To increase detection sensitivity and specificity on hepatitis C virus (HCV) is vital for prevention and controlling of the disease. To establish a more reliable detection method for HCV diagnosis, the full gene fragment of ns3 (non-structural protein of HCV) from recombinant plasmid of J6/JFH1 2a was amplified and then connected into the pET-28a prokaryotic expression vector, and the latter was subsequently transformed into Escherichia coli BL21 (DE3) to have the target protein expression. As a result, a protein with a molecular weight of 72 kDa was obtained and visualized in 10% SDS-PAGE. The purified NS3 protein was used as immunogen to inoculate BALB/c mice and the sera was collected after the fourth immunization. The antibody titer of serum is determined to be about 1:256000 with ELISA. Western blotting and indirect immunofluorescence analysis showed that the mouse polyclonal antibody could react specifically with the native NS3 protein in Huh 7.5.1 cells infected with HCV. These findings may provide basis for further preparation of monoclonal antibodies against NS3 and the development of related detection kit.
Animals ; Antibodies, Viral ; immunology ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; Hepacivirus ; Mice ; Mice, Inbred BALB C ; Plasmids ; Viral Nonstructural Proteins ; biosynthesis ; immunology

Tree shrews get more and more concerns due to many of its physiological , biochemical and anatomical characteristics similar to those of human beings .Therefore, tree shrews models of human diseases such as viral diseases , neurological diseases and tumors attract more and more attention of researchers .In this article we will review the recent ad-vances in application of tree shrew models in research on human viral diseases .

Viral hepatitis is a major liver disease caused by virus infection .Viral hepatitis is popular in China , mainly caused by hepatitis B and hepatitis C viruses .Experimental animal model is a necessary platform for the research on mechanism of viral infection and pathogenicity , for treatment and vaccine development .Up to date, a great progress in the development of viral hepatitis animal models has been achieved in spite of the most of findings are limited to hepatitis B and C.Here, we summarize the recent findings of viral hepatitis animal models , focusing on the tree shrew animal model and its modeling strategy .

Objective To investigate expression and regulatory mechanism of phosphatase and tensin hemology deleted on chromosome ten gene (PTEN) and cyclooxygenase-2 (COX-2) in human myeloid leukemia cells.Methods Thirty patients was collected from the First Hospital of Baoding and Second Affiliated Hospital of Hebei Medical University,including 10 chroni myeloid leukemiac (CML)patients in chronic phase (CML-CP),10 CML patients in blast crises (CML-BC) and 10 normal controls.The recombinated adenovirus containing green fluorescent protein (GFP) and PTEN ( Ad-PTEN-GFP) or empty vector (Ad-GFP) was transfected into human CML K562 cells.The growth of K562 cells and cell adhesion ability was observed by 3-[4,5-dimethyl-2-thiazolyl]-2,5-dip (MTF) assay; PTEN and COX-2 messenger ribonucleic acid (mRNA) levels were detected by real-time fluorescent relative-quantification reverse transcriptional PCR (FQ-PCR).PTEN and p-Akt protein levels were detected by western blot,and COX-2 protein were measured with cytochemical staining.ResultsThe mRNA expression levels of PTEN in CML-BC patients (0.022 ±0.021 ) were lower than CML-CP patients (1.134 ± 1.124) and normal control ( 1.059 ±0.595).The mRNA expression levels of COX-2 in CML-BC patients (0.761 ±0.418) were higher than CML-CP (0.211 ± 0.158) and normal control (0.165 ± 0.152).The growth of K562 cells was suppressed markedly,and the maximum growth inhibition rate was 38.67％ ± 4.30％ after transfected with PTEN gene,which was higher than Ad-GFP group (5.34％ ±0.31％,t =13.39,P ＜0.01 ).COX-2 mRNA expression levels in Ad-PTEN-GFP(0.013 ± 0.001 )were significantly lower than Ad-GFP group (0.199 ±0.018) and untransfected group (0.217 ± 0.021,F =499.45,P ＜ 0.01 ),and p-Akt as well as COX-2protein were also down-regulated after K562 cells transfected ( MOI =200) with wild type PTEN in three days.ConclusionPTEN may inhibit proliferation and adhesion ability of leukemia cell in myeloid leukemia via down-regulating COX-2 expression.

With the in-depth exploration of all stages in early-stage embryos, in particular zygotic genome activation and first cell lineage differentiation, researchers have found that early embryonic epigenetics follows a strict pattern of temporal and spatial modification. Previous studies have determined the inhibitory effect of H3K9me3 and H3K27me3 on genomic expression, and found that they are involved in many core biological events in the genome such as chromatin reprogramming, genomic imprinting, maintenance of embryonic stem cell pluripotency and somatic cell nuclear transfer, though the detailed molecular mechanism has remained elusive. From the point of developmental biology and epigenetics, this article has expounded the research progress on the methylation of H3K9 and H3K27 histones in early-stage embryos, which may provide a clue for the complex mechanism of embryonic development and improvement of culture method for embryos in vitro.
Chromatin ; Embryonic Development ; Epigenesis, Genetic ; Female ; Gene Expression Regulation, Developmental ; Histones/metabolism* ; Humans ; Methylation ; Pregnancy

Objective: To evaluate the new compensation mechanism for primary healthcare institutions in Zhejiang province, in terms of fairness, performance, incentive mechanism and sustainability in pilot areas. Methods: Evaluation indicators were constructed based on stakeholder theory, fairness theory, expectation theory and sustainable development theory.Focus group interviews were conducted with stakeholders and quantitative data were collected through questionnaires. Meanwhile, the financial compensation, income and expenditure and work equivalent data were collected from such institutions of the four pilot areas, with quantitative data subject to descriptive analysis. Results: This study found the reform used reasonable proportion of funds allocated(the proportion of basic salary for employees was lower than 50%)and adjustment factors(1.0-1.8)of different primary healthcare institutions to guarantee the fairness of the reform; the increase of work equivalents(the per capita work equivalents of medical staff in pilot counties had increased from 38.435 million in the previous year to 42.590 million work equivalents)reflected the performance outcomes of the reform. The incentive and sustainability of the reforms were the weak parts. These were mainly due to the fact that the internal distribution system of primary healthcare institutions failed to make corresponding reforms. Conclusions The reform of the compensation mechanism based on the equivalent method has changed medical staff′s perception of the distribution of funds. The principle of" more pay for more work" and the use of information technology to capture work equivalents have improved the enthusiasm of primary medical staff and the operational efficiency of these institutions, thus, making reform generally scientific and reasonable.

Objective To investigate the differences in the expression of PHF20 and Bax and their correlations in non-small cell lung cancer before chemotherapy.Methods An immunohistochemical method was used to detect the expression of PHF20 and Bax in non-small cell lung cancer,and to analyze the clinical significance of PHF20 and the possible correlation between PHF20 and Bax proteins.Results PHF20 protein is expressed in the cytoplasm of non-small cell lung cancer cells.Moreover,it is highly expressed in squamous cell carcinoma,less expressed in adenocarcinoma,and closely related with cell differentiation,TNM staging,and lymph node metastasis.The expression of PHF20 and Bax was positively correlated with squamous cell lung carcinoma.Conclusion The expression of PHF20 in non-small cell lung cancer is closely associated with tumor progression and the expression of Bax.PHF20 may be a new target for the treatment of non-small cell lung cancer.

Higher alcohols are one of the main by-products of Saccharomyces cerevisiae in brewing. High concentration of higher alcohols in alcoholic beverages easily causes headache, thirst and other symptoms after drinking. It is also the main reason for chronic drunkenness and difficulty in sobering up after intoxication. The main objective of this review is to present an overview of the flavor characteristics and metabolic pathways of higher alcohols as well as the application of mutagenesis breeding techniques in the regulation of higher alcohol metabolism in S. cerevisiae. In particular, we review the application of metabolic engineering technology in genetic modification of amino transferase, α-keto acid metabolism, acetate metabolism and carbon-nitrogen metabolism. Moreover, key challenges and future perspectives of realizing optimization of higher alcohols metabolism are discussed. This review is intended to provide a comprehensive understanding of metabolic regulation system of higher alcohols in S. cerevisiae and to provide insights into the rational development of the excellent industrial S. cerevisiae strains producing higher alcohols.
Alcoholic Beverages ; Alcohols/analysis* ; Fermentation ; Saccharomyces cerevisiae/metabolism* ; Saccharomyces cerevisiae Proteins/metabolism*

OBJECTIVE：To study the effects of berberine on mic e macrophage polarization based on TLR 4-MyD88-NF-κB signaling pathway. METHODS ：Using mice RAW 264.7 macrophage as the object ，atorvastatin calcium as positive control ， inflammatory cell model was induced by lipopolysaccharide （LPS）；ELISA method was used to detect the contents of TNF-α，IL-6 and NF-κB in cell culture medium after treated with low，medium and high doses of berberine （5，10，20 μmol/L）for 24 h. The real-time fluorescence quantitative PCR was conducted to determine the mRNA expression of TLR 4 and MyD 88 in cells. Western blotting assay was used to detect the protein expression of TLR 4，MyD88，iNOS and CD 206 in cells. RESULTS ：Compared with blank control group ，the contents of TNF-α，IL-6 and NF-κB in cell culture medium，mRNA expression of TLR 4 and MyD 88， protein expression of TLR 4，MyD88 and iNOS in cells were increased significantly in LPS induction group （P＜0.05）. Compared with LPS induction group ，the contents of TNF-α and IL-6，mRNA and protein expression of TLR 4 and MyD 88 in atorvastatin calcium group ，berberine medium-dose and high-dose groupsas well as the content of NF-κ B and protein expression of iNOS in administration groups were decreased significantly ， while the content of NF-κB in berberine high-dose group was significantly lower than atorvastatin calcium group （P＜0.05）. The protein expressions of CD206 in atorvastatin calcium group and berberine high-dose group were increased significantly ，while the protein expression of CD 206 in berberine high-dose group was significantly higher than atorvastatin calcium group （P＜0.05）. CONCLUSIONS ：Different doses of berberine can intervene in mice macrophage polarization to different extents ，the mechanism of which may be associated with the regulation of TLR4/MyD88/NF-κB signaling pathway.