[Effect of Angelica sinensis polysaccharide on the osteogenic differentiation of bone marrow mesenchymal stem cells of rats with high glucose levels].

OBJECTIVE: This study aims to evaluate the effect of Angelica sinensis polysaccharide (ASP) on the osteogenic differentiation of the bone marrow mesenchymal stem cells (BMSCs) of rats with high glucose levels. METHODS: Rat BMSCs were isolated and identified by osteogenic and adipogenic differentiation. Then, the BMSCs were divided into three groups as follows: normal control group (5.5 mmol·L⁻¹ glucose), high glucose group (25.5 mmol·L⁻¹ glucose), and ASP+high glucose group (25.5 mmol·L⁻¹ glucose +40 mg·L⁻¹ ASP). The proliferation activities of the BMSCs were detected by CCK8. Alizarin red staining, and alkaline phosphatase activity were used in the examination of osteogenic activity. Quantitative real time-polymerase chain reaction was used to detect the expression levels of the osteogenic genes (Runx2, Osx, OCN, Col-Ⅰ) and the key factors of Wnt/ß-catenin signal pathway (CyclinD1, ß-catenin). In vivo, a type 2 diabetes rat model was established. The rats were divided into three groups, namely, the normal control group (normal rats), diabetes group (diabetic rats), diabetes+ASP group (diabetic rats, ASP feeding). Then, the tibia bone defect was established. The repair of bone defects in each group was observed through histological examination. RESULTS: The proliferation of BMSCs was higher in the high glucose group and ASP+high glucose group than in the normal control group (P<0.05). No significant difference was observed between the high glucose group and ASP+high glucose group (P>0.05). The number of calcium nodules of BMSCs; alkaline phosphatase activity; and the mRNA expression of Runx2, OCN, Osx, Col-Ⅰ, CyclinD1, ß-catenin in the high glucose group were lower than those in the normal control and ASP+high glucose groups (P<0.05). No significant difference was observed between the normal control and ASP+high glucose groups (P>0.05). The bone mass was significantly lower in the bone defect of the diabetes group than in the bone defect of the normal control or diabetes+ASP group (P<0.05). No statistical difference was found between the normal control and diabetes+ASP groups (P>0.05). CONCLUSIONS: ASP can promote the osteogenic differentiation of rat BMSCs under high glucose culture and induce bone regeneration in rats with type 2 diabetes. These features may be related to the activation of the Wnt/ß-catenin signaling pathway.

[Effect of TGF-ß3 transfected with adipose derived stem cells and OGP-HA-chondroitin sulfate scaffold on repair of condylar cartilage in rabbits].

PURPOSE: To investigate the feasibility of repairing damaged cartilage of rabbit condylar with TGF-ß3-transfected adipose stem cells combined with OGP (osteogenesis peptide)-HA (hyaluronic acid)-ChS (chondroitin sulfate) scaffold. METHODS: Rabbit ADSCs were isolated and cultured. The expression vector carrying TGF-ß3 gene was constructed using recombinant adenovirus and transfected into rabbit ADSCs. The fluorescence expression of the cells was observed after 14 days and the efficiency of virus transfection was calculated. Western blot was used to detect the expression of TGF-ß3 protein. Fifty rabbits were randomly divided into 5 groups: group A as blank group, group B as TGF-ß3 transfected ADSCs group, group C as OGP-HA-ChS scaffold group, group D as ADSCs complex OGP-HA-ChS group, E group was TGF-ß3 transfected ADSCs with OGP-HA-ChS. After the rabbit temporomandibular joint osteoarthrosis model was established, the animals were transplanted according to the experimental design. The animals in each group were sacrificed after the 3rd and 9th weeks of transplantation. Scanning electron microscopy, histological observation and real-time fluorescent quantitative PCR were performed accordingly. SPSS17.0 software package was used for analysis of variance and t test. RESULTS: Scanning electron microscopy and histological observation showed that the repair of cartilage lesions in group D and E was better than those in group B and C. Compared with group B, the results of real-time quantitative PCR showed that the expression of MMP-3 in group E was similar to that in group A, but significantly lower than that in group C and D (P<0.05). The expression of TIMP-1 in group E was similar to that of group A, but significantly higher than group C and D (P<0.05). CONCLUSIONS: TGF-ß3 transfected ADSCs composite OGP-HA-ChS scaffold has repairing effect on rabbit condylar damaged cartilage.

[Effects of PRF and released three growth factors on migration of rat adipose tissue-derived stem cells].

PURPOSE: To analyze the effects of PRF and released three growth factors on migration of rat adipose tissue-derived stem cells and to investigate the mechanism of migration. METHODS: The inguinal adipose tissue of rat was excised at aseptic condition to obtain primary ADSCs by enzyme digestion. Multi-directional differentiation was used to identify the ADSCs. PRF membrane was acquired through one time centrifuge. The cell migration was examined by Transwell assay and wound healing assay. The mRNA expression of MMP2 and MT1-MMP was tested by real-time PCR. Statistical analysis was performed using SPSS 13.0 software package. RESULTS: Cell migration test showed that the migration of rat ADSCs in PRF group were significantly higher than those in the negative group（P<0.05） and inhibitor group（P<0.05）. The ADSCs migration effects in three growth factors group at different concentrations showed significant difference（P<0.05）. Real-time PCR showed that gene expressions of MMP2 and MT1-MMP were significantly higher in PRF group than control group (P<0.05). PCR showed that gene expressions of MMP2 and MT1-MMP were significantly higher in three growth factors group than control group (P<0.05). CONCLUTIONS: PRF and three growth factors consistently enhanced the migration of rat ADSCs in a dose-response manner. The migration increase of rat ADSCs may be associated with the up-regulation of MMP2 and MT1-MMP gene expression.