ABSTRACT
BACKGROUND:Although there is a certain progress in the preparation of tissue-engineered bone tissue using a variety of materials, some deficiencies have appeared such as mismatching between scaffold degradation rate and new bone formation rate, slow tissue growth, toxic metabolites. OBJECTIVE:To build a new type of inducible bone repair composite scaffold with bionic bone structurematerials and to evaluate its physicochemical and biological properties. METHODS: Icarin encapsulated by chitosan was used to prepare drug-loaded microspheres, and the drug release rate of the microspheres was detected. Chitosan microspheres were mixed with colagen to build the core part of scaffold materials. Hydroxyapatite (HA), polycaprolactone (PCL) and colagen were mixed in hexafluoride isopropanol (HFIP) to prepare the HA/PCL/colagen outer part of composite scaffold material at the rate of 0:3:3, 1:3:3, 2:3:3, 3:3:3. Each proportional electrospinning was used for one layer, and finaly the 4-layer outer tube of the scaffold was produced. The tube core and outer tube were crosslinked by 1% genipin. Universal material testing machine, surface contact angle meter, infrared spectroscopy, scanning electron microscope, water absorption, permeability, porosity,in vitro degradation tests for cross-linked and uncross-linked were used to observe the structure and characteristics of tubular materials. Bone marrow mesenchymal stem cels were seeded on the surface of cross-linked and uncross-linked bone repair materials to evaluate the biocompatibility of the scaffolds. Cross-linked and uncross-linked bone repair materials were implanted subcutaneously into Wistar rats to evaluate the histocompatibility of the scaffolds. RESULTS AND CONCLUSION:The drug in the scaffold had a suitable release; the bone scaffold material had good uniformity, and cross-linked scaffolds materials had better mechanical properties, water absorption and permeability than the uncross-linked(P < 0.05). The degradation rate of the cross-linked group was significantly lower than that of the uncross-linked group (P< 0.05). Hematoxylin-eosin staining showed that the bone marrow mesenchymal stem cels could adhere wel to the cross-linked and uncross-linked materials. No inflammatory reactions occurred after subcutaneous implantation of cross-linked and uncross-linked materials. These findings indicate that the cross-linked scaffold for inducible bone tissue engineering has good biocompatibility and mechanical properties.
ABSTRACT
BACKGROUND:Our previous studies have shown that salidroside can induce bone marrow mesenchymal stem cells directly into neuron-like cells, and Ca2+signal is one important way to achieve its biological signal transduction. OBJECTIVE:To investigate the role and mechanism of the calcium/calmodulin (Ca 2+/CaM) signaling pathway inducing bone marrow mesenchymal stem cells to differentiate into nerve cells. METHODS:Bone marrow mesenchymal stem cells were divided into two groups:control groups and salidroside groups. Salidroside groups were induced with different concentrations of salidroside (5, 10, 20, 50 and 100 mg/L) for 24 hours and 100 mg/L salidroside was added to culture cells for different time (12, 24, 48 and 72 hours). Western blot assay was used to detect the expression levels of neural cellmarker, microtubule-associated protein 2, and the important protein of Ca2+/CaM signaling pathway:CaM and calmodulin dependent kinase II (CaMK II). Then Ca2+/CaM signaling pathway specific blockers were applied to cells respectively for 30 minutes, including 500 μmol/L EGTA (Ca 2+chelator), 1 mmol/L Nifedipine(L-type Ca2+channel blocker) and 10 mmol/L LY294002 (PI3K inhibitor). Then, 100 mg/L salidroside was added and cultured for 24 hours. Western blot assay was used to detect the expression of neuron-specific enolase and CaM in the Ca2+/CaM signaling pathway. RESULTS AND CONCLUSION:(1) After inducing with salidroside, the expression of microtubule-associated protein 2 were upregulated (P<0.01), indicating that salidrosid can induce the neuronal differentiation of bone marrow mesenchymal stem cells. (2) After different concentrations of salidrosid induced bone marrow mesenchymal stem cells for 24 hours, the expressions of CaM and CaMK II were significantly upregulated in the 10 mg/L group ( P<0.01);For the 100 mg/L salidrosid that was added for cellinduction for different time, the expressions of CaM and CaMK II were significantly downregulated in 72-hour group (P<0.01). (3) After blocking extracellular Ca2+and PI3K signaling pathway, the expressions of neuron-specific enolase and CaM were higher than those in salidrosid groups (P<0.05). These results suggest that salidrosid can induce bone marrow mesenchymal stem cellto directly differentiate into nerve cells by inhibiting the Ca2+/CaM signaling pathway.