To analyze the initial displacement of the upper central incisor and stress distribution of periodontal ligament under different torque values of upper incisors under the action of a four-curved auxiliary arch to provide a reliable basis for the safety of clinical application of four-curved auxiliary arches.

A three-dimensional finite element model for torque control of upper anterior teeth with a homemade quadrilateral auxiliary arch was established. Four different states were analyzed: molar ligation without extraction space (group A), microimplant ligation without extraction space (group B), molar recovery with extraction space closure (group C) (the adductive traction force was set at 115 g) and microimplant recovery with extraction space closure (group D) (the adductive traction force was set at 115 g). When four types of torque (0.5 N, 1.0 N, 1.5 N, and 2.0 N) were applied. The initial displacement of upper central incisors and the stress distribution of periodontal ligament in 16 groups (A1-A4, B1-B4, C1-C4, D1-D4) were observed.

Under different conditions, as the strength of the four-curve auxiliary arch increases, the maxillary anterior teeth has crown labial inclination and a root lingual inclination. The displacement of the incisor tip increases with the increase in the loading force of the torque auxiliary arch, and the displacement of the incisor root apex increases as the force increases. The difference in incisor-apex displacement distance in A1-A4, B1-B4, C1-C4, D2 and D4 groups increased as the torque force increases, while the difference between the D3 group and D1 and D2 groups decreased slightly. The stress of the cervical periodontal ligament of the upper central incisor did not exceed the stress of the periodontal ligament in the following groups: A1, A2, B1, B2, B3, C1, C2, D1, and D2. The stress of the lip side of the upper central incisor did exceed the stress of the periodontal ligament in the following groups: A3, A4, B4, C3, C4, D3, and D4. In other words, when using the four-curved auxiliary arch as an implant anchorage, the force applied in the absence of extraction space should not exceed 1.5 N, and the force applied in the adduction of extraction space should not exceed 1.0 N. When using the nonimplant anchorage, the force applied in the absence of extraction space and the adduction of extraction space should not exceed 1.0 N. In addition, the range of force should not exceed the maximum stress of the periodontal ligament in the cervical region such that the effective and safe torque movement can be achieved. Under other stress conditions, the stress of the labial and cervical periodontal ligament of the upper central incisor exceeded the stress value (2.6 × 10 ^{- 2} MPa). The stress value of periodontal ligament was 2.6 × 10 ^{- 2} MPa in all groups.

A four-curved auxiliary arch has a significant effect on the upper anterior teeth, and the use of microimplants can better control root movement such that the crown of upper central incisors cannot be excessively lip inclined.

To investigate the effect of melanoma associated antigen D1 (Mage-D1) on mouse femoral bone mass and mineralization ability of mouse bone marrow mesenchymal cells (BMSCs) and its potential molecular mechanism.

Female Mage-D1 gene knockout heterozygous mice and male wild-type (WT) mice were subjected as parent mice to breed Mage-D1 gene knockout homozygous (Mage-D1 KO) mice. PCR and agarose gel electrophoresis were used to identify male Mage-D1 knockout (Mage-D1 KO) mice and littermate male wild-type (WT) mice. Micro-CT scanning was performed to observe mouse femoral bone mass, and ELISA and chemical assay were employed to detect serum levels of calcium, phosphorus, calcitonin, and parathyroid hormone in mice. After primary cultured BMSCs were identified with flow cytometry, immunofluorescence staining was utilized to detect the expression of Mage-D1 in BMSCs. BMSCs were infected by Mage-D1 silencing lentivirus, and then the cells were divided into negative control group (sh-NC) and silencing group (sh-Mage-D1). Cell scratch assay was conducted to detect the migration ability of BMSCs, and flow cytometry and CCK-8 assay were conducted to detect the cycle change and proliferation ability of BMSCs. After mineralization induction, alkaline phosphatase (ALP) staining and alizarin red staining were performed; RT-qPCR and Western blotting were used to measure the expression levels of ALP, Runx2 and Col1. RT-qPCR was used to detect mineralization-related genes p75NTR and Msx1.

Compared with the WT mice, the femoral cortical bone thickness, cortical bone mineral content, cancellous bone mineral content, trabecular number, and cancellous bone surface density were decreased, and trabecular separation was increased in the Mage-D1 knockout homozygous mice (P<0.05). There were no significant changes in the serum levels of calcium, phosphorus, calcitonin and parathyroid hormone in mice after Mage-D1 knockout. Mage-D1 was expressed in the whole BMSCs and was highly expressed in the nucleus and perinuclear regions. Compared with the sh-NC BMSCs, the sh-Mage-D1 group had decreased proliferation ability (P<0.01), enhanced migration ability (P<0.01), and decreased expression of ALP, Runx2 and Col1 genes (P<0.05) and protein (P<0.01) after mineralization induction, milder ALP and alizarin red stain, and lower expression levels of p75NTR and Msx1.

Mage-D1 knockout can significantly reduce femur bone mass in mice. It can promote the proliferation and inhibit migration of BMSCs, and positively regulate their mineralization in vitro, and the p75NTR-Dlx1/Msx1 signaling axis may be involved in the regulation of bone metabolism by Mage-D1.

To investigate the regulative effect of neurally differentiated EC-cell-derived protein, Necdin, on the odontogenic differentiation and mineralization of rat dental germ ectomesenchymal stem cells（EMSCs）.

Immunofluorescence staining was used to detect the expression and distribution of Necdin in E13.5d and E19.5d rat dental germ. EMSCs derived from E19.5d rat dental germ were isolated and cultured, and identified with flow cytometry for cell surface antigens. The EMSCs were stably transfected with lentiviral vector of Necdin silencing. Then, the apoptosis of EMSCs was detected by flow cytometry in the negative control group（sh-NC）and silenced group（sh-Necdin）. The proliferation capacity of EMSCs was detected by CCK-8 assay, and the migration ability of EMSCs was detected by cell scratching assay. After mineralization induction, the regulative effect of Necdin on the odontogenic differentiation and mineralization of EMSCs was observed by alkaline phosphatase staining, alizarin red staining, RT-PCR and Western blotting.

The immunofluorescence results showed that Necdin was strongly expressed and concentrated in the inner enamel epithelium, outer enamel epithelium and in the contact area between epithelium and mesenchyme. After Necdin silencing, the EMSCs showed higher apoptotic rate, enhanced proliferation ability but weaker migration ability when compared with the cells from the sh-NC group（all P<0.01）. What's more, Necdin silencing also resulted in lighter alkaline phosphatase stain and less mineralized nodule formation, and up-regulations of the molecules related to tooth differentiation and mineralization at mRNA and protein levels（P<0.05）in the cells from the sh-Necdin group than those from the sh-NC group.

Necdin is specifically expressed in the future odontogenic differentiation and mineralization regions in rat dental germ. Its silencing promotes the apoptosis and proliferation, but inhibits the migration and the abilities of odontogenesis differentiation and mineralization in EMSCs. Necdin might play an important role in tooth development and mineralization.

To investigate the dynamic expression profile of mdanoma associated antigen D1 (Mage-D1) at different tooth development stages in mice after birth, and explore the effect of Mage-D1 knockout on tooth development in vivo.

C57-BL/6 wild-type suckling mice were grouped according to their age (1, 4, 7, 11 and 15 d after birth, n=3 in each group). Tissue samples were prepared and the developmental dynamics of the first mandibular molars in mice were observed by HE staining. Immunohistochemical staining was used to observe the expression of Mage-D1 as well as the homologous box genes Dlx1 and Msx1 at different stages of postnatal tooth development in mice. The Mage-D1 homozygous knockout mice (KO) and wild-type mice (WT) aged 1 and 3 months (n=6 in each group) were selected for phenotypic observation, and micro-CT scanning for mandible was performed to analyze the data of teeth and mandible.

Morphological observation showed that on postnatal day 1, the first mandibular molar was in the late bell-shaped stage without obvious hard tissue formation; on day 4, dentin and enamel alternatively formed and crown began to develop; on day 7, epithelial root sheaths shaped and root development initiated; on day 11, the enamel and crown dentin were almost completed, with the root about half developed; on day 15, the root of the tooth further developed, reaching more than 2/3, and the cementum was formed. Mage-D1 and the homeobox genes Dlx1 and Msx1 were positively expressed in both ameloblasts and odontoblasts of mice throughout the period from 1 d to 15 d after birth. As compared with WT mice, the body weight of KO mice was increased (P<0.05); though the number of teeth was the same in the 2 groups, the teeth appearance in KO group was slightly different, with incisors less stained and molars more easily worn. Scanning results indicated no significant difference in the density of teeth and alveolar bone.

Mage-D1, Dlx1 and Msx1 may contribute together to the formation of tooth enamel and dentin in mice, and the mice with Mage-D1 gene knockout present obesity signs and altered tooth phenotype.