Objective To analyze the changes of Blood Cell Count and D-dimer concentration in lung cancer patients with thrombosis.Methods ①554 cases of patients with lung cancer were included between January 2012 and November 2013,75 patients with other diseases which lead to high coagulation state were excluded,blood cell count and D-dimer concentration were compared between 64 lung cancer patients were the cases with thrombosis with 415 lung cancer patients were the con-trol group without thrombosis.②Color doppler ultrasonography was taken as the golden standard,receiver operating charac-teristic curves (ROC)were drawn for indexes which had significance in method.Results ①Compared with lung cancer pa-tients without thrombosis,lung cancer patients without thrombosis had decreased red blood cell count (RBC),increased white blood cell count (WBC)and D-dimer.And there were no significant change in platelet (PLT).②The area under curve (AUC)of ROC in RBC,WBC and D-dimer were 0.662,0.637 and 0.896,respectively.By thecut-off values of RBC>4.06× 1012 L,WBC<5.37×109/L and D-dimer<4.02 mg/L,the negative predictive values of RBC,WBC and D-dimer for diagno-sis of lung cancer patient with thrombosis were 93%,93% and 96%,respectively.By the cut-off value of D-dimer>4.02 mg/L,the positive predictive value of D-dimer for diagnosis of lung cancer patient with thrombosis was 6 6%.Conclusion RBC,WBC and D-dimer were related with the progress of thrombosis in lung cancer patients.RBC,WBC and D-dimer have good negative prediction effect on lung cancer patients with thrombosis.D-dimer had positive effect for diagnosis of lung cancer with thrombosis.

BACKGROUND:Cel replacement therapy as an effective strategy for reconstruction of the central nervous system has very broad application prospects.
OBJECTIVE:To investigate the effect of stereotactic transplantation of neural stem cels into the brain on the neuromotor function of craniocerebral trauma rats.
METHODS:Twenty male Sprague-Dawley rats were equivalently randomized into study and control groups. Animal models of craniocerebral trauma were made using the improved free-fal method in the rats. Then, model rats in the study and control groups were given parenchymal transplantation of embryonic neural stem cels and the same volume of culture medium with no stem cels at 1 day after injury, respectively. Neuromotor function of rats was assessed based on the neurological severity scores. At 2 weeks after transplantation, brain tissues were taken for hematoxylin-eosin staining, anti-BrdU, glial fibrilary acidic protein, β-tubulin III and tyrosine hydroxylase immunohistochemistry staining.
RESULTS AND CONCLUSION:The neurological severity scores in the study group were significantly lower than those in the control group at 1 and 2 weeks after injury (P< 0.05). In the study group, there were many BrdU-positive neural stem cels in the brain tissues, some of which were positive for glial fibrilary acidic protein, β-tubulin III and tyrosine hydroxylase; while in the control group, there was no BrdU-positive cel in the brain tissues. Experimental findings show that neural stem cels stereotacticaly transplanted into the brain can proliferate and differentiate in the brain lesion, and thereby notably improve the neuromotor function of rats with craniocerebral trauma.

Objective During pregnancy , exosomes can be released from the placenta into maternal circulation and play im-portant roles in normal pregnancy or placenta-related diseases .We aimed to establish a simple and efficient method for isolating and i-dentifying placental exosomes from maternal serum and lay a foundation for the studies of pregnancy -related diseases . Methods Using sucrose gradient centrifugation with 8% PEG6000 precipitation twice , we isolated and purified placenta-derived exosomes from normal maternal serum and detected their molecular markers CD 63 , CD81 and PLAP by Western blot , followed by silver staining anal-ysis of the protein profile of the exosome pellet .We identified the morphology of the placenta-derived exosomes by transmission electron microscopy ( TEM) and measured the size and distribution of the particles by dynamic light scattering ( DLS) . Results Silver stai-ning of the protein profiles of the exosomes after sucrose gradient centrifugation clearly revealed the bands of the protein molecules . Western blot showed the expressions of CD 63, CD81, and PLAP in the 21-34%density layer, which demonstrated the presence of serum placental exosomes mainly in the 1.09-1.16 g/mL density layer.TEM exhibited that the placenta-derived exosomes were round or oval cup-shaped, specifically expressing PLAP, and the particles were uniform in size, with a mean diameter of (41.79 ±11.94) nm. Conclusion A simple, fast, and efficient method was successfully established for isolating placenta-derived exosomes from ma-ternal serum, which provides a basis for studying the roles of placental exosomes in normal pregnancy and placenta -related diseases.

Objective:The aim of this study was to identify fibrinogen ( FG ) on the development of intimal hyperplasia ( IH ), using an organ culture model. Methods：Segments(n=9 ) of human saphenous vein ( HSV ) wereharvested during coronary artery or infrainguinal vein bypass surgery. The culture medium supplemented with FG (from0 mg/ml to 5 mg/ml ). The proliferation of smooth muscle cell ( SMC ) quantified by 5′-Bromodeoxyuridine (5′-BrdU) uptake in the final four days of the culture period. Histologic analysis and computerized morphometric analysis were used to determine intimal and medial thickness and area，then the intima/media thickness ratio and intima/media area ratio were calculated. Monoclonal antibodies to 5′-BrdU were used as an immunohistochemical maker for proliferating SMC. Results：Addition of FG ( 2.5 mg/ml ) to the cultured medium caused a significant increase in median ( range ) of intima/media thickness ratio and intima/media area ratio of these segments when compared with the normal cultured vein segments ( Wilcoxon paired rank test )：0.387versus 0.215（P=0.017 ）and 0.396 versus 0.229（P=0.015 ），respectively. Addition of FG ( 5.0 mg/ml ) to the cultured medium also caused a significant increase in median ( range ) of intima/media thickness ratio and intima/media area ratio of these segments when compared with the normal cultured vein segments: 0.421 versus 0.215（P=0.008 ）and 0.382 versus 0.229 （P=0.011 ），respectively. However，there were no significant differences in the two vein segments which 2.5 mg/ml or 5.0 mg/ml FG in cultured medium (P>0.05 ).In addition, there was no significant difference in the median ( range ) of intima/media thickness ratio and intima/media area ratio of the segments which FG ( 0.5 mg/ml ) in cultured medium when compared with the normal cultured vein segments ( P>0.05 ). These were supported by SMC proliferation index using staining with 5′-BrdU. Conclusion：High concentration FG at local preianastimotic area may an important factor for IH and early postoprative vein graft restenosis or occlusion.

Nanobody is a kind of antibody from camel, which misses light chain. Nanobody has the same antigen binding specificity and affinity as mAb. Moreover, because of its small molecular weight, high stability and easy preparation, nanobody has great value of biomedical applications. In this study, we successfully prepared highly pure antiEGFR nanobody in E.coli using genetic engineering techniques. Cell proliferation assay (CCK-8 assay) and migration experiments (cell scratch test and Transwell assay) indicated that the recombinant antiEGFRnano can significantly inhibit the proliferation and migration of endometrial cancer cells. These results provide a new way of thinking and methods for EGFR-targeted therapy of endometrial cancer.

OBJECTIVE To screen small molecule compounds from traditional Chinese medicine that can enhance recombinant adeno-associated virus (rAAV) transduction. METHODS Recombinant adeno-associated virus (rAAV) has been established as a powerful tool for in vivo gene transfer and achieved much promise in gene therapy applications. However, widespread clinical use has been limited by transduction efficiency.In the current study,we screened a panel of small molecule compound from traditional Chinese medicine focused on AAV intracellular trafficking process and found salvianolic acid B can significantly enhance rAAV2 transduction. RESULTS Salvianolic acid B caused a dose-depen-dent increase in rAAV2 transduction regardless of vector dose,genome architecture,and over a broad range of cell line from various cell type and species(HEK293,HeLa,HepG2,Huh-7,CHO-K1,LO-2). Salvianolic acid B treatment redirected rAAV2 particles toward large vesicles positive for late endosomal (Rab7)and lysosomal(LAMP1)markers.Furthermore,salvianolic acid B acted to increase accumulation of viral particles at the perinuclear region. CONCLUSION In summary, our results suggest that salvi-anolic acid B redirects rAAV2 toward more productive trafficking pathways and stabilizes perinuclear accumulations of vectors,facilitating productive nuclear trafficking.

BACKGROUND:The high level of matrix metalloproteinase 9 (MMP9) is thought to slow down the healing of diabetic foot ulcers. Whether it can influence the biological behaviors of skin fibroblasts and affect wound healing is stillunclear. The present study aimed to observe changes in the biological behaviors of rat dermal fibroblasts induced by high expression of MMP9 and to clarify the possible mechanisms of wound healing for diabetic foot. METHODS:A cellmodel of skin fibroblast with high expression of MMP9 was established by co-culture of high glucose (22.0 mmol/L) and homocysteine (100 μmol/L). A control group was incubated with normal glucose (5.5 mmol/L). Realtime PCR, ELISA and gelatin zymography were used to detect the MMP9 mRNA, protein expression and activity of MMP9. Flow cytometry, CCK-8, ELISA assay, scratch test and transwellwere used to detect cellproliferation, viability, collagen (hydroxyproline) secretion, horizontal migration and vertical migration of cells. The data were expressed as mean±SD. P value less than 0.05 was considered statistically significant. RESULTS:The expression of MMP9 mRNA, protein levels and the activity of MMP9 were much higher in the high MMP9 group than in the control group (7.05±1.02 vs. 1.00±0.00, 206.9±33.6 pg/mL vs. 40.4±5.9 pg/mL, and 1.47±0.13 vs. 0.57±0.12, respectively,P<0.01). The proportion of S-phase cells, proliferation index, cellviability, collagen (hydroxyproline) secretion, horizontal migration rate and the number of vertical migration cells were lower in the high MMP9 group than in the control group (P<0.01). CONCLUSION:Fibroblasts with a high expression of MMP9 decreased proliferation, activity, secretion and migration of collagens, suggesting that MMP9 may inhibit the biological behaviors of fibroblasts.

Abstract: Objective　To detect the distribution of CYP2A6∗2, CYP2A6∗10, CYP2A6∗17, CYP2B6∗4, CYP2B6∗6, and CYP2B6∗18 loci affecting the metabolism of artemisinins in Kazak population in Xinjiang. To explore the pharmacogenetic background of the Kazak population in Xinjiang for artemisinin drugs and provide clinical decision support for the treatment and prevention of malaria based on artemisinin drugs. Methods　Six SNPs including CYP2A6∗2, CYP2A6∗10, CYP2A6∗17, CYP2B6∗4, CYP2B6∗6, and CYP2B6∗18 were selected for the sequencing experiment. 330 whole blood samples were collected from the Kazak population in Xinjiang. After extracting the whole blood DNA genome, multiplex PCR and high-throughput sequencing were used for genotyping. The allele frequencies were analyzed using the Hardy-Weinberg equilibrium. Results　In this study all SNPs follow the Hardy-Weinberg equilibrium (P>0.05), there was no significant difference in the distribution of SNPs between different genders (P>0.05). The number of successfully sequenced samples of CYP2A6∗2, CYP2A6∗10, CYP2A6∗17, CYP2B6∗4, CYP2B6∗6, and CYP2B6∗18 were 326, 319, 328, 318, 322 and 328 respectively. The frequencies of variant alleles of CYP2A6∗2, CYP2A6∗10, CYP2A6∗17, CYP2B6∗4, CYP2B6∗6, and CYP2B6∗18 in Kazak population are: 0.61%, 0%, 0%, 30.97%, 22.98%, 0%. Conclusions　Mutation alleles affecting the metabolism of artemisinins exist in the Kazak population in Xinjiang. When using artemisinins, the relationship between the drug effect and individual pharmacogenetic background should be further explored.

To construct a scFv library by phage display technique from the spleen cells of mice immunized with B3HM cells. Three mice were immunized with B3HM cells, and their spleen cells were harvested. The genes of VH and Vk were amplified by RT-PCR from the cDNA of the immunized spleen cells and a scFv-phage display antibody library was constructed. The capacity of library was measured,and the variety of the library was analyzed by digesting with restriction endonuclease BstNI.ScFv phage clones were randomly picked and identified phage-scFv clone by binding B3HM cells using immunofluorescein.A scFv library containing 5?106 individual clones which showed different patterns after digested with restriction endonuclease BstNI was produced. Individnal phage-scFv clone showed B3HM cells positive using immunofluorescein. A scFv library of anti-B3HM cell surface molecules has been constructed. It will be useful for finding out some novel genes of causing leukemia, and establishs the infarctate foundation of clarifying the pathogenesis of leukemiagenesis.

OBJECTIVE: To identify sarcosaphagous flies and their larvae, pupa. METHODS: Sarcosaphagous flies and their larvae, pupas were collected from human corpses and their surroundings in the Weifang city. A 304 bp region in COI gene was analyzed by mtDNA sequencing. RESULTS: The studied region showed no sequence divergence within same species and significant difference were found between different species in all samples. CONCLUSION It is a practical approach to identify these Sarcosaphagous flies and their larvae, pupas by sequence analysis of the 304bp region of the COI in mtDNA.
Animals ; Base Sequence ; China ; DNA Primers ; DNA, Mitochondrial/genetics* ; Diptera/genetics* ; Electron Transport Complex IV/genetics* ; Forensic Medicine ; Genes, Insect ; Humans ; Larva/genetics* ; Phylogeny ; Polymerase Chain Reaction/methods* ; Pupa/genetics* ; Sequence Analysis, DNA/methods* ; Species Specificity