BACKGROUND:Whether transplanted bone marrow mesenchymal stem cells under hypoxic conditions can survive is crucial for the successful celltransplantation. Therefore, studies on the growth of bone marrow mesenchymal stem cells under hypoxic conditions in vitro can provide experimental evidence for in vivo celltransplantation. OBJECTIVE:To observe the expression of vascular endothelial growth factor in bone marrow mesenchymal stem cells under hypoxia. METHODS:Rat bone marrow mesenchymal stem cells were obtained and cultured, and observed under light microscopy. Passage 3 cells were cultured under normoxia (21%O2) and hypoxia (3%O2 hours. Then cellcounting kit-8 assay and flow cytometry were employed to detect cellproliferation in the two groups. Western blot assay was adopted to detect the expression of hypoxia-inducible factor-1αand vascular endothelial growth factor in the two groups. RESULTS AND CONCLUSION:(1)Rat bone marrow mesenchymal stem cells were obtained and cultured successful y, which were fusiform cells and had uniform shape under the light microscope. (2)The results of cellcounting kit-8 assay showed that the number of cells in the hypoxic group was higher than that in the normoxic group at each time point, and cellviability increased significantly at hours 36 and 48 (P<0.05). (3)The results of flow cytometry demonstrated that the proportion of cells in S phase and cellproliferation index in the hypoxic group were significantly increased, compared with the normoxic group (P<0.05). (4)Western blot results showed ), respectively, for 72 that there was a smal amount of the expression of hypoxia-inducible factor-1αand vascular endothelial growth factor in the normoxic group, but the expression of these two proteins in the hypoxic group was increased in a time-dependent manner (P<0.05). These findings suggest that hypoxia can induce proliferation of rat bone marrow mesenchymal stem cells cultured in vitro, and also raise hypoxia-inducible factor-1αand vascular endothelial growth factor expression in a time-dependent manner.

AIM: To investigate the effects of polidatin on phospholipase A_2(PLA_2),nitric oxide(NO),and endothelin-1(ET-1) levels in serum and lung homogenate of hypoxic pulmonary hypertension rats.METHODS: 29 healthy Spraugue-Dawley rats were randomly divided into normal control group,chronic hypoxic group and hypoxic plus polidatin group.The model of chronic hypoxic pulmonary hypertension was made by method of intermittent isobaric hypoxia for 21 days.The mean pulmonary arterial pressure(mPAP) was measured by inserting a microcatheter into the pulmonary artery.The ratio of right ventricular wall and that of left ventricular wall and ventricular septum weight(R/L+S) was measured as well.RESULTS: After exposed to hypoxia for 21 days,the mPAP,R/L+S,PLA_2 and ET-1 activities in blood and lung homogenate increased significantly. The NO concentration decreased significantly.Pretreatment with PD attenuated these changes mentioned above.CONCLUSION: Pretreatment with PD effectively prevents the formation of chronic hypoxic pulmonary hypertension.Its mechanism may be related to the inhibition of PLA_2 and ET-1 and promotion of NO production.

Dengue virus (DENV) envelope [E] protein is the major surface protein of the virions that indued neutralizing antibodies. The domain III of envelope protein (EDIII) is an immunogenic region that holds potential for the development of vaccines; however, the epitopes of DENV EDIII, especially neutralizing B-cell linear epitopes, have not been comprehensively mapped. We mapped neutralizing B-cell linear epitopes on DENV-1 EDIII using 27 monoclonal antibodies against DENV-1 EDIII proteins from mice immunized with the DENV-1 EDIII. Epitope recognition analysis was performed using two set of sequential overlapping peptides (16m and 12m) that spanned the entire EDIII protein from DENV-1, respectively. This strategy identified a DENV-1 type- specific and a group-specific neutralizing epitope, which were highly conserved among isolates of DENV-1 and the four DENV serotypes and located at two regions from DENV-1 E, namely amino acid residues 309-320 and 381-392(aa 309-320 and 381-392), respectively. aa310 -319(310KEVAETQHGT319)was similar among the four DENV serotypes and contact residues on aa 309 -320 from E protein were defined and found that substitution of residues E309 , V312, A313 and V320 in DENV-2, -3, -4 isolates were antigenically silent. We also identified a DENV-1 type-specific strain-restricted neutralizing epitope, which was located at the region from DENV-1 E, namely amino acid residues 329-348 . These novel type- and group-specific B-cell epitopes of DENV EDIII may aid help us elucidate the dengue pathogenesis and accelerate vaccine design.
Amino Acid Sequence ; Animals ; Antibodies, Neutralizing ; immunology ; Dengue ; virology ; Dengue Virus ; chemistry ; genetics ; immunology ; Epitope Mapping ; Epitopes, B-Lymphocyte ; chemistry ; genetics ; immunology ; Humans ; Mice ; Molecular Sequence Data ; Viral Envelope Proteins ; chemistry ; genetics ; immunology

AIM:To investigate the affecting factors of detecting platelet activation by flow cytometry (FCM). METHODS:Using decoagulant of natrium citricum,anticoagnlated peripheral venous bloods from 6 healthy donors were labeled with the method of three-colour immunofluorescence assay. Platelet activation markers fibrinogen receptor (Fib-R,PAC-1) and P-selectin (CD62P) were measured. In the same time,the reproducibility of FCM was assessed.RESULTS:The platelet activation markers PAC-1 and CD62P at each time point showed significant difference(P

Molecularly imprinted polymers (MIPs) with high selectivity to tilmicosin (TIM) were prepared using tylosin(TYL) as dummy template, methacrylic acid(MAA) as monomer, ethylene glycol dimethacrylate (EGDMA) as cross-linker.The effects of 4 porogens including dimethyl formamide, methanol, acetone, and chloroform on the recognition capability of MIPs were investigated.Orthogonal test was used to optimize the preparation of MIPs, and the optimal composition was as follows; 1.0 mmol TYL, 8.0 mmol MAA, 20.0 mmol EGDMA, 6.0 mL chloroform, 20.0 mg azobisisobutyronitrile.The solid phase extraction condi tions and characteristics of MIPs as adsorptive material for the selective extraction and enrichment of TIM were also studied.The recovery of TIM was above 90% when the following procedure was applied to MIPs cartridge: conditioning with methanol and water(pH 9.0), loading with acetonitrile, cleaning with methanol and chloroform respectively, and eluting with 3 mL methanol-ammonia(95:5, V/V).The recovery of TIM on non-imprinted polymers cartridge was only 32%.

Objective To construct a recombinant expression vector for expression of the function-al domains of dengue virus serotype 1 ( DENV1 ) envelope ( E ) protein in native soluble form. Methods The genes encoding the functional domains of DENV1-E protein (1-394 aa) were amplified with PCR and then cloned into the Psectag2B-Fc eukaryotic expression vector.The 293T cells were transfected with the recombinant vector by cationic lipid-based delivery.The cell clones expressing the fusion DENV1-E-Fc protein were screened out with 2 mg/ml of Zeocin.Immunofluorescence assay ( IFA) was performed to analyze the antigenicity and integrity of the fusion protein.The fusion proteins were purified from cell lysate with Protein-G and further identified by Western blot assay.Results The soluble form of fusion protein with a molecular weight of about 90×103 was obtained at a yield of about 25 μg per 1×107 cells.The results of IFA indicated that the fusion protein kept its integrity with right conformational epitopes.The fusion protein was successfully expressed with the advantage of good specificity as indicated by IFA and Western blot assay. Conclusion The recombinant fusion protein in soluble form was successfully expressed in eukaryotic ex-pression system, which paved the way for further investigation on the function of DENV1 E protein and its protective epitopes.

Objective To retrospectively investigate the disease spectrum of inpatients aged over 65 year and cost constitution in Sichuan Provincial People's Hospital from 2010 to 2014,so as to provide baseline data for further study.Methods The inpatients'disease spectrum and costs were collected from hospital information system.The diseases were classified according to the International Classification of Diseases(ICD-10).The data were analyzed using SPSS 18.0 software.Results The total number of old inpatients was 111,935,and male (55.2 ％) was more than female (44.8 ％).The average age was (74.5 ±6.8)years.The top four systematic diseases of primary diagnosis were circulatory system disease (21.0 ％),respiratory system disease (13.7 ％),digestive system disease (12.7％)and neoplasms (12.1％).The total number of male inpatients was more than the female inpatients.The average cost per capita was increased from (￥)18,778.1 yuan to (￥)23,391.9 yuan since 2010.The proportion of all costs accounted for by drugs in elderly inpatients was decreased from 45.5％ to 38.9％ since 2010.Nosocomial infection was decreased from 3.19％ to 0.16％ since 2010.Conclusions The number of elderly inpatients are more in male than in female in Sichuan Provincial People's Hospital from 2010 to 2014.The most common systematic disease is circulatory system diseases,and followed by respiratory system diseases,digestive system diseases and neoplasms.The proportion of all expenditures accounted for by drugs was declined in five years,while the average cost per capita is increased.

Objective To study CT and MRI appearance of transient hepatic attenuation difference (THAI)) .to reveal the cause of THAD),and to avoid false positive and misdiagnosis.Methods 10 cases appearing THAD in CT and 5 cases appearing THAI) in MRI were reviewed and all were processed with plain scan and dy- namic contrast with CT or MRI.Results 7 cases appeared transienl hypertransfusion of CT,4 cases appeared tran- sient hypertransfusion of MRI;3 cases appeared transient Hypoperfusion of CT,1 case appeared transient hypoperfu- sion of MRI.Conclusion The appearance of THAD in CT and MRI,was related to the quick-scan with CT and MRI only sufficient comprehension of the characteristics of blood supply in normal physiology and pathology of liver, combined with plain scan,could make correct decision possible in the final diagnosis when it occurred regional perfu- sion difference in liver.

Effects of different drying methods including sun drying, steamed, boiled, constant temperature drying (at 40, 50, 60 °C) on appearance, hardness, rehydration ratio, dry rate, moisture, total ash, extractive and polysaccharides contents were studied to provide the basis of standard processing method for Tulipa edulis bulbus. The results showed that the treatments of sun drying and 40 °C drying showed higher rehydration ratios, but lower dry rate, higher hardness, worse color, longer time and obvious distortion and shrinkage in comparison with other drying methods. The treatments of 60 °C constant temperature drying resulted in shorter drying time, lower water and higher polysaccharides content. Drying time is shorter and appearance quality is better in the treatment of steaming and boiling compared with other treatments, but the content of extractive and polysaccharides decreased significantly. The treatments of 50 °C constant temperature drying led to similar appearance quality of bulb to commercial bulb, and it resulted in lowest hardness and highest dry rate as well as higher rehydration ratio, extractive and polysaccharides content, moderate moisture and total ash contents among these treatments. Based on the results obtained, 50 °C constant temperature drying is the better way for the processing of T. edulis bulbus.
Color ; Desiccation ; methods ; Plant Stems ; chemistry ; Polysaccharides ; analysis ; Quality Control ; Tulipa ; chemistry ; Water ; analysis

Objective To construct a novel enhanced green fluorescent protein (EGFP) and glial cell line-derived neurotrophic factor (GDNF) recombinant baculovirus. Methods The target gene(EGFP and GDNF) was cloned into baculovirus transfer vector pFastBacDual, pFB-EGFP-GDNF was constructed and restriction enzyme analysis was conducted. pFB-EGFP-GDNF was transposited with baculovirus shuttle vector (Bacmid) into DH10Bac competent cells, and recombination baculovirus vector Bacmid-EGFP-GDNF was constructed. The plasmid was extracted and PCR was performed for identification. Bacmid-EGFP-GDNF was transfected with Sf9 insect cell package virus by liposomal transfection method. Immunofluorescent staining was employed to detect the expression of EGFP and GDNF protein in St9 cells. Results The target gene fragment was correctly cloned into pFastBaeDual vector, and recombinant Bacmid was constructed. Bacmid-EGFP-GDNF was successfully transfected, and higher virus titer was obtained. The coexpression of GDNF and EGFP protein in Sf9 cells was identified by immunofluorescent staining. Conclusion The recombinant baculovirus Bacmid-EGFP-GDNF can be successfully constructed, and the protein of EGFP and GDNF is coexpressed in St9 cells, which paves a way for the research of GDNF gene therapy.