ObjectiveTo employ the vascular endothelial growth factor (VEGF165) gene transfected rabbbit osteoplast to combine with the porous nano-hydrxyapatite crystal (n-HA)/polyam ide 66(/PA66) so as to evaluate the osteogenesis and rapid revascularization early after repair of the rabbit radius bone defects with the tissue-engineered bone (n-HA/PA66/osteoplast/VEGF165).MethodsThe animal models of bilateral radius bone defects were created in 56 New Zealand white rabbits that were then randomly divided into Group A and Group B.In Group A, the animals were implanted with n-HA/PA66on the left bone defects (Group Al) and with n-HA/PA66/VEGF165 composite materials on the right bone defects (Group A2). In Group B, the animals were implanted with n-HA/PA66/osteoplast/VEGF165 composite materials on the left side (Group B1) and with n-HA/PA66/osteoplast on the right side (Group B2).Gross, digital radiography, histological sections, vessel count and scanning electron microscopy (SEM) were performed at 2, 4, 8 and 12 weeks after operation.ResultsThe osteogenesis and revascularization in Group B1 was superior to that in the other groups at each time point, with statistical difference (P ＜0.05).The revascularization and osteogenesis in Groups B1 and B2 was far better than that in Groups A1 and A2, with no statistical difference between Group A1 and Group A2.ConclusionsThe new tissue-engineered bone (n-HA/PA66/osteoplast/VEGF165) has a perfect osteogenetic effect and can promote rapid revascularization and the bone healing in the early stage after repair of the bone defects.

BACKGROUND:Bone marrow mesenchymal stem cells are considered as commonly used seed cells to construct tissue-engineered for repair of bone and cartilage defects. It is of great significance for cytology and tissue engineering experiments to study the common problems existing in the basic operation and how to avoid these problems in a timely manner. OBJECTIVE:To summarize the common problems existing in the process of operation in order to provide reliable methods about separation, culture and identification of bone marrow mesenchymal stem cells for beginners and researchers. These can reduce or avoid some errors and problems during operation. METHODS:Sixteen New Zealand white rabbits were selected as experiment objects, and bone marrow mesenchymal stem cells were separated from rabbits by iliac puncture, purified and augmented by using density gradient centrifugation combined with adherent culture method. Then cellmorphology was observed by inverted phase contrast microscope, growth curve detected by MTT method and cellphenotype identified by flow cytometry. RESULTS AND CONCLUSION:We encountered some problems in the process of separation and culture, when we operated the first five rabbits. After careful y summarizing and analysis of the reasons, the operation was successful y completed on the rest 11 rabbits. Bacteria pol ution and cellaging were not found in the process of cellculture. What is more, the cells at passage 3 appeared with high-expression of CD29, and CD44, but low expression of CD14 and CD34. The cellgrowth curve showed that the proliferation activity of cells at passages 3 and 5 was higher than that at passage 10. Although the technology of separation, culture and identification of bone marrow mesenchymal stem cells is mature, the failure wil be happen if we do not pay attention to the details of operation. By strictly carrying out normal operations, we can get high purity of bone marrow mesenchymal stem cells, which lays a good foundation for celland animal experiments in the future.

Objective To investigate the preliminary clinical result of thoracolumbar fracture combined with posterior ligamentous complex injuries repaired by posterior transpedicular screw fixation.Methods A retrospective review was performed on 22 patients with thoracolumbar flexion-distraction fracture combined with posterior ligamentous complex injuries treated with transpedicular screw fixation from July 2008 to March 2013.There were 16 males and 6 females with mean age of 39 years (range,23-62 years).After medically stable,posterior pedicle screw fixation was performed under intravenousinhalational anesthesia.According to the degree of fracture displacement and types of ligament injury,posterolateral bone grafting or intervertebral fusion at the level of injury was conducted.Vertebral height restoration,Cobb' s angle and American Spinal Injury Association (ASIA) score were reviewed preoperatively,at postoperative 3 days and at the last follow-up.Results All the patients were operated on smoothly.There were no complications during operation.All the patients were followed up for 5-51 months (mean,26.5 months).Fracture reductions were satisfied with the closure of vertebral posterior element.Mean anterior vertebral height and Cobb' s angle improved by 20.6％ and 10.60°respectively after operation (P ＜0.01).Eight patients with neurological dysfunction showed some recovery after operation with the mean sensory score improved by 20.7％ (P ＜ 0.05) and mean motor function score improved by 30.9％ (P ＜ 0.0l).All bone grafts were healed,without pain,loosening or breakage in the fixation system.Conclusions Posterior pedicular screw fixation attains good short-term outcome for thoracolumbar flexion-distraction fracture combined with posterior ligamentous complex injuries.The surgery provides satisfactory reduction and instant spinal three-column stability for the unstable spine fracture.Sufficient bone graft is the guarantee to permanent stability.

With the organic combination of rapid on-site evaluation (ROSE) and interventional pulmonary diagnostic technology, we can build a complete The System of Diagnostic Interventional Pulmonology Based on Rapid on-site Evaluation. With the help of ROSE, changing the ways, methods and modalities of interventional pulmonary diagnostic technology to obtain the target lesions is the core of this system. In this statement, the most commonly used standard operating techniques in The System of Diagnostic Interventional Pulmonology Based on Rapid on-site Evaluation are described in detail, including double-hinge curette operating technique, transbronchial lung biopsy (TBLB) technique, and transbronchial brushing technique.

Objective To understand the primary structure and potential antigenic epitopes of antigen Pf332(Ag332) of P.falciparum iso late FCC1/HN.Methods　Based on the published Pf332 gene sequence , nine pairs of primers were designed for the PCR amplification of the Pf332 gen e fragments from genomic DNA of P.falciparum isolate FCC1/HN. The amplified gene fragments were subcloned into pMD-18T vectors and sequenced. The sequences were aligned using DNAstar software to obtain the full-length sequence of the gene Pf332. The primary structure and sequence homology of Ag332 were analyzed by SAPS, Tmpred, SingalP and Blastn programs. Three fragments, R0, R1 and R2, cor responding to nt#9595-10083, nt#10339-10767 and nt#10855-11247 of Pf332 gene were subcloned into the eukaryotic expression vector pcDNA3-S separately. The Balb/c mice were immunized with pcDNA3-S-R0, pcDNA3-S-R1 and pcDNA3- S-R2 separately, and the expressions of the recombinant proteins were detected by immunohistochemistry assay. The protective immune responses elicited by DNA I mmunization were analyzed by ELISA and parasite growth inhibition tests in vitro .Results　Nine Pf332 gene fragments were specifically amplif ied, subcloned into pMD-18T vectors and sequenced. Pf332 gene of the P.falci parum isolate FCC1/HN was 16,377 bp in length, encoding a protein of 5,458 ami no acids, about 615.28kDa. The Ag332 contains 17 regions of highly degenerated Glu-rich repeats, with 30.18% Glu in total amino acids of Ag332. Ag332 of P.falciparum isolate FCC1/HN and 3D7 exhibited 94.55 % homology in amino acid residues. The results of immunohischemistry assay showed that R0, R1 and R2 were expressed in mice muscle tissue. The amount of IgG antibody of the groups immu nized with pcDNA3-S-R0, pcDNA3-S-R1 and pcDNA3-S-R2 were higher than those of blank and pcDNA3 groups (P＜0.05). The result of parasite growth inhibition test showed that the immunized sera at 1∶5 dilution of groups of pcDNA3-S-R0, pcDNA3-S-R1 and pcDNA3-S-R2 had an incomplete inhibitor y effect on P.falciparum growth. Conclusion　The antigen Pf332 is an large protein containing highly degenerated Glu-rich repeats. Pf332 gene fragments, R1 and R2 encoding potent antigenic epitope repeats.

Objective:To study the differential gene expression and immune cell infiltration of gout patients,to find the key genes and immune cells of gout pathogenesis,and to explore the relationship between immune cells and gout.Methods:The gout chip GSE160170 was downloaded from the GEO database,and the differential gene expression analysis was carried out with the help of R language.Then,the STRING database was used to analyze the differential gene,and the Cytoscape software was used to screen the key genes,and then carry out enrichment analysis.At the same time,the infiltration of immune cells were analyzed.Results:The study found that IL-6,IL-1β,TNF,CCL3,CXCL8 and CXCL1 were key genes in the pathogenesis of gout,which were mainly exerted by IL-17,Toll-like receptor,NOD-like receptor,NF-κB and other signaling pathways.Processes such as cellular responses to lipo-polysaccharides,bacteria-derived molecules,and biological stimuli lead to disease;immune infiltration results indicate that memory B cells,activated NK cells,activated dendritic cells,activated mast cells and eosinophils were involved in the disease.It was signifi-cantly expressed in gout patients;the correlation analysis between immune cells showed that the expression of follicular helper T cells were positively correlated with the expression of activated mast cells,and the expression of unactivated NK cells and monocyte were negatively correlated.Conclusion:Key genes and differentially expressed immune cells are closely related to the pathogenesis of gout,providing new ideas for the study of the molecular mechanism of gout.