OBJECTIVE: To construct a VEGFR1 and VEGFR2 bi-targeting oligopeptide-based fusion protein A7/G6-LDP and investigate its anti-angiogenic activity and the mechanism of action. METHODS: PCR and overlap PCR were used to construct the fusion protein A7/G6-LDP expression vector that consists of the gene encoding G6, A7, LDP, and the linker peptide. The product was purified by affinity chromatography and analyzed by SDS-PAGE and HPLC. The binding activity to endothelial cells was examined by ELISA and immunocytochemical staining. The inhibition of HMEC-1 proliferation was determined with CCK-8 assays. The phosphorylation of AKT and c-Raf was detected by Western blotting. HMEC-1 migration was determined using a wound healing assay and tube formation was measured after incubation on Matrigel. RESULTS: The data of DNA sequence confirmed that the A7/G6-LDP fusion protein was correctly constructed. The fusion protein was recovered in high yield (up to 20 mg·L-1) and high purity after His-tag purification. A7/G6-LDP bound to HMEC-1 and HUVEC, respectively; in addition, it inhibited endothelial cell proliferation more effectively than LDP alone when used higher concentration. Moreover, A7/G6-LDP disrupted the formation of endothelial tube structures and inhibited endothelial cell migration. The mechanistic study showed that A7/G6-LDP decreased the phosphorylation of AKT in HMEC. CONCLUSION: The engineered VEGFR1 and VEGFR2 bi-targeting oligopeptide-based fusion protein A7/G6-LDP effectively inhibits anti-angiogenesis. It might serve as a drug delivery carrier in targeted cancer therapy.

Objective To study the diagnosis and treatment of neuroendocrine carcinoma of the nasal cavity and paranasal sinuses, and analyse the influencing factors of suvival and prognosis. Methods We retrospectively analyzed the diagnosis and treatment process of 14 patients with sinonasal neuroendocrine carcinoma (SNEC) admitted in The First Affiliated Hospital of Kunming Medical University from 2007 to 2011. All patients were followed up to learn the survival status of them.Results All patients were followed up for one year up to six years except 2 patients who gave up treatment. Five patients died and six survived with good tumor control in the followed up period. Two patients received only endoscopy surgery, and one of them died from lung metastasis in 21 months after operation, and the other one survived with good tumor control, the disease free survival (DFS) was 9 months . Eight patients were treated by endscopy surgery and /or chemo-radiotherapy, three cases died in following-up period, and five of them survivied with good tumor control, and the disease free survival was 20.25 months.Two patients with transcatheter arterial infusion chemotherapy survivied with good tumor control within the follow up period,and the DFS was 25.5 months.Five patients had moderately differentiated SNEC and DFS was 25.5 months. Seven patients had poorly differentiated SNEC with DFS 14.6 months. Six patients were T4N0M0, four patients were T3N0M0, two patients T2N0M0, and their DFS were 19 months, 12.8 months and 33 month, respectively. Conclusions Surgery with radiotherapy and/or chemotherapy is the current treatment method for sinonasal neuroendocrine carcinoma. Small cell neuroendocrine carcinoma with poor differiation displays highly aggressive and poor prognosis. Diagnosis and treatment in early stage is important for good prognosis.

OBJECTIVE:To ensure the stability of electronic data capture(EDC)system in drug clinical trials and to improve the quality of drug clinical trials. METHODS:The quality control system for EDC system was established and introduced from the formulation of quality control process,establishment of data standard,trial project management,daily management,trial project design,system operation,system function,etc. RESULTS & CONCLUSIONS:Data standard have been achieved through estab-lishing EDC quality control system by our hospital based on attributable,legible,contemporaneous,original and accurate principle. The management of trial project and daily management are conducted through data registration,staff training,the formulation of da-ta management plan,fault emergency treatment,database backup;multiple verification of support data,data lock and export,trial report autogeneration and other functions have been realized by formulating related standard operation instruction,program file,op-eration manual and quality record. Those aspects improve facticity,accuracy and integrality of data in clinical trials,and lay a foun-dation for further data mining.

OBJECTIVE:To promote the supervision and management of clinical trial by institution. METHODS:The structure and function of clinical trial management system(CTMS)developed by our hospital and other enterprise together were analyzed to evaluate the application and operation result of CTMS. RESULTS & CONCLUSIONS:CTMS of our hospital is made up of foun-dation,efficiency and strategy. It is equipped with role allocation,information exchange and report,information warning,drug tracking,clinical trial process control,quality control of electronic record,electronic signature and integration and connection with other system,etc. Relevant operation procedure is established to promote standardization and institutionalization of CTMS. Due to the application of CTMS,the cooperation among departments become smoother,and management level have been enhanced in dai-ly management,pharmacy management,subjects and document administration. It also simplifies the work of researcher and reduc-es the human error by the autogeneration of trial records and tables with the system. Consequently,the monitor coveraged through-out all the trial process.

Mutation (substitution, deletion, insertion, etc.) in nucleotide acid causes the maximal sequence lengths of exact match (MALE) between paralogous members from a duplicate event to become shorter during evolution. In this work, MALE changes between members of 26 gene families from four representative species (Arabidopsis thaliana, Oryza sativa, Mus musculus and Homo sapiens) were investigated. Comparative study of paralogous' MALE and amino acid substitution rate (d(A)<0.5) indicated that a close relationship existed between them. The results suggested that MALE could be a sound evolutionary scale for the divergent time for paralogous genes during their early evolution. A reference table between MALE and divergent time for the four species was set up, which would be useful widely, for large-scale genome alignment and comparison. As an example, detection of large-scale duplication events of rice genome based on the table was illustrated.
Amino Acid Sequence ; Amino Acid Substitution ; Animals ; Arabidopsis ; genetics ; Chromosome Mapping ; methods ; Conserved Sequence ; genetics ; Evolution, Molecular ; Humans ; Mice ; Molecular Sequence Data ; Oryza ; genetics ; Proteome ; genetics ; metabolism ; Sequence Alignment ; methods ; Sequence Analysis, Protein ; methods ; Sequence Homology, Amino Acid ; Species Specificity

Objective To explore the effect of vascular endothelial growth factor antisense oligonucleotides(AS-VEGF)on HL60 cell and HL60/VCR multidrug resistance cell and analyze the function of P-gp and the expression of related multidrug resistance genes including Bcl-2,Mcl-1,MDR1,MRP,GST?,TopoⅡ? and TopoⅡ?.Methods A vector AS-VEGF which expressed in eukaryotic cell was established,then transfected the vector into HL60 and HL60/VCR by limposome transfection technology,observed and drew the growth curve by Tapanlan taining,RT-PCR was used to detect the expression of Bcl-2,Mcl-1,MDR1,MRP,GST?,TopoⅡ? and TopoⅡ? in mRNA level after transfected 24 h and 48 h.Western Blot was used to detect the expression of P-gp in proteinum level after transfected 24 h and 48 h.Results The growth of HL60 and HL60/VCR was inhibited by AS-VEGF(1.25 mmol/L).Between HL60 and HL60/VCR,AS-VEGF decreased the expression of MDR1,MRP,GST? and TopoⅡ? but could not influence the expression of Bcl-2,Mcl-1 and TopoⅡ?,and the expression of P-gp was also obviously decreased in 48 h compared with that in 24 h.Conclusions AS-VEGF can inhibite the growth of HL60 and HL60/VCR and reverse multidrug resistance by changing cell microenvironment and the cell membrane correlated protein transportating channel,reduce the cell disintoxicating and the self-repair ability.

Objective To investigate the effect of human cytomegalovirus (HCMV) on proliferation of colony forming unit T-lymphocyte (CFU -TL)and its interference methods. Methods Normal CFU - TL culture was used as blank control. Normal CFU- TL culture system plus inactivated HCMV fluid as inactivated HCMV control. The dilution of 1:10,1:100,1:1000 were added into CFU -TL colonies culture system directly as infected group. Astragalus (AMI) and ganciclovir(GCV) were added into culture system with HCMV dilution of 1:10 as experimental group. By methylcellulose semi-solid culture, different concentrations of HCMV - AD1699 affect CFU-TL and interfered by astragalus AMI, GCV. CFU - TL were surveyed. The effect of HCMV on CFU-TL proliferation was measured by MTT; HCMV-AD169 DNA in CFU-TL was found by PCR. Results 1. Compared with control group, the numbers of CFU -TL in the HCMV infection groups decreased significantly(P

OBJECTIVE: To observe the expression pattern of caspase-3 and HCLS1-associated protein X-1 (HAX-1) at different time after cerebral contusion in rat, and explore the new method for estimating the injury interval. METHODS: The cerebral contusion model was established using adult SD male rats. Then the rats were randomly allocated into 8 groups: 2 h, 6 h, 12 h, 1 d, 3 d, and 7 d after cerebral contusion, sham-operation and normal control. Expression of caspase-3 and HAX-1 protein after cerebral contusion in rat was detected by Western blotting. Laser scanning confocal microscope was used to observe the number of HAX-1 positive cells and TUNEL-stained cells after cerebral contusion. RESULTS: The expression of caspase-3 increased parallelly with the time after cerebral contusion and reached the peak value on 3 d. The expression of caspase-3 decreased gradually and still maintained a high level expression on 7 d (P < 0.05). The expression of HAX-1 positive cell went up after injury, and reached the peak value at 6 h (P < 0.05), then turned down gradually after 12 h and went out of detection after 3 d. The number of TUNEL-stained cells increased obviously at 2 h and reached the peak value on 3 d. The number of TUNEL-stained apoptotic cells decreased gradually and still maintained a high level expression on 7 d (P < 0.05). CONCLUSION The expression of caspase-3 and HAX-1 after cerebral contusion has time sequential regularity, which may provide new evidence for forensic diagnosis of cerebral contusion interval.
Animals ; Blotting, Western ; Brain Injuries/pathology* ; Carrier Proteins/metabolism* ; Caspase 3/metabolism* ; Cerebellum/pathology* ; In Situ Nick-End Labeling ; Intracellular Signaling Peptides and Proteins ; Male ; Rats ; Rats, Sprague-Dawley

To explore the correlation between the ecological factors and the contents of podophyllotoxin and total lignans in root and rhizome of Sinopodophyllum hexandrum, podophyllotoxin in 87 samples (from 5 provinces) was determined by HPLC and total lignans by UV. A correlation and regression analysis was made by software SPSS 16.0 in combination with ecological factors (terrain, soil and climate). The content determination results showed a great difference between podophyllotoxin and total lignans, attaining 1.001%-6.230% and 5.350%-16.34%, respective. The correlation and regression analysis by SPSS showed a positive linear correlation between their contents, strong positive correlation between their contents, latitude and annual average rainfall within the sampling area, weak negative correlation with pH value and organic material in soil, weaker and stronger positive correlations with soil potassium, weak negative correlation with slope and annual average temperature and weaker positive correlation between the podophyllotoxin content and soil potassium.
Berberidaceae ; chemistry ; China ; Chromatography, High Pressure Liquid ; Climate ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; Ecosystem ; Lignans ; chemistry ; isolation & purification ; Podophyllotoxin ; chemistry ; isolation & purification ; Soil ; chemistry ; Temperature

Objective To study the foxml gene and its protective effect on the lung tissue of rats with acute lung injury (ALI) induced by lipopolysaccharide (LPS), and to observe the dexamethason' s (DEX) impacts on foxml gene and the prognosis of ALI. Method Seventy-two healthy mice were randomly(random number) divid-ed into three groups: control group (A group, n = 24), model group (B group, n = 24) and DEX treatment group (C group, n = 24). The observing intervals were respectively set in 24 h, 48 h and 72 hours. At each ob-serving interval, the foxml protein in lung tissue of mice was detected by using immunohistochemistry (IHC), and the expression of foxml gene in lung tissue was detected by using RT-PCR, as well as to observe the pathological changes in lung tissue. Results Comparisons were made between paired groups at 24 h,48 h and 72 h intervals in which the expression of foxml mRNA and the level of foxml protein in lung tissue of mice in C group were signifi-cantly higher than those in B group (P < 0.05), and those in B group were significantly higher than those in A group (P < 0.05). The expression of foxml mRNA and the level of foxml protein in lung tissue of mice in B group at 48 h interval were significantly higher than those both at intervals of 72 h and 24 h (P < 0.05), and the those at 72 interval were significantly higher than those at 24 h interval (P < 0.05). Compared with B group, the pathologi-cal changes in lung tissue of mice in C group were lessened. Conclusions In both model group and dexamethasone treatment group, the expression of foxml mRNA and the level of foxml protein in lung tissue of mice are increased significantly. Dexamethasone lessens the injury of both vascular endothelial cells and alveolar epithelial ceils of lung tissue, and it also significantly increases the expression of foxml mRNA and the level of foxml protein.