95%). Conclusion Immunomagnetic separation can effectively isolate and purify PSCs.

Using gene fusion technology, polypeptide fusion tags can be engineered into target proteins at the genetic level.The resultant recombinant proteins may possess the biochemical properties of the imported fusion tags.Therefore, it is possible to take advantage of fusion tags to improve and evaluate protein expression, to detect and track protein targets, and to purify and characterize proteins.However, it is necessary to eliminate any influence of the fusion tag in structural characterization experiments or in isolating pharmaceutical proteins.Scientists must therefore remove fusion tags prior to structural and functional analyses when fusion tags are suspected of interfering with the biological activity of a protein or influencing its behavior.The fusion tag can be removed by several methods including harsh chemical treatment, mild enzymatic cleavage by endoprotease or exoprotease, and intein-mediated self-cleavage.Here the literature is reviewed in relation to principles, applications, and approaches of each method.

Objective To investigate the clinical features,diagnosis,treatment and prognosis of gastric neuroendocrine carcinoma (G-NEC).Methods Clinical data of 52 G-NEC cases were analyzed.Follow-up was conducted by telephone.The survival curves were drawn using Kaplan-Meier method.Univariate analysis was performed by the Log-rank test and multivariate analysis was performed by COX proportional hazards model.Results The median overall survival rate was 19 months (range 6 to61 months),and the overall 1,3,5-year survival rates were 74％,16％ and 5％ respectively.Tumor stage surgery,vascular nerve involvement and Ⅲ and Ⅳ phase chemotherapy were related to prognosis (x2 =24.254,10.005,7.261,8.790,all P ＜ 0.05).Multivariate analysis showed tumor stage and vascular nerve involvement were independent prognostic factors (x2 =17.170,5.810,all P ＜ 0.05).Conclusion G-NEC is a highly malignant tumor with poor prognosis.Preoperative diagnosis rate is low.Surgery is the treatment of first choice.Definite diagnosis depends on postoperative pathology and immunohistochemical examination.

To investigate the technique of sorting high-purity precartilaginous stem cells from rat's perichondrium, neonatal rat's perichondrium cells suspensions were incubated with monoclone antibody of anti-fibroblast growth factor receptor-3 (anti-FGFR-3), and the labeled cells were separated from the suspension in the magnetic field by immuno-beads coated with the second antibody. Purity of the sorted neural stem cells was found to be 93.0%-99.0%, with living cells amounting to 80% -85 %. The magnetic cell sorting system could effectively separate precartilaginous stem cells from perichondrium cell suspensions.

Herpes simplex virus (HSV) is a double-stranded DNA virus that can infect skin and mucosal epithelial cells. It can establish latency in sensory neurons and sporadically reactivate from these cells. In order to reply to attacks of the host and evade the immunity surveillance during infection and reactivation, HSV has developed a multitude of clever strategies. Dendritic cells (DCs), one of the most important antigen-presenting cells (APC), can recognize pathogens at the infection sites and activate specific T cells, thus playing a crucial role in the host immunity against virus infection. This paper reviewed the mechanism of the host immunity against HSV, especially the role of DCs in HSV-induced immune responses and the future research perspective.

BACKGROUND:There is a lack of study on material properties and parameters of foot finite element models in China. Vernier caliper is a common method for measuring the width and thickness of ligaments and tendons to calculate the cross-sectional area.
OBJECTIVE:To design a new ligament cross-sectional area measuring instrument to improve the measurement accuracy.
METHODS:The cross-sectional area of the five fresh cadaver ankle ligaments was respectively measured using the new instrument and vernier caliper, and then a comparative analysis of the two measurement methods was performend.
RESULTS AND CONCLUSION:The cross-sectional area of anterior talofibular ligament, calcaneofibular ligament, tibionavicular ligament and calcaneotibial ligament was (20.61±7.52), (22.38±11. 49), (33.09±9.91) and (28.20±10.88) mm2, respectively measured by the vernier caliper, and (17.59±4.03), (20.77±7.91), (28.08±8.14) and (30.39±7.98) mm2 by the new ligament cross-sectional area measuring instrument. These results suggest that this new measuring instrument is accurate, reliable and easy to operate, which can be used as a special instrument to measure ligament cross-sectional area, but further studies wil be necessary.

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.

In order to improve the efficacy of vibration expectoration and prevent secondary injury in patients with chest trauma,the body surface marking method was used in vibration expectoration.The exact injured locations and areas were identified via CT ot X-ray results.Then,the restricted and focused areas of expectoration were marked with red and green colors,which could guide the machines to avoid the fracture sites,reduce pain,prevent secondary injury and administrate effective expectoration in the areas of sputum and atelectasis.However,the dynamic CT or X-ray results must be used for reference to ensure the accuracy of the body surface marking.

Objective To investigate the role of nerve growth factor (NGF) in the pathogenesis of diabetic neuropathy (DNP) and the correlation of NGF level with,sensory nerve conduction velocity (SNCV) and duration of DNP.MethodsThe 41 patients with DNP,35 diabetics without DNP and 33 healthy controls were enrolled.And the serum level of NGF was measured with ELISA,the SNCV of left medial nerve,ulnar nerve,peroneal nerve and tibial nerve were measured too. Then the correlation analysis was completed. ResultsThe serum level of NGF was significantly lower in DNP patients [(665.18±188.32) ng/L] than in healthy controls [(976.44±159.07)ng/L] and in diabetics without DNP [(943.32±167.33) ng/L,F=9.316,P<0.001].The NGF level of DNP patients was positively correlated with SNCV of left medial nerve (r=0.810,P<0.001 ),left peroneal nerve (r=0.760,P<0.001) and duration of DNP (r=0.542,P<0.001).Conclusions The serum level of NGF is lower in DNP patients than in healthy controls.The decrease of the production of NGF may play a role in the pathogenesis of DNP.

Immortalized human precartilaginous stem cells (IPSCs) were established to provide stable cell resource for the study of the molecular mechanism of gene targeting on the differentiation of PSCs. Plasmid pCMVSV40T/PUR containing simian virus 40 large T antigen gene (SV40Tag) was transfected into human PSCs by using lipofectin transfection. Colonies were isolated by puromycin selection and expanded by multiple passages. Immunohistochemistry, RT-PCR and Southern blotting were used to identify the transfected cells and to detect the expression and integration of SV40Tag in expanded cell lines. The positive colonies were isolated and subcultured, designated immortalized precartilaginous stem cells (IPSCs), which were confirmed as fibroblast growth factor receptor-3 (FGFR-3) positive cells by immunohistochemistry and RT-PCR. SV40Tag cDNA was found in cultured IPSCs of passage 8 by Southern blotting, and the expressions of SV40Tag mRNA and protein were confirmed by RT-PCR. These findings suggested that IPSCs strain with SV40Tag was constructed successfully.
Cartilage/*cytology ; Cell Proliferation ; Cell Transformation, Viral ; Cells, Cultured ; Fetus ; Simian virus 40/*genetics ; Stem Cells/*cytology ; Transfection