ObjectiveTo optimize the nutrient components of yeast-derived serum-free medium(SFM)and determine the composition and concentration by Plackett-Burman design and response surface methodology(RSM).MethodsBased on the SFM selected in the early stage of experiment,10 nutrient components were screened by PBD,using Chinese hamster ovary(CHO)cells as the research object,which were optimized secondarily by RSM to elucidate the interaction among the variables and determine the best medium composition for the growth of CHO cells,after the factors promoting the growth of CHO cells were screened out. CHO cells in logarithmic growth stage were re-suspended in the optimized culture medium,and then subcultured and amplified.ResultsAmong the 10 variables screened by Plackett-Burman experiment,yeast extract(YE)FM888 and trace elements effectively increased the specific growth rate of cells;Non-essential amino acids and FM888 showed significant effects on improving cell survival days;FM888,essential amino acids,non-essential amino acids and trace elements significantly increased the density of viable cells. Based on this,three factors,FM888,trace elements and non-essential amino acids,were comprehensively selected for RSM optimization test. When cultured in the optimized medium in suspension,the maximum specific growth rate of CHO cells was 0. 025/h,the survival days were 6d,and the maximum viable cell density was 7. 187 × 10~6cells/mL. The optimized medium was used for cell passage test,based on which,the cells were stably passaged for 12 passages,and the viable cell density and cell viability were close to those of the control group(commercial SFM).ConclusionThe yeast-derived SFM with clear components and supporting the high-density growth of CHO cells was obtained,which provided a reference and basis for the application of YE in animal cell culture and the development of high-efficiency SFM.

Objective To investigate the therapeutic efficacy and side effect of combination of intra-articular injection of hyaluronate and extracorporeal shock wave therapy(ESWT) on patients with knee osteoarthritis.Methods50 patients with knee osteoarthritis were treated by combination of intra-articular injection of hyaluronate and ESWT(study group, n=22) or combination of intra-articular injection of hyaluronate and pain-point injection of betamethasone(control group, n=28). The parameters included pain intensity (Numeric Rating Score, NRS) at rest and at exercise, pain relief rate, the time when pain disappeared and times of betamethasone administration or ESWT. The side effects were also evaluated.ResultsNRS decreased in the two groups after treatment, while NRS was not different between the two groups(P>0.05) except 1 week after the treatment(P<0.01).ConclusionThe combination of intra-articular injection of hyaluronate and extracorporeal shock wave is a safe and effective treatment for knee osteoarthritis.

Objective:To analyze the correlated factors on the non-public hospitals efficiency performance and compare different types of them in order to provide reference for improving efficiency.Methods:Based on the result of 160 non-public hospitals total efficiency,questionnaire surveys were taken for collecting the information of region,quality,form of ownership,whether invested by the government,whether under the medical insurance,the total amount of employers,bed and service amount as the independent variables,while the total efficiency as the dependent variable to conduct the Tobit multi regression analysis.Results:Access policy,investment policy,nature of ownership,the number of beds,the proportion of physicians,the per capita services,assets and equipment had significant impact.Conclusion:The development of non-public hospital was related to the policy environment,operation mode and internal management.

Encouraging and providing guidance to social forces to invest in health care industry is one of the key elements determined to deepen the health care reform.From four aspects of the unclear conception and undefinable functions of non-public hospitals, ambiguity of property rights of non-profit non-public hospitals, gradual reduction of traditional management measures, and deficiency of effective supervision on daily operations, this paper summarizes the major policy issues in non-public hospitals.To solve these problems, this paper conducts a systemic analysis of the historical experiences of the United States, Germany, Japan, and Taiwan.On this basis, this paper suggests that the government should clearly determine the connotation and extension of non-public hospitals, improve non-profit non-public hospital policy system, play a guiding role in planning and health insurance guidance, and strengthen the supervision of business operations.

Objective:To evaluate the technical efficiency and scale efficiency of non-public hospitals in China to provide references for improving efficiency of non-public hospitals with the comparison with the differences among different non-public hospitals with different types.Methods:Input and output data of 160 non-public hospitals in 2014 were collected;C2R and BC2 models were adopted to evaluate the annual total efficiency,technique efficiency and scale efficiency.Results:There were 16 total efficient hospitals(10.00％),41 technical efficiency hospitals(25.63％) and 20 scale efficiency(12.50％),43 at IRS stage,99 at DRS stage.Conclusion:The overall efficiency of non-public hospitals was low relatively and most of them were lower than public hospitals at same level.Scale was an important factor affecting the efficiency of non-public hospitals.

In this paper, 172 non-public hospitals have been investigated covering east, central and western regions with structured and semi-structured questionnaire and group interviews. We found that the health insurance policies were the most concerned policies by non-public hospitals, accounting for 26.5% of the total. We also found that non-public hospitals from different areas, levels profit nature have different policy expectations, for example Eastern regional institutions wanted to make a breakthrough of the planning policies, the central region wanted to change the health personnel policy, while the western region is considered to be limited by land policy. Tertiary hospitals were more concerned about research projects and land policy, and secondary hospitals were more concerned about access policy. There were 44.8% of the surveyed institutions considered that regulatory policywasmore stringent public hospitals and public hospitals, and there were no difference between profit and non-profit hospitals. According to the survey results, this paper puts forward suggestion that we should implement more targeted policies, purchasing more services from non-public hospitals and enhanced supervision. Considering the survey results research team puts forward sensitive advice as: provide preferential policies in according with different kinds of institutions reinforce purchasing service and enhance non-public hospitals supervision.

Objective To investigate of human esophageal squamous carcinoma cells(Eca-109) in which cyclooxygenase (COX)-2 gene was knocked down by adenovirus delivered siRNA.Methods Based on the plasmid pSUPER cloned with RNA polymerase Ⅲ-dependent promoter HI,the interfering plasmid psiRNA/COX-2 targeting human COX-2 mRNA was constructed,The siRNA/COX-2 fragment was derived from psiRNA/COX-2 digested by Not I and Xho 1.and was cloned into the shuttle plasmid pAdTrack.Then pAdTrack/siRNA/COX-2 was obtained and co transfected into the E.coli strain BJ5183 with the bone plasmid pAdEasy-1,the recombinant adenovirus Ad/siRNA/COX-2 was generated by homologous recombination.Having been packaged and amplified in cells 293,Ad/siRNA/COX-2 was transfected into Eca-109 cells.The PGE2 concentration in the cells culture supernatant was determined by ELISA,and the level of COX-2 mRNA in the cells was tested by real time PCR.Moreover,cell cycle and apoptosis were determined by flow cytometry,and cells growth curve was protracted.Results The recombinant adenovirus Ad/siRNA/COX-2 was successfully constructed.Ninty six hours after Ad/ siRNA/COX 2 transfecting into Eca 109 cells,COX-2 mRNA was reduced by 71.7%,and PGE2 concentration in the cells culture supernatant was decreased by 62.0%.Correspondingly,the growth of cells slowed down.At the same time,the cells in G0-G1 phase was increased by 32.24%,and those in S phase and G2-M phase were reduced by 16.38% and 15.86%.respectively.And cells apoptosis index was increased by 9.19%.Conclusion The adenovirus based-RNAi was capable of knocking down remarkably COX-2 of human esopbageal carcinoma cells,which lead to growth of cells slowing down.

Air ; analysis ; Hydrobromic Acid ; analysis ; Ion-Selective Electrodes ; Workplace

Objective To investigate the eftect of VEGF gene transtection on endothelial progenitor cell derived from murine bone marrow.Methods Wistar rat's bone marrow was obtained, mononuclear cell isolated,and endothelial progenitor cells(EPS)were cultured in EGM-2MV.EPCs were identified by immunocytochemistry and electron microscope.EPCs were transfected by liposome mediated pcDNA3.0-hVEGF165.VEGF protein level was determined in the cultural medium supernatant after VEGF transfection by ELISA.Cultural medium supernatant was used to co-culture with ECV304,VEGF protein activity was evaluated by MTT.EPCs expression of vWF,VEGF,FLK-1 was detected by immunocytochemistry.Results EPCs were effectively enriched by EGM-2MV,and the EPCs obtained express the typical cell surface markers such as CD34,CD133,FLK-1.The concentration of VEGF protein in supernatant reaches 1280 pg/ml in the 7th day after pcDNA3.0-hVEGF transfection.No influence of EPCs proliferation could be found after transfeetion.The cell surface marker expression of VEGF,FLK-1, vWF became higher with time,and the ratios of positive cell were 88.52%,82.65% and 95.97% respectively.Conclusions pcDNA3.0-hVEGF165 transfeet EPCS mediated by liposome could excrete a high concentration of functional VEGF protein.It is helpful for EPC to maintain the characters of endothelial cell after VEGF gene transfection and differentiate to mature endothelial cell.

ObjectiveTo observe the spinal cord pathological change and hindlimbs motor function of rats after decompression on chronic spinal cord injury.Methods 30 Wistar rats were divided randomly into conrol group ( n =5 ),compression group ( n =5 ) and decompression group ( n =20 ),then the decompression group was subdivided into 4 parts as 1 week,2 weeks,4 weeks,6 weeks after decompression.Decompression was made after 30 days of chronic spinal cord copression injury,rats'motor function was detected by inclined plate experiment,the histopathological change,Nissl body and neural cells apoptosis in different spinal cord sections were assessed by the stainings of HE,Nissl and TUNEL.ResultsCompared with control group,the behavior testing showed all rats in compression group presented with weakness of muscle power in their hindlimbs (P ＜ 0.01 ),but the hindlimbs motor functions recovered from the first week after decompression and the difference had statistical significance compared with the compression group(P＜ 0.01 ).Then the rats hindlimbs functions recovered gradually later on.The staining of HE,Nissl and TUNEL showed that the injured spinal cord section performed no improvement at the first 2 weeks after decompression,the neural apoptosis index(24.31 ± 4.73 )％ decreased until the forth week after decompression(P＜0.05).The spinal cord segments closed to the injured part recovered in early stage.At 1 week after decompression,lots of Nissl bodys were observed in spinal anterior horn,the neural apoptosis index in the 2 sections closed to the injured part were ( 15.21 ± 4.81 ) ％ and ( 16.21 ± 3.98 ) ％,which declined observably compared with compression group(P ＜ 0.05 ).ConclusionAfter decompression on chronic spinal cord injury,the recovery of rats'hindlimbs motor function in early stage is probably benefited by the functional compensation of the spinal cord segments closed to the injured section.