Wnt/ß-Catenin Promotes the Osteoblastic Potential of BMP9 Through Down-Regulating Cyp26b1 in Mesenchymal Stem Cells.

BACKGROUND: All-trans retinoic acid (ATRA) promotes the osteogenic differentiation induced by bone morphogenetic protein 9 (BMP9), but the intrinsic relationship between BMP9 and ATRA keeps unknown. Herein, we investigated the effect of Cyp26b1, a critical enzyme of ATRA degradation, on the BMP9-induced osteogenic differentiation in mesenchymal stem cells (MSCs), and unveiled possible mechanism through which BMP9 regulates the expression of Cyp26b1. METHODS: ATRA content was detected with ELISA and HPLC-MS/MS. PCR, Western blot, and histochemical staining were used to assay the osteogenic markers. Fetal limbs culture, cranial defect repair model, and micro-computed tomographic were used to evaluate the quality of bone formation. IP and ChIP assay were used to explore possible mechanism. RESULTS: We found that the protein level of Cyp26b1 was increased with age, whereas the ATRA content decreased. The osteogenic markers induced by BMP9 were increased by inhibiting or silencing Cyp26b1 but reduced by exogenous Cyp26b1. The BMP9-induced bone formation was enhanced by inhibiting Cyp26b1. The cranial defect repair was promoted by BMP9, which was strengthened by silencing Cyp26b1 and reduced by exogenous Cyp26b1. Mechanically, Cyp26b1 was reduced by BMP9, which was enhanced by activating Wnt/ß-catenin, and reduced by inhibiting this pathway. ß-catenin interacts with Smad1/5/9, and both were recruited at the promoter of Cyp26b1. CONCLUSIONS: Our findings suggested the BMP9-induced osteoblastic differentiation was mediated by activating retinoic acid signalling, viadown-regulating Cyp26b1. Meanwhile, Cyp26b1 may be a novel potential therapeutic target for the treatment of bone-related diseases or accelerating bone-tissue engineering.

Caseinate-gelatin and caseinate-hydrolyzed gelatin composites formed via transglutaminase: chemical and functional properties.

BACKGROUND: Treatment of food proteins by enzymatic crosslinking and other reactions can confer modified properties on the treated proteins. Bovine gelatin and hydrolyzed bovine gelatin were used to generate two caseinate-based composites via transglutaminase, and potential useful properties to food processing were investigated for both composites. RESULTS: Caseinate-gelatin and caseinate-hydrolyzed gelatin composites contained 33.4 and 10.3 g kg(-1) protein of 4-hydroxyproline, respectively. Caseinate conjugation with gelatin and hydrolyzed gelatin resulted in two composites with stronger absorption at five wavenumbers during Fourier transform-infrared analysis, demonstrating that they were rich in hydroxyl and carboxyl groups. Both composites exhibited higher viscosity values in aqueous dispersions, lower thermal stability (i.e. higher mass loss) during thermogravimetric analysis and worse emulsifying properties than original caseinate, owing to conjugation and crosslinking via transglutaminase. However, confocal laser scanning microscopy (CLSM) analysis revealed that both composites actually had better emulsion stability after 2 weeks of storage. CONCLUSION: The composites generated were different in chemical characteristics and better in viscosity and emulsion stability than original caseinate. They might have potential as protein thickeners and emulsifiers. CLSM is a better technique to assess emulsion stability of food proteins than the classic turbidity method.