Difference of Graft Maturity of Anterior Cruciate Ligament Reconstruction between Oval-Shaped Tunnel Technique and Round-Shaped Tunnel Technique / 中国运动医学杂志

Objective To compare the graft maturity of the anterior cruciate ligament reconstruction using the oval-shaped tunnel technique and round-shaped tunnel technique by signal/noise quotient (SNQ) of MRI postoperatively.Methods Forty patients diagnosed as the anterior cruciate ligament rupture between August 2015 and June 2016 were included according to the inclusion and exclusion criteria.Patients were randomized into a round-shaped group and an oval-shaped group,undergoing traditional round-shaped tunnel and oval-shaped tunnel reconstruction of the anterior cruciate ligament respectively.One year postoperatively,the MRI was conducted,and three intra-articular regions of interest (ROI) were selected to compare the graft maturity by calculating SNQ.Results None of the forty patients experienced complications of bone tunnel blowout,graft getting through difficulties and neurological or vascular injuries.SNQ of the round-shaped group were 3.72 ± 2.29,significantly higher than that of the oval-shaped group(P＜0.001).Moreover,SNQ of proximal ROI and distal ROI of ovalshaped group were 1.97 ± 1.30 and 2.76 ± 1.75,significantly lower than the round-shaped group with proximal site of 3.53 ± 2.11(P=0.008) and distal site of 4.46 ± 2.28(P=0.012).Conclusion Comparing MRI signal intensity one year after the treatment,we have found the graft SNQ after oval-shaped tunnel reconstruction was lower than the round-shaped reconstruction,with better graft maturity.

A cell-based high-throughput approach to identify inhibitors of influenza A virus

Influenza is one of the most common infections threatening public health worldwide and is caused by the influenza virus. Rapid emergence of drug resistance has led to an urgent need to develop new anti-influenza inhibitors. In this study we established a 293T cell line that constitutively synthesizes a virus-based negative strand RNA, which expresses Gaussia luciferase upon influenza A virus infection. Using this cell line, an assay was developed and optimized to search for inhibitors of influenza virus replication. Biochemical studies and statistical analyses presented herein demonstrate the sensitivity and reproducibility of the assay in a high-throughput format (Z' factor value>0.8). A pilot screening provides further evidence for validation of the assay. Taken together, this work provides a simple, convenient, and reliable HTS assay to identify compounds with anti-influenza activity.

Establishment and drug susceptibility test of isoniazid resistant Mycobacterium smegmatis / 药学学报

With the emergence of drug resistant tuberculosis, it is very urgent to find novel anti-tuberculosis drugs, especially novel anti-drug-resistant tuberculosis drugs. Because of the slow growth and the need to work in a biosafty environment of Mycobacterium tuberculosis, the development of evaluation of drug effect is severely impeded. In order to solve these issues, non-pathogenic fast-growing Mycobacterium smegmatis is introduced as test organism. The inhA is one of a target of isoniazid (INH) overexpression or mutation of this gene in Mycobacterium tuberculosis conferring resistant to INH. A recombinant plasmid bearing inhA was constructed and electroporated into Mycobacterium smegmatis, using shuttle expression vector pMV261. Transformants were induced to express a protein of inhA, identified by SDS-PAGE. Results show that Mycobacterium smegmatis containing inhA plasmids exhibited 100-fold or greater increased resistance to INH, but it conferred no increased resistance to others first-line anti-tuberculosis drugs. Resazurin microtiter assay plate testing of Mycobacterium smegmatis susceptibility to drugs is a rapid, simple, and inexpensive method and could decrease color background of drugs by detecting fluorescence. It will be benefit for high-throughout screening of drugs of anti-isoniazid-resistant Mycobacteria.

Studies on antibacterial activity and endotoxin neutralization of murine BPI N-terminal functional fragment(muBPI_(25) protein)in vitro / 中国免疫学杂志

Objective:To establish an experimental model for intracellular antibacteria and endotoxin neutralization in vitro to detect the antibacterial and endotoxin neutralization activity of the muBPI_(25) protein.Methods: RAW264.7 cells were transfected with pcDNA3.1(+)muBPI_(36-259), and then were infected with intracellular bacterial of either G ~+/G~-to establish the experimental model of intracellalar antibacteria.The RAW264.7 cells were co-transfected with the pSecTag2B-muBPI_(36-259) and dual-luciferase reporter gene plasmids for establishment of the experimental model of endotoxin neutralization.Results:The experimental model of intracellular antibacteria confirmed that the muBPI_(25) protein could inhibit/kill Salmonella typhi.The experimental model of endotoxin neutralization indicated that the muBPI_(25) protein could neutralize endotoxin.Conelusion: We firstly demonstrate that murine BPI N-terminal functional fragment(muBPI_(25) protein)can inhibit/kill Salmonella typhi,and can neutralize, its lysating product, endotoxin.

Establishment and application of a screening anti-HIV-1 drug model targeted nuclear trafficking of virus RNA / 药学学报

The HIV-1 Rev protein facilitates nuclear export of unspliced and singly spliced viral transcripts containing RRE RNA through the CRM1 export pathway. Inhibition of Rev-mediated RNA nuclear export can arrest HIV-1 transcriptional process, which clearly, reveals a target for anti-HIV drug development. In this work, a cell-based assay has been established for screening anti-HIV compounds targeting the Rev-mediated RNA nuclear export. This assay utilized a codon-optimized green fluorescent protein (GFP) as reporter gene, which expression is in a Rev-dependent manner. Any compound that inhibits the Rev-mediated RNA nuclear export is identified by reducing emission of GFP. The Z' score of this model is 0.8220. Three thousands compounds were screened and the positive rate was 9.3% with a cutoff at 50% inhibition. IMB7C7, one of the positive compounds, efficiently inhibits viral production from HIV-1 infected cells.

Establishment and application of a high-throughput screening assay for premature activation of HIV-1 precursors / 药学学报

Strict regulation of HIV-1 PR function is critical for efficient production of mature viral particles. During viral protein expression and viral assembly, HIV-1 PR located within Gag-Pol precursor must be inactive to prevent premature cytoplasmic processing of the viral Gag and Gag-Pol precursors. Premature activation of HIV-1 precursors leads to major defects in viral assembly and production of viral particles. A cell-level premature activation of HIV-1 precursors assay using bioluminescence resonance energy transfer (BRET) was established. Three thousand compounds were screened to evaluate this assay. The results showed that the assay is sensitive, specific and stable (Z' factor is 0.905).

Cloning, expression and activity identification of human osteoprotegerin-mycobacteria heat shock protein 70 fusion protein / 中国组织工程研究

AIM: To solve the two difficulties of bone resorption and inflammation in rheumatoid arthritis, clone the recombinant human osteoprotegerin (OPG) and mycobacteria heat shock protein 70 (HSP70) functional gene,and study the expression and activity of OPG-HSP70 fusion protein in E.coli. METHODS: Experiments were performed at the Laboratory of Department of Immunology, Capital Medical University from May 2006 to September 2007. Complementary DNA encoding full length OPG protein was amplified by reverse transcription-polymerase chain reaction (RT-PCR) from human osteosarcoma cell line MG63 and cloned into pGEMT-Easy vectors. Then, using the recombinant plasmid as the template,the DNA encoding the fusion protein OPG-HSP 70 was amplified by PCR,and was inserted into prokaryotic expression vector pET-28a. Construct pET-28a-OPG-HSP 70 was used to transform into competent E.coli. BL21(DE3) which were induced by isopropyl B-D-thiogalactopyranoside (IPTG),and the fusion protein from above E.coli was collected. Sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) and Western-blotting were performed to identify OPG-HSP 70 fusion protein. The experiments of osteoclast inhibition and restraining inflammation were used to detect the bioactivity of fusion protein. RESULTS: ①The complementary DNA encoding full length OPG protein was obtained. OPG-HSP 70 fusion gene obtained in this experiment was successfully inserted into pET-28a vector. OPG-HSP70 fusion protein was expressed when transformed into E.coli.BL21(DE3). ②SDS-PAGE indicated that the fusion protein was large expressed at the molecular weight of Mr22 000, but there were no band in the total lysate of bacteria harboring pET-28a-OPG-HSP70 without IPTG induction group. ③Western-blotting indicated that the OPG-HSP70 fusion protein could specifically react with anti-human OPG monoclonal antibodies. ④The osteoclast inhibition test demonstrated that the fusion protein could reduce the number of osteoclast, and had the ability to inhibit bone absorptions in vitro. ⑤The experiment of restraining inflammation showed that the fusion protein could significantly reduce the inflammation of delayed type hypersensitivity (DTH) mice, which explained that HSP70 in fusion protein had the inflammation inhibitory bioactivity. CONCLUSION: OPG-HSP70 fusion protein is expressed in E.coli.BL21(DE3),and function study in vitro illustrates the bioactivity of fusion protein.

Feasibility of Detecting Bacterial Endotoxins in Low Molecular Weight Heparins Calcium Injection by TAL / 中国药房

OBJECTIVE:To explore the feasibility of detecting bacterial endoxins in low molecular weight heparins calcium injection LMWHC by tachypleus amebocyte lysate(TAL).METHODS:Bacterial endotoxins in samples were detected qualita?tively and quantitatively by kinetic turbidimetric assay and gel-clot method.RESULTS:Diluted to the concentration of25IU/ml,LMWHC had an obvious interference action to the TAL test,while not at the concentration of12.5IU/ml.The contents of bacterial endotoxins in all samples were less than0.01EU/IU.CONCLUSION:It is feasible to detect bacterial endotoxins in LMWHC by TAL test.

Problems in continuing medical education in grass-roots hospitals and solutions / 中华医院管理杂志

With the burgeoning of continuing medical education, grass-roots hospitals are confronted with quite a number of problems in the implementation of the task, such as a general rush into action with each department going its own way, incontrollable educational expenses that are beyond the hospitals, difficulties in developing programs at the hospital level because of the limitation of academic disciplines, and the buying and selling of credits. In view of the above problems, the authors put forward some suggestions for formulating a series of continuing medical education management strategies that both conform to the situation in our country and suit the needs of health professionals at grass-roots institutions.