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Objective:To construct BimS lentivirus RNA interference(RNAi) vector and to study its infection efficiency by using RNAi technique.Methods:Three interference targets were designed according to the BimS sequence.The single chain primer was annealed into double-stranded oligo sequences,and then connected with vector linearized with Age Ⅰ and EcoR Ⅰ enzyme.The recombinant plasmid was packaged,and the infection efficiency was observed by infecting ACC-2 cells.Results:After amplification,a 337 bp band was appeared in the electrophoresis results of positive clones.Sequence of inserted fragments were identical with the result of DNA sequencing.Restructuring lentivirus was packed in 293T cells,the virus titer was 2 × 108 TU/ml,MOI =20,and the transfection efficiency was 85％.The BmS mRNA relative expression of pFU-GV-BmS-1,pFU-GV-BMS-2 and control group was 0.743 ±0.025,0.466 ±0.023 and 1.266 ±0.042 respectively(between each 2 groups,P ＜0.05).Conclusion:BimS lentivirus virus RNA interference vectors can be constructed,and can efficiently infect ACC-2 cells.
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Objective To establish the method of isolation,culture and identify biological characterization of mesenchymal stem cells from human umbilical cord (hUCMSCs);and study their multiple differentiation potency.Methods Stem cells from human umbilical cord were cultured by enzyme Wharton jelly method in vitro.The surface markers were identified by flow cytometry.Multi-differentiation capacity was identified by osteogenic and adipogenic differentiation.ALP was detected with Calcium cobalt staining.The mineralized ability in vitro was measured with Alizarin red staining.Theadipocyte differen-tiation ability was measured with oil red-O staining.Results Flow cytometry analysis revealed that CD73 (92.45%),CD90 (95.45%)and CD105 (96.45%)were highly expressed on these cells’surface,while CD34 (1.07%)were negative ex-pressed.Cells were cultured with induced-osteogenic medium after 3 weeks,ALP staining in the cytoplasm of black parti-cles,and a large amount of mineralized nodules within cells was observed after 4 weeks.Cells were cultured with induced-adi-pogenic medium after 2 weeks,the majority of these cells were round,oil red O staining of lipid droplets generated within cells was observed.Conclusion Mesenchymal stem cells from human umbilical cord have the potential of multi-directional differentiation.These cells could be induced to differentiate into adipocytes and osteoblasts,which laid the foundation for clinical stem cell therapy research source of seed cells.
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BACKGROUND:Studies have found that combination of two of chitosan (CS), nano-hydroxyapatite (nHA) and poly(lactide-co-glycolide) (PLGA) can improve the mechanical properties and biocompatibility of the composite stent in certain extent as wel as improve osteogenic differentiation of the cels, but there is a certain distance from the ideal bone tissue engineering scaffolds. OBJECTIVE:To study biocompatibility and osteoinductive activity of nHA/CS/PLGA scaffolds with different proportions in vitro. METHODS: nHA/CS/PLGA scaffolds were prepared at mass ratio of 10:10:80, 10:20:70, 20:10:70 respectively by particle leaching method. And human bone marrow stem cels (hBMSCs) were co-cultured with these scaffolds in vitro. Adhesion, proliferation, and osteoinductive activity of these scaffolds were examined qualitatively and quantitatively by growth curve of hBMSCs on scaffolds. Gene expression of alkaline phosphatase activity and osteocalcin was detected by RT-PCR. RESULTS AND CONCLUSION: hBMSCs could be attached, proliferated, and osteoinduced better on the nHA/CS/PLGA scaffold with the mass ratio of 20:10:70, compared to the other two groups of scaffolds. The differences were significant statisticaly (P< 0.05). Alkaline phosphatase and osteocalcin expressions were respectively higher in the scaffold with the mass ratio of 20:10:70 after 9-27 days of co-culture and 15-27 days of co-culture, in comparison with the other two groups of scaffolds. These findings indicate that the nHA/CS/PLGA scaffolds with the mass ratio of 20:10:70 demonstrated preferable biocompatibility and osteogenic inductivity, which is expected to be a promising scaffold material for bone tissue engineering.
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Overtreatment is not only an economic phenomenon, but also an ethics problem at the same time. It is precisely a medical - social problem. This paper analyzes the reasons of overtreatment in oral disease from the view of medical elthics. Furthermore, it provides the strategy which the doctor, the patient, the medical establishment and the social shoud be adopted on overtreatment in oral disease.
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Objective To examine the changes of matrix metalloproteinase-2,9 (MMP-2,9) in gingival crevicular fluid(GCF) after phase 1 periodontal treatment of adult patients with periodontitis. Methods GCF was sampled with filter paper strips by intra-pocket method to determine MMP-2,9 levels. Forty teeth of forty adult patients with periodontitis and forty teeth of forty periodontally healthy persons were included in this study. Assays for MMP-2,9 in GCF were performd by ELISA. Results Contents of MMP-2,9 were higher in AT group than those in controls(P