WNT10A (Wingless-type MMTV integration site family, member 10A) plays a crucial role in tooth development, and patients with biallelic WNT10A mutation and mice lacking Wnt10a show taurodontism. However, whether epithelial or mesenchymal WNT10A controls the initiation of the root furcation formation remains unclear, and the functional significance of WNT10A in regulating root morphogenesis has not been clarified. Here, we investigated how Wnt10a affects tooth root development by generating different tissue-specific Wnt10a conditional knockout mice. Wnt10a knockout in the whole tissue ( EIIa-Cre;Wnt10aflox/flox) and in dental epithelium ( K14-Cre;Wnt10aflox/flox) led to an absence of or apically located root furcation in molars of mice, a phenotype that resembled taurodontism. An RNAscope analysis showed that the dynamic epithelial and mesenchymal Wnt10a expression pattern occurred during root development. Immunofluorescent staining of E-cadherin and EdU revealed decreased epithelial cell proliferation at the cervical region of the molar in K14-Cre;Wnt10aflox/flox mice at postnatal day 0 (PN0), just before the initiation of root morphogenesis. Interestingly, we found increased pulpal mesenchymal cell proliferation in the presumptive root furcating region of the molar in K14-Cre;Wnt10aflox/flox mice at PN4 and PN7. RNA-seq indicated that among the Wnt ligands with high endogenous expression levels in molars, Wnt4 was increased after epithelial knockout of Wnt10a. The RNAscope assay confirmed that the expression of Wnt4 and Axin2 in the dental papilla of the presumptive root furcating region, where dental pulp overgrowth occurred, was increased in K14-Cre;Wnt10aflox/flox molars. Furthermore, after suppression of the elevated Wnt4 level in K14-Cre;Wnt10aflox/flox molars by Wnt4 shRNA adenovirus and kidney capsule grafts, the root furcation defect was partially rescued. Taken together, our study provides the first in vivo evidence that epithelial Wnt10a guides root furcation formation and plays a crucial role in controlling the organized proliferation of adjacent mesenchymal cells by regulating proper Wnt4 expression during root furcation morphogenesis.
Background/purpose Bioactive glass (BG), one type of bioceramics, shows similar or better characteristics to calcium silicate which has been regarded as a promising root filling material in endodontics. This study aimed to develop a novel BG-based root canal sealer for endodontics. Materials and methods The novel BG-based root canal sealer was composed of phytic acid derived bioactive calcium phosphosilicate glass named PSC mixed with zirconium oxide (ZrO 2 ) as powder, and phosphate solution (PS) dissolved with sodium alginate (SA) named PS-SA as liquid. Moreover, the physicochemical properties, mineralization, sealing ability and biocompatibility of the novel BG-based root canal sealer were evaluated. Results This study developed a novel BG-based sealer named BGS-SA-Zr which contained the powder of PSC and ZrO 2 and the liquid of PS-SA. Results indicated that the flow, film thickness and radiopacity of BGS-SA-Zr conformed to ISO 6876:2012. The setting time and solubility of BGS-SA-Zr were 53.7 ± 1.5 min and 21.46 ± 0.54%, respectively. The pH value of the simulated body fluid (SBF) immersed with BGS-SA-Zr raised slightly up to 7.70. The CCK-8 assay indicated that BGS-SA-Zr had no cytotoxic effects on MG-63 cells. After immersion in SBF for 4 weeks, dense hydroxyapatite crystals were observed on the surface of BGS-SA-Zr. Furthermore, there was no difference in the sealing ability between BGS-SA-Zr and the bioceramic sealer iRoot SP whether setting at 1 day or immersed in SBF for 4 weeks ( P > 0.05). Conclusion Our results suggest that the novel BG-based sealer may be a promising sealer for endodontic treatment.
Early angiogenesis is important to facilitate biomaterials’ osteogenic effects and avoid the bone regeneration failure for large-sized bone defects. Bioactive glasses (BG) have been widely utilized as a biomaterial for bone repair. However, the early angiogenesis of BG may be inadequate. In this study, we explored the effects of photobiomodulation (PBM) combined with BG on early angiogenesis to solve this bottleneck problem of insufficient early angiogenesis. In vitro, human umbilical vein endothelial cells (HUVECs) were cultured with BG extracts and treated with PBM using 1 J/cm2. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, real-time reverse transcription-polymerase chain reaction (real-time RT-PCR) and tubule formation assay were utilized to detect HUVECs’ proliferation, vascular growth factor genes expression and tubules formation. In vivo, bone defects at the femoral metaphysis in Sprague-Dawley rats were treated with BG particulates and PBM at 120 J/cm2. Immunohistochemical staining was applied to observe the vascular-like structure formation. In vitro results showed that PBM combined with BG significantly promoted HUVECs’ proliferation, genes expression and mature tubules formation. On days 2, 4 and 7, the VEGF gene expression in BG+PBM group was 2.70-, 2.59- and 3.05-fold higher than control (P<0.05), and higher than PBM and BG groups (P<0.05). On days 4 and 7, the bFGF gene expression in BG+PBM group was 2.42- and 1.82-fold higher than control (P<0.05), and also higher than PBM and BG groups (P<0.05). Tube formation assay showed that mature tubules formed in BG+PBM and PBM groups after 4 hours. The tubules number in BG+PBM group was significantly higher than other groups (P<0.05). In vivo results further confirmed that PBM induced early angiogenesis. More vascular-like structures were observed in BG+PBM and PBM groups 2-week post surgery. In conclusion, with the optimum PBM fluence and BG concentration, PBM combined with BG exerted additive effects on enhancing early angiogenesis.
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