury group(all P<0.05).After silencing ROR2 expression,compared with the pcDNA-circ+siRNA-NC+dental pulp injury group,the expression of ROR2,p-ERK1/2,BSP,ALP,OCN,DSPP and DMP-1 and the activity of ALP in the pcDNA-circ+siRNA-ROR2+dental pulp injury group were down-regulated(all P<0.05).Compared with the pcDNA-circ+dental pulp injury group,the expression of p-ERK1/2,ALP,BSP,OCN,DSPP and DMP-1 and the activity of ALP were down-regulated in the pcDNA-circ+PD98059+dental pulp injury group(all P<0.05).Conclusion:CircRNA_079813 promotes osteogenic differentiation in dental pulp injury rats through activation of the ERK1/2-MAPK signaling pathway by promoting ROR2 expression.