Objective To observe the effect of Sox9 gene overexpressian on chondrogenesis of esenchymal stem ceils (MSCs) in vitro. Methods Rabbit MSCs were obtained and purified by gradient centrifuge and adhesion culture in vitro. MSCs were transfected by recombinant Sox9 plasmid. RT-PCR and Western blot analysis were used for transfection confirmation. Chondrogenic molecules such as type I1 collagen,aggrecan and Sox9 were detected by immunohistology, RT-PCR and western blot. Cell proliferation was tested by MTT assay. Results MSCs were successfully transfected with Sox9 gene and verified by RT-PCR and Western blot analysis. The overexpression of Sox9 had no effect on the proliferation of MSCs. Transfeeted cells expressed more chondrogenic markers than the control groups. Conclusion Sox9 gene overexpression can enhance chondrogenic differentiation of MSCs.

BACKGROUND: Different isolation and culture methods for bone marrow mesenchymal stem cell (BMSCs) will result in varying cell activity and purity, which will influence repair effect of tissue engineering. OBJECTIVE: To investigate the optimized methods of isolation and culture of BMSCs in vitro and validate the efficacy of cell acquisition. DESIGN, TIME AND SETTING: In vitro cytology trial was performed at the laboratory of Department of Orthopedics, Second Hospital of Shanxi Medical University from October to December 2007. MATERIALS: Two 3-month-old New Zealand rabbits were provided by Shanxi Institute of Livestock. METHODS: Lymphocyte isolation solution was added to DMEM culture solution containing fresh bone marrow. BMSCs were obtained and purified by gradient centrifuge and adhesion culture in vitro. Non-attached cells were moved to new culture flask every 8 hours. The solution was changed firstly after 5 days of culture. Trypsinization was conducted at cell confluence of 90%. The first, third and fifth passages of cells were harvested. MAIN OUTCOME MEASURES: The morphology of BMSCs was observed with phase contrast microscope; the growth curve was drawn to determine doubling time. The percentage of the wel1 growth P2 cells were identified by CD44 staining by flow cytometry. RESULTS: Primary cultured BMSCs were oval, spindle-shaped or polygonal, and adhered to plastic surface within 48 hours, exhibiting spindle-shaped or polygonal with clear nucleus, and reached 90% confluence within 14 days. The first, third and fifth passages of BMSCs showed S-type, and were in latent phase at 1-3 days, in log phase 3 days later, and cells came into platform phase at 7-8 days. The mean doubling time was 24.22 hours. CD44-positive cells were found in the second passage, with positive rate of 93.0%. CONCLUSION: BMSCs are easily isolated and cultured in vitro with good growth and high purity by gradient centrifuge and adhesion culture with lymphocyte isolation solution.

Objective To observe the change of collagen in osteoarthritis and study the pathogenesis of osteoarthritis.Methods The right hind limb of twenty-four adult New Zealand White rabbits were immobilized with plaster cast in extension position for 10,20,30 and 40 days,respectively.Six animals without immobilization served as control.The articular cartilage of the medial femur condyle was harvested for transmission electron microscopy,in-situ hybridization of collagenⅡ,and immunohistochemistry of collagenⅠ,Ⅱ,Ⅲ.Results Transmission electron microscopy showed articular cartilage was destructed from the fine collagen fiber network of tangential zone.The fine collagen fiber network did not contain chondrocyte.Immunohistochemistry and in-situ hybridization showed that in earlier period of osteoarthritis,the collagen typeⅡand its gene expression firstly increased,then decreased with destruction of ultrastructure,and chondrocytes enhanced type Ⅱ collagen expressing and synthesizing mainly in transition zone and upper deep zone.In articular cartilage of osteoarthritis there was type Ⅲ collagen,instead of typeⅠcollagen.Conclusion In osteoarthritis,articular cartilage degenerated from tangential layer,in which collagen can not be repaired after destruction,this may contribute to the chondral degeneration.

Through introducing mutations into ribosomes by obtaining spontaneous drug resistance of microorganisms, ribosome engineering technology is an effective approach to develop mutant strains that overproduce secondary metabolites. In this study, ribosome engineering was used to improve the yield of butenyl-spinosyns produced by Saccharopolyspora pogona by screening streptomycin resistant mutants. The yields of butenyl-spinosyns were then analyzed and compared with the parent strain. Among the mutants, S13 displayed the greatest increase in the yield of butenyl-spinosyns, which was 1.79 fold higher than that in the parent strain. Further analysis of the metabolite profile of S13 by mass spectrometry lead to the discovery of Spinosyn α1, which was absent from the parent strain. DNA sequencing showed that there existed two point mutations in the conserved regions of rpsL gene which encodes ribosomal protein S12 in S13. The mutations occurred a C to A and a C to T transversion mutations occurred at nucleotide pair 314 and 320 respectively, which resulted in the mutations of Proline (105) to Gultamine and Alanine (107) to Valine. It also demonstrated that S13 exhibited genetic stability even after five passages.
Genetic Engineering ; Macrolides ; metabolism ; Point Mutation ; Ribosomal Proteins ; genetics ; Ribosomes ; metabolism ; Saccharopolyspora ; metabolism

OBJECTIVE: To establish a cone beam computed tomography (ECBCT) system for high-resolution imaging of the extremities. METHODS: Based on three-dimensional X-Ray CT imaging and high-resolution flat plate detector technique, we constructed a physical model and a geometric model for ECBCT imaging, optimized the geometric calibration and image reconstruction methods, and established the scanner system. In the experiments, the pencil vase phantom, image quality (IQ) phantom and a swine feet were scanned using this imaging system to evaluate its effectiveness and stability. RESULTS: On the reconstructed image of the pencil vase phantom, the edges were well preserved with geometric calibrated parameters and no aliasing artifacts were observed. The reconstructed images of the IQ phantom showed a uniform distribution of the CT number, and the noise power spectra were stable in multiple scanning under the same condition. The reconstructed images of the swine feet had clearly displayed the bones with a good resolution. CONCLUSIONS The ECBCT system can be used for highresolution imaging of the extremities to provide important imaging information to assist in the diagnosis of bone diseases.
Algorithms ; Animals ; Artifacts ; Calibration ; Cone-Beam Computed Tomography ; instrumentation ; methods ; Equipment Design ; Extremities ; diagnostic imaging ; Image Processing, Computer-Assisted ; methods ; Phantoms, Imaging ; Radiographic Image Enhancement ; instrumentation ; methods ; Swine

OBJECTIVE: To develop a digital breast tomosynthesis (DBT) imaging system with optimizes imaging chain. METHODS: Based on 3D tomography and DBT imaging scanning, we analyzed the methods for projection data correction, geometric correction, projection enhancement, filter modulation, and image reconstruction, and established a hardware testing platform. In the experiment, the standard ACR phantom and high-resolution phantom were used to evaluate the system stability and noise level. The patient projection data of commercial equipment was used to test the effect of the imaging algorithm. RESULTS: In the high-resolution phantom study, the line pairs were clear without confusing artifacts in the images reconstructed with the geometric correction parameters. In ACR phantom study, the calcified foci, cysts, and fibrous structures were more clearly defined in the reconstructed images after filtering and modulation. The patient data study showed a high contrast between tissues, and the lesions were more clearly displayed in the reconstructed image. CONCLUSIONS This DBT imaging system can be used for mammary tomography with an image quality comparable to that of commercial DBT systems to facilitate imaging diagnosis of breast diseases.
Algorithms ; Artifacts ; Breast ; diagnostic imaging ; Female ; Humans ; Mammography ; methods ; Phantoms, Imaging ; Radiographic Image Enhancement ; methods

OBJECTIVE: We propose a sparse-view helical CT iterative reconstruction algorithm based on projection of convex set tensor total generalized variation minimization (TTGV-POCS) to reduce the X-ray dose of helical CT scanning. METHODS: The three-dimensional volume data of helical CT reconstruction was viewed as the third-order tensor. The tensor generalized total variation (TTGV) was used to describe the structural sparsity of the three-dimensional image. The POCS iterative reconstruction framework was adopted to achieve a robust result of sparse-view helical CT reconstruction. The TTGV-POCS algorithm fully used the structural sparsity of first-order and second-order derivation and the correlation between the slices of helical CT image data to effectively suppress artifacts and noise in the image of sparse-view reconstruction and better preserve image edge information. RESULTS: The experimental results of XCAT phantom and patient scan data showed that the TTGVPOCS algorithm had better performance in reducing noise, removing artifacts and maintaining edges than the existing reconstruction algorithms. Comparison of the sparse-view reconstruction results of XCAT phantom data with 144 exposure views showed that the TTGV-POCS algorithm proposed herein increased the PSNR quantitative index by 9.17%-15.24% compared with the experimental comparison algorithm; the FSIM quantitative index was increased by 1.27%-9.30%. CONCLUSIONS The TTGV-POCS algorithm can effectively improve the image quality of helical CT sparse-view reconstruction and reduce the radiation dose of helical CT examination to improve the clinical imaging diagnosis.