0.05).The size of PCR fragments ranged from 144 to 188bp containing 19～41 CA repeats and the frequency of (CA)n ranged from 0.34%～14.1%.The heterozygosity(H) and polymorphism information content(PIC) were 0.85 and 0.83 respectively.Significant differences were found in frequencies of eleven alleles including 22-repeats,23-repeats,26-repeats,27-repeats,28-repeats,30-repeats,31-repeats,32-repeats,33-repeats,34-repeats and 35-repeats between Chinese and Caucasians.Conclusion:This site is referred to as a genetic marker with high polymorphism information content.And the distribution of this genetic marker is different in different ethnic groups.

[Objective]To prove the feasibility of using the distance between sagittal plane of the spinal process and the enter point to individualize the placing of pedicle screw.[Method]Thirty spine specimen were collected and divided into two groups,data were measured,such as the width of the pedicle,distance between the enter point and anterior border of the vertebra,distance between sagittal plane of the spinal process and the enter point,angle from the longitudinal axis of the pedicle to sagittal axis of the vertebra,angle from the longitudinal axis of the pedicle to vertical line of the operating table.In group one the pedicle screws were placed with the help of the distance between sagittal plane of the spinal process and the enter point,the other by the method advised by Ebraheim.CT scan was applied to evaluate the place of the screws,according to the perforation extent,they were classified into 4 grades:A=totally in the pedicle;B=perforation extent4mm.[Result]The individualized group showed much lower perforation rate than the traditional method group in T3~10,and similar in T1,T2,T11,12.[Conclusion]It can obviously improve the accuracy of the pedicle screw placement to use the distance between sagittal plane of the spinal process and the enter point to localize the enter point,especially when anatomic landmark such as articulationes zygapophysiales and transverse process change.

Background and purpose:The screening of the genes being related closely with the mechanism of osteosarcoma metastasis was a difficult point in the realm of orthopaedics.We screened differential expression gene of human osteosarcoma MG-63 cell sublines with different metastatic capabilities with cDNA microarray,and studied the molecular mechanism of osteosarcoma metastasis.Methods:Total RNA of human osteosarcoma MG-63 cell sublines A1 and A2 was extracted,purified to mRNA and then reversely transcripted to cDNA probe respectively.The cDNA probe of A1 was labelled with Cy3 and the cDNA probe of A2 was labelled with Cy5.The two samples were hybridized with the cDNA microarray.The hybridization signals were scanned by Agilent Scanner and obtained data were analyzed using Ima Gene 3.0 software and Genespring software.Results:222 differential expression genes were found between cell sublines A1 and A2 by analyzing gene expression profile.There were 119 upregulated genes and 103 downregulated genes in cell sublines A1.All differential expression genes belonged to six main function groups and 49 genes of these had very obvious differentce in expression.Conclusions:There were many differently expressed genes between A1 and A2 cell sublines and only part of them were closely associated with mechanism of osteosarcoma metastasis.The technology of cDNA microarray could analyze effectively gene expression profile of human osteosarcoma MG-63 cell sublines, and supply a new approach to study the mechanism of osteosarcoma metastasis

Objective To screen promoter DNA-binding protein of inducible nitric oxide synthase gene by using phage display technique from human liver cDNA library, and to study the expression and regulation mechanism of iNOS gene. Methods The sequence of iNOS promoter was identified in GenBank by bioinformatics based on the open reading frame(ORF) of iNOS gene and amplified from HepG2 genome by polymerase chain reaction (PCR). The amplified product was subcloned into pCAT3-Basic reporter vector, named as pCAT3-iNOSp. The HepG2 cell line was then transfected with pCAT3-Basic to serve as negative control, and pCAT3-promoter which contains the promoter region of CMV served as the positive control subject, and pCAT3-iNOSp served as the test subject, respectively. The choloraphenical acetyltransferase(CAT)activity was determined by enzyme linked immunosorbent assay(ELISA) kit. The T7 Select human liver cDNA library was biopanned and positive clones were selected. After screening, positive plaque was performed to amplify and PCR products were sequenced. Results The expression of CAT in transfection of pCAT3-PS1TP1p was 4.2 times as higher as pCAT3-Basic plasmid. Sequence analysis was performed in 12 positive plaque, which were the iNOSp binding protein. Conclusion The iNOS gene promoter identified in this study has shown to have transcription activity,and iNOS promoter DNA-binding proteins havs been screened. The results will be useful for further study of the expression and regulation mechanism of iNOS in liver cell.

[Objective]To investigate the relationship between degeneration of cartilage and the expressions of Matrix Metalloproteinase 7(MMP-7),Matrix Metalloproteinase 9(MMP-9),Matrix Metalloproteinase 13(MMP-13)and Tissue Inhibitor of Matrix Metalloproteinase 1(TIMP-1)in osteoarthritis.[Method]The histological changes of cartilages by hematoxyllin-eosin staining and immunohistochemical expression of Matrix Metalloproteinase 7(MMP-7),Matrix Metalloproteinase 9(MMP-9),Matrix Metalloproteinase 13(MMP-13)and Tissue Inhibitor of Matrix Metalloproteinase 1(TIMP-1)in osteoarthritis.were studied in 20 osteoarthritis cases and 2 normal controls.All data were statistically analyzed by Mann-Whitney U test and correlation analysis.[Result]The osteoarthritis cartilage underwent fibroplasias and tearing.The quantity of chondrocyte increased and the clustered and hypertrophic cells came into being.Little immunostaining of MMP-7 and MMP-13 was observed in normal cartilage,while their expressions increased in degenerated cartilage(P0.05),superficial layer of the moderate and end-stage osteoarthritis.However,it significantly increased in deep layer(P

[Objective] To study the bone metabolic biochemical index of the aged patients with hip fracture,for better predicting the future risk of the old people' s hip fracture.[Method]50 cases of sufferers(over 60 years old) with hip fracture(28 males,and 22 females) and 30 cases of healthy aged people(15 males,and 15 females) were selected to analyze Ⅰ Collagen crosslinked c-telopeptide(ICTP),deoxypyridino line(Dpd) in urine,and serum bone glaprotein(BGP).[Result](1)The mean level of ICTP and Dpd in urine in aged hip fracture group was higherthan that of the control group(P0.05).[Conclusion]Bone absorbability in the aged hip fracture patients is higher than in the aged healthy people.The analysis of ICTP and Dpd in urine may /might give some reference value in preventing and treatlng aged hip fracture patients.

PBL,problem based learning,is a kind of teaching model for resolving problems. Based on the present teaching situation of our university,my experience in NewYork University and my teaching experience for overseas students,this article elucidated the importance and necessity of PBL teaching model in overseas student education and put forward a suitable PBL teaching method.

Objective To explore the relationship of the (CA)n dinucleotide repeats polymorphism of the eNOS gene with type 2 diabete mellitus (DM) and the serum NO level in DM patients. Methods The eNOS genotype was assessed in 179 unrelated Chinese of Han nationality,including 93 patients with DM and 86 normal controls(NC).Genotype was determined by polymerase chain reaction(PCR) and denature polyacrylamide gel electrophoresis (PAGE) with silver stain.Fasting serum NO level was measured by nitrate reductase and fasting serum NOS level was measured by spectrophotometer. Results Twenty two different alleles were identified containing 19-40 CA repeats,in which the frequency of (CA) 30 (166bp)was the highest. The heterozygosity (H) and polymorphism information content (PIC) were 0.85 and 0.83 respectively. Frequency of allele (CA) 34 was higher in DM (7.5%) than in NC (2.9%) significantly. Compared with NC,the serum NO level was significantly lower in T2DM with genotype (CA)n 1 (CA)n 2 when n 1≥34 or n 2≥34. Conclusions The allele(CA) 34 is associated with the susceptibility to DM and may be involved in the development of DM. The eNOS gene polymorphism is connected with the serum NO level.

Objective To study the impacts of dynamic compressive stress on the mRNA expression of osteopontin ( OPN ),runt related gene 2 ( Runx2 ),osteocalcin ( OC ),osterix,alkaline phosphatase (ALP) and bone morphogenetic protein 2 (BMP-2) in the osteoblasts of Sprague-Dawley (SD) rats. Methods Osteoblasts extracted from skull periosteum tissue of neonatal SD rats were digested using trypsin and collagenase （Ⅰ）,then were subcultured and amplified in vitro.ALP staining and alizarin red staining were performed to identify the purified cells.The cells were treated with compressive stress at 20,50 or 100 mmHg for 24 h.The expression levels of OPN,Runx-2,OC,osterix,ALP and BMP-2 were measured and quantitatively analysed using a real-time quantitative polymerase chain reaction. Results Under 20 mmHg of dynamic compressive stress the expression levels of OPN,Runx2,OC,osterix,ALP and BMP-2 all were elevated compared with the control group.The peak expression oecured under 50 mmHg pressure. The expression levels did not change significantly compared with the control group under 100 mmHg pressure. Conclusions Moderate dynamic compressive stress can promote the expression of OPN,Runx-2,OC,osterix,ALP and BMP-2 mRNA in osteoblasts,which might be an important mechanism for promoting the union of fractures.

BACKGROUND: The precartilaginous stern cells are limited regarding in vitro proliferative capacity, but the immortalized cell lines can provide a large number of stable immortalized cells, and simian virus 40 large T antigen gene (SV40Tag) is one of gene fragments which are commonly used and effective in vitro immortalized ceils. OBJECTIVE: To construct human immortalized precartilaginous stem cells (IPSCs) using human precartilaginous stem calls induced by SV40LTAg gane. METHODS: The human immortalized precartilaginous stem calls were isolated from aborted fetus and purified with enzyme digestion and immunomagnetic beads screening method. By using liposome-mediated gene transfection technology, plasmid pCMVSV40T/PUR containing SV40Tag was transfected in primary embryonic precartilaginous stem cells, while non-transfected cells sewed as negative controls. Positive clones were cultured to observe the cell morphology and the passage recovery, to calculate cell survival rata and population doubling time, to drew call growth curve. Immunofluorescence cytochemistry was used to detect the expression of IPSCs fibroblast growth factor receptor 3, the expressions of SV40Tag and fibroblast growth factor receptor 3 in the human precartilaginous stem cells were determined by RT-PCR. RESULTS AND CONCLUSION: Morphology of human IPSCs seemed coincidence with primary human precartilaginous stem cells. The survival rate of human IPSCs was not influenced by subculture, freezing and recovery, but the survival rate was descended in the human precartilaginous stem cells at the 6~(th) and 10~(th) passages (P < 0.01). Compared with cells at the 6~(th) and 10~(th) passages, the proliferation of human IPSCs was greater, with short population doubling time and high growth rate (P < 0.01). The immunofluorescence showed that fibroblast growth factor receptor 3 was positive in human IPSCs at the second passage, and the RT-PCR results of fibroblast growth factor receptor 3 revealed a specific amplification band at 400 bp,.while that of SV40Tag revealed at 560 bp. No band was seen in the primary cells. It is indicated that SV40Tag human IPSCs can be constructed successfully using immunomagnatic bead screening technology and liposome transfection technique.