The study of traditional Chinese medicine and its complex prescription gradually adopted the methods of serum pharmacology, which has become a new researching thought and a gained more attention. In this article, the background of relevant researches, methodological discussions, analysis of the advantages and problems of serum pharmacology were reviewed. The serum pharmacology, for its unique value, will certainly has a brilliant prospect in its application to the research of traditional Chinese medicine and its complex prescription.

The goal of this work is to understand the difference between vertical structure and horizontal structure on data management. We take adverse event report as an example to find out the relationship between the storage space and the factors related to the trail. From the study we find out the difference of storage space between horizontal structure and vertical structure goes up with the increase of the storage space of each adverse event record and the number of adverse event allowed to record in the horizontal structure and goes down with the increase of the happening rate and the average times of the adverse event. When using the most widely used study design, vertical structure always takes less storage space than the horizontal structure. This vertical structure database is more suitable for the data management than the horizontal structure database.

Objective To explore the changes of cytokines in myasthenia gravis children and the effect of glucocorticoid on the serum levels of them.Methods Forty cases of myasthenia gravis children were the observation group which was divided to group one (pre-glucocorticoid therapy)and group two (post-glucocorticoid therapy),and 20 cases of healthy children in the same period as the normal control group.The serum levels of cytokines INF-γ,TGF-β1,IL-10 and IL-18 of the observation group one and two and the normal control group were detected by ELISA and were compared between the observation group one and the normal control group and the observation group one and two.Results The serum levels of cytokine INF-γand IL-18 were higher and IL-10 and TGF-β1 were lower in the observation group than in the normal control group,the differences were statistically significant (t =4.45 ~16.72,P <0.01 ).Significant difference of INF-γ,TGF-β1 and IL-18 was found between the observation group one and two (t =8.12 ~10.68,P <0.01)and the sera level of IL-10 had no significant difference (P >0.05).Conclusions Cytokines INF-γ,TGF-β1,IL-10 and IL-18 are involved and probably play different roles in the pathogene-sis of myasthenia gravis in children;Glucocorticoid could affect the secretion of cytokines IFN-γ,TGF-β1 and IL-18 of myasthenia gravis children,which would ultimately to achieve the aim of interfering and con-trolling the clinical symptom of myasthenia gravis in children.

Objective The purpose of this study was to evaluate the disinfection effectiveness and properties of acrylic resin via microwave irradiation. Methods Forty acrylic resin base dentures fabricated in a standardized procedure were chosen and divided into Group A,B,C and D randomly. Group A,B and C were immersed in 200ml distilled water and submitted to microwave irradiation at 700W for 3,4,5 minutes individually. Group D was used as positive control. Bacteria specimens from each group were got for culture and numbers were calculated. Then the four groups were tested for the flexural strength and impact strength by universal test machine before and after microwave irradiation sterilization. Results The germicidal ratio of Group B was more than 90% ,while the ratio of group C was 100%. There was no significant difference in flexural strength and impact strength between group C and group D. Conclusions Microwave irradiation for 5 minutes at 700W produced sterilization of dentures contaminated with all bacteria whereas the denture strength is not affected. Microwave irradiation at 700W solution for 5 minutes was effective to sterilize acrylic resin base dentures.

Objective To investigate the proof of fetal cells passing through the placental barrier into the maternal peripheral blood to provide laboratory data for the non invasive prenatal gene diagnosis of genetic diseases. Methods A total of 22 samples of placental tissues delivered(male fetus: 12, female fetus: 11) were divided into two groups for parallel section. HE staining was used to find the distribution of fetal cells in chorionic villi. In situ hybridization (ISH) technique with SRY DNA probes was used to identify the existence of fetal cells in placental villi, particularly in intervillous space. Results Light microscope examination revealed that there were fetal cells that passed through the capillary endothelium of villi and trophoblast basement membrane in the placental tissue sections of the 22 samples. ISH with SRY DNA probe also revealed that there were positive signals in the capillary of villi, at the edge of trophoblast basement membrane and in intervillous space in the placental tissue sections of the 12 placentas, but no signals were found in 10 female placentas. Conclusion This study demonstrates that the distribution of the fetal cells in the chorionic villi and intervillous space could be identified. The detection of fetal DNA in maternal circulation is one of the candidate approaches for non invasive prenatal gene diagnosis.

Kruppel-like factor 6 (KLF6) was reported as tumor suppressor in multiple cancers. However, loss of chromosomal locus spanning KLF6 is relatively infrequent in previous published studies. To explore the role of KLF6 in hepatocellular carcinoma (HCC), we examined the gene for expression change, loss of heterozygosity (LOH) and mutation in 26 HCC samples. The expression levels of KLF6 were significantly down-regulated in HCCs, as detected by qRT-PCR. LOH occurred in 11 (52%) of 21 tumors, and all the samples with LOH showed KLF6 down-regulation. The mutational frequency was 24%, and sequence changes located in activation domain of KLF6. Furthermore, MTT assay showed a significant antiproliferative effect of the wt KLF6 transfected in HepG2 hepatoblastoma cells. Fluorescence-activated cell sorting analysis revealed that KLF6 could induce apoptosis. These findings indicate that deregulation of KLF6, together with genetic abnormalities of allelic imbalance and mutations, may play a role in HCC pathogenesis.

Objective To prepare quality control samples for St.Louis encephalitis virus(SLEV)molecular detection by constructing pseudovirus containing target sequences of SLEV.Methods According to the principles of armored RNA technique, the prM gene sequence of SLEV was cloned into the prokaryotic expression vector to generate recombinant plasmid pSE380-MS2-SLEV.Then, recombinant E.coli transformed with the corresponding plasmid was induced with IPTG to produce recombinant pseudovirus particles.The particles were purified by chloroform and further characterized by double enzyme digestion and transmission electron microscopy.The temperature sensitivity experiments and quantitative RT-PCR were performed to validate the potential of these pseudovirus particles as quality control samples.Results PCR amplification and sequencing analysis confirmed that the prM gene sequence of SLEV was cloned into vector pSE380-MS2.Transmission electron microscopy showed that homogenous spherical particles with a diameter of about 25 nm were produced upon IPTG induction.The SLEV genomic RNA within the pseudovirus particles was resistant to DNaseⅠand RNase A digestion, and remained stable for 20 days at 37℃.These samples were validated with quantitative RT-PCR for SLEV.Conclusion The RNase-resistant and stable pseudovirus particles containing prM fragment of SLEV are constructed successfully, which can be used as positive quality control samples for RNA extraction and molecular detection.

Objective To investigate the expression and genetic alterations of KLF6 in hepatocellular carcinoma (HCC) and explore their functional mechanisms in the oncogenesis and development of HCC. Methods Real-time quantitative-PCR, direct sequencing and LOH approaches were used to detect KLF6 genetic abnormalities in HCC. The experiment had 2 groups, an experimental group and a control group. In the experimental group, the transfected plasmid pcDNA3.0 was recombined with KLF6 and tranfected into HCC HepG2 cells. MTT, flow cytometry and Western blotting were used to observe the effect of anti-oncogene wild type KLF6 on HepG2 cells by transgenic method for 48 h.Results Expression levels of KLF6 were significantly downregulated in HCCs(P＜0. 01), as detected by qRT-PCR. LOH occurred in 11 (52%) of the 21 tumors, and all the samples with LOH showed KLF6 down-regulation. The mutational frequency was 29%, and sequence changes located in activation domain of KLF6. Meanwhile, MTT assay showed a significant antiproliferative effect of the transfected wtKLF6 on HepG2 cells(42.7%, P＜0.05). Fluorescence-activated cell sorting analysis revealed that KLF6 induced apoptosis. Conclusion The deregulation of KLF6 together with genetic abnormalities of allelic imbalance and mutations may play an important role in HCC pathogenesis.

Objective To construct p66shc gene interfering lentivirus vectors recombination and transfect it to 293T cells ,RNA interfering was carried out to induce p66shc gene silence ,so as to provide basis for further study of the p66shc function .Methods Screening of three RNA targets which were named after p66shc‐shc1 ,p66shc‐shc2 ,p66shc‐shc3 ,cloned into the pLenR‐GPH vec‐tor ,which contained green fluorescent protein(GFP) and transformed into DH5αcells .The positive clone were picked out for right sequencing and transfected to 293T cells with pRsv‐REV ,pMDlg‐pRRE ,pMD2G .The expression of GFP in inverted fluorescence microscope confirmed the virus packaging success .Fluorescence quantitative PCR and Western blot technology were used to investi‐gate the expression of p66shc at the molecular and protein levels ,p66shc‐shc1 target of effective silencing p66shc gene was selected to prepare for subsequent tests .Results The shRNA lentivirus vector was constructed which could express p66shc and was trans‐fected into 293T cells successfully .Fluorescence quantitative PCR and Western blot technology were used to investigate p66shc gene silence by RNA interference .Conclusion The lentivirus RNAi vector of targeted expression p66shc could induce p66shc gene si‐lence at the molecular and protein levels after transfected into 293T cells by RNA interference .

ObjectiveTo develop an antibody-array system for multiple detection of antibodies against Japanese B encephalitis virus (JEV),Tick-borne encephalitis virus (TBV),Dengue virus ( DENV ),West Nile virus (WNV),Western equine encephalitis virus (WEEV) and East Equine encephalitis virus (EEEV).MethodsRecombined antigens were spotted on array as capture antigens.Specific antibodies were detected by using a sandwich ELISA format.Rabbit antiserum was employed to select and confirm the specificity of antigens and to optimize the conditions of the assay.The detection efficiency of the system was validated by 40 clinical suspected serum samples and compared with the relative ELISA assays.ResultsEleven recombined antigens were selected as diagnostic antigens with high specificity.Better detection could be achieved when scale of antigen concentrations were within 0.125-0.900 mg/ml and the serum dilutions were 1:100-1:1000.When detecting the 26 clinical suspected TBE serum samples,20 were IgG positive (76.9％),and 17 were IgM positive (65.3％) which was 96.1％ and 84.6％ consistent with the relevant ELLSA tests,the 8 clinical suspected JEV serum samples,4 were IgG positive (50.0％),and 5 were IgM positive (62.0％),which was 86.3％ and 90.1％ consistent with the relevant ELLSA tests.As for the 22 DEN serum samples,13 were IgG positive (60％) and 15 were IgM positive (68％) which was 85％ and 93％ consistent with ELISA.The specificity of the assay was 100％ and the sensitivity was higher than the relative ELISAs.ConclusionThe developed antibody-array is highly specific and reliable,which could be used for the detection of antibodies against the 6 arboviruses.