Canonical Wnt signaling acts synergistically on BMP9-induced osteo/odontoblastic differentiation of stem cells of dental apical papilla (SCAPs)

Abstract

Dental pulp/dentin regeneration using dental stem cells combined with odontogenic factors may offer great promise to treat and/or prevent premature tooth loss. Here, we investigate if BMP9 and Wnt/β-catenin act synergistically on odontogenic differentiation. Using the immortalized SCAPs (iSCAPs) isolated from mouse apical papilla tissue, we demonstrate that Wnt3A effectively induces early osteogenic marker alkaline phosphatase (ALP) in iSCAPs, which is reduced by β-catenin knockdown. While Wnt3A and BMP9 enhance each other’s ability to induce ALP activity in iSCAPs, silencing β- catenin significantly diminishes BMP9-induced osteo/odontogenic differentiation. Furthermore, silencing β-catenin reduces BMP9-induced expression of osteocalcin and osteopontin and in vitro matrix mineralization of iSCAPs. In vivo stem cell implantation assay reveals that while BMP9-transduced iSCAPs induce robust ectopic bone formation, iSCAPs stimulated with both BMP9 and Wnt3A exhibit more mature and highly mineralized trabecular bone formation. However, knockdown of β-catenin in iSCAPs significantly diminishes BMP9 or BMP9/Wnt3A-induced ectopic bone formation in vivo. Thus, our results strongly suggest that β-catenin may play an important role in BMP9-induced osteo/ondontogenic signaling and that BMP9 and Wnt3A may act synergistically to induce osteo/odontoblastic differentiation of iSCAPs. It’s conceivable that BMP9 and/or Wnt3A may be explored as efficacious biofactors for odontogenic regeneration and tooth engineering.

Pro-Inflammatory Cytokine TNF-α Attenuates BMP9-Induced Osteo/ Odontoblastic Differentiation of the Stem Cells of Dental Apical Papilla (SCAPs)

“…The 293pTP line was used for adenovirus amplification. Both 293pTP and iSCAPs were maintained in complete Dulbecco's Modified Eagle's Medium (DMEM) containing 10% fetal bovine serum (FBS, GIBCO,CA), 100 units of penicillin and 100 mg of streptomycin at 37°C in 5% CO 2 .Recombinant adenoviruses expressing BMP9 and/or GFP were constructed as previously reported [25, 26]. Polybrene (10µg/ml) was used to enhance infection efficiency for all adenoviral infections [28].…”

“…Serial sections of the embedded specimens were deparaffinized and stained with hematoxylin & eosin (H & E). Trichrome staining was also carried out as previously described [25, 26, 29]. …”

“…Stem cell implantation-mediated ectopic bone formation was performed as described [25, 26, 29]. Briefly, the iSCAP cells were infected with AdBMP9 or AdGFP, and treated with TNF-α (10ng/ml) condition for 36h, collected and resuspended in PBS with TNF-α (10ng/ml) for subcutaneous injection (5×10 6 cells/injection) into the flanks of athymic nude (nu/nu) mice (5 animals per group, 4-6 week old, female, acquired from the Laboratory Animal Center, Chongqing, China).…”

“…We have recently established the reversibly immortalized mouse stem cells of apical papilla tissue of mouse lower incisor teeth (iSCAPs), which were shown to retain the ability to differentiate into multiple lineages [25, 26]. We also demonstrated that BMP9 can effectively induce osteo/odontoblastic differentiation of the iSCAPs [25, 26]. However, considering that tissue regeneration may occur under the inflammation phase of periapical periodontitis, the effect of high dose of TNF-α on BMP9-induced osteogenic differentiation of iSCAPs remains unclear.…”

The Role of Pannexin3-Modified Human Dental Pulp-Derived Mesenchymal Stromal Cells in Repairing Rat Cranial Critical-Sized Bone Defects

“…Therefore, it is reasonable to conclude that Wnt/β-catenin signaling plays a positive role in osteogenic differentiation of hDPSCs, but the promotive effect of Panx3 on differentiation of hDPSCs may not be attributed to suppression of the Wnt/β-catenin signaling pathway. This finding is in agreement with a previous study showing that Wnt/β-catenin promotes odonto/ osteogenic differentiation of stem cells of dental apical papilla [29].…”