Résumé
Objective To investigate the improvement effect of Jiegu Qili Capsules on the fracture healing of radius in rats by activating the bone morphogenetic protein(BMP)/Smad signaling pathway.Methods(1)A rat model of radius fracture was constructed,and the serum levels of alkaline phosphatase(ALP),calcium(Ca)and phosphorus were detected,and the pathological changes in the fracture gap was observed.(2)Human osteosarcoma SaOS-2 cells were used to measure ALP activity and mineralization level.The quantitative real-time polymerase chain reactionn(qRT-PCR)method was used to detect the cellular osteogenesis-related genes ALP,collagen I(COL-I),ornithine transcarbamylase(OTC),Osterix,osteoblastin(OPN),Runt-related transcription factor 2(RUNX2)and BMP2.The expression of key proteins in BMP/Smad signal pathway was detected by Western Blot.Results Jiegu Qili Capsules effectively promoted fracture healing of radius in rats,enhance ALP activity,increase Ca and P levels Jiegu Qili Capsules stimulate the formation of mineralized nodules in SaOS-2 cells in rats.,and promoted the expression levels of COL-I,OTC,Osterix,BMP2 and OPN in SaOS-2 cells in a dose-dependent manner.Jiegu Qili Capsules up-regulated the levels of Smad1/5 phosphorylation of the BMP/Smad signaling pathway in SaOS-2 cells,as well as the levels of BMP2 and RUNX2.Noggin,an inhibitor of the BMP/Smad signaling pathway,inhibited osteogenic differentiation of SaOS-2 cells induced by Jiegu Qili Capsules.Conclusion Jiegu Qili Capsules can promote fracture healing by activating the BMP/Smad signaling pathway to increase the expression of osteogenesis-related genes.
Résumé
BACKGROUND:Congenital clubfoot mainly manifests as abnormal bone itself and abnormal cartilage development.The bone morphogenetic protein(BMP)/drosophila mothers against decapentaplegic protein(Smad)signaling pathway can direct the development of bone and cartilage during embryonic period,but its role in the field of clubfoot etiology has not been confirmed in animal experiments. OBJECTIVE:To explore the mechanism by which the BMP/Smad signaling pathway is involved in the regulation of foot and ankle chondroplasia in a rat congenital clubfoot model. METHODS:Sprague-Dawley rats at 10 days of gestation with the same growth condition were randomly divided into experimental and control groups.The experimental group was intragastrically given 135 mg/kg retinoic acid to make the clubfoot model,while the control group was given the same amount of vegetable oil.The fetal rats were taken out after 21 days of gestation by cesarean section.In the experimental group,the 27 of 41 fetal rats had clubfoot,with a deformity rate of 65.9%;in the control group,no clubfoot was observed in all the 36 fetuses.The ankles tissues of the rats were collected for hematoxylin-eosin staining.Western blot assay,RT-qPCR and immunohistochemistry were performed to detect the expression levels of Smad5 and P-Smad5,the core proteins of the BMP/Smad signaling pathway,as well as SP7 and Sox9,the downstream proteins of the BMP/Smad signaling pathway. RESULTS AND CONCLUSION:Compared with the control group,hematoxylin-eosin staining showed that the cartilage matrix in the foot and ankle tissues increased and the gap between the bones increased in the experimental group.Immunohistochemical findings showed that the expression levels of Smad5 and SP7 decreased in the experimental group,while the mRNA expression of Sox9 increased.RT-qPCR results showed that the mRNA expression of Smad5 and SP7 decreased,while the mRNA expression of Sox9 increased in the foot and ankle tissues of rats in the experimental group.Western blot results showed that P-Smad5/Smad5 expression and SP7 expression were decreased and Sox9 expression was increased in the ankle of rats in the experimental group.To conclude,the occurrence of cartilage abnormalities in the foot and ankle of the rat model of congenital clubfoot is associated with impaired transmission of the BMP/Smad signaling pathway.