Objective To clone and express prokaryotic recombinant plasmid of nucleoside triphosphate hydrolase (NTPase) gene of Toxoplasma gondii, and analyze its antigenicity. Method NTPase gene was amplified by PCR from RH strain of T. gondii and cloned into pGEM-T Easy vector. Positive clones were screened and identified by BglⅡ, HindⅢ digestion and sequenced. The target gene was then subcloned into prokaryotic expression vector pBAD-HisB and transformed into E. coli BL21(DE3). The expressed recombinant protein was purified with Ni-NTA agarose and further analyzed by sodium dodecyl sulfate polyacrylamide gel electropheresis (SDS-PAGE) and Western blotting. Results NTPase-Ⅱ gene was specifically amplified, and the homology of DNA sequence was 100% to that in the GenBank. SDS-PAGE showed that the recombinant NTPase protein with correct molecular weight was expressed highly in E.coli BL21(DE3). Western blotting testified that the purified recombinant protein could be specifically recognized by mouse serum immunized with T. gondii and mouse anti-recombinant protein serum. Conclusion The NTPase-Ⅱ gene has been cloned and expressed in E.coli BL21(DE3), and the purified protein of NTPase-Ⅱ gene displays a specific antigenicity.

TCM syndrome differentiation system (TCM-SDS) possesses a complex nonlinear characteristic, which determines the falt that the evaluation of TCM efficacy should not be simplified to a causal or linear relationship. Previous selected indexes and methods for evaluating curative effect of Child's pneumonia have their limitations. This paper mainly introduces several thoughts and methods formed in the exploratory researches, employing data mining technology in this field. We believe that efficacy evaluation research of Child's pneumonia on the basis of standardized study of TCM-SDS is a scientific work. Applying data mining method in processing enormous and multilayer information of TCM differentiation has strong advantages in screening evaluating indexes of Child's pneumonia. Meanwhile, some major concerning technical categorizations and implementing strategies for the research are also introduced.
Algorithms ; Automatic Data Processing ; methods ; Child ; Child, Preschool ; Data Interpretation, Statistical ; Diagnosis, Differential ; Drugs, Chinese Herbal ; therapeutic use ; Female ; Humans ; Infant ; Male ; Medicine, Chinese Traditional ; methods ; standards ; Outcome Assessment (Health Care) ; methods ; statistics & numerical data ; Phytotherapy ; Pneumonia ; diagnosis ; drug therapy

Objective To investigate the clinical features and pathogenic genes of a familial hypokalemic periodic paralysis ( HOKPP).Methods PCR amplification and DNA sequencing were used to screen candidate genes of the HOKPP family members (CACNA1S, SCN4A, KCNE3), and the clinical features were carefully analyzed at the same time.Results The sequencing analyses of the SCN4A gene in the proband identified three nucleotide sequence mutations, which influenced the amino acid sequence of the skeletal sodium channel.One of the mutations was identified as a C/T heterozygous pattern at the 2111th nucleotide position in exon 13, resulting in a change from Thr to Met at the 704th amino acid position of the sodium channel protein.All affected patients carried the Thr704Met mutation, whereas unaffected family members did not.Clinical symptoms in this family followed an autosomal dominant inheritance pattern.Muscles weakness, pain and hypokalemia in the period between attacks were seen in all patients.Paralytic symptoms occurred early, lasted longer and recurred frequently, while cold was the main predisposing factor.With the progress of the disease, patients represented persistent weakness and atrophy in proximal muscles.Conclusions Mutation (Thr704Met) in the SCN4A gene should be responsible for this family.This mutation causes severe HOKPP and progressive muscle atrophy.

To analyze and compare the protective effects of active components in different ethyl acetate extracts (EAEEPs) from Eclipta prostrate, in order to study the comparison of materials bases protecting normal human bronchial epithelial (NHBE) cells. The MTT assay was taken to compare the protective effect of different EAEEPs on cigarette smoke extracts (CSE) -induced NHBE cells. The ultra-performance liquid chromatography (UPLC) was applied to analyze the content of phenolic acid, coumaric grass ether and flavonoid in EAEEPs. According to the results, all of the eight EAEEPs (0-200 mg x L(-1)) showed certain protective effect on NHBE cells, with statistical difference. Specifically, the total mass of EAEEP VII (89.15 mg x L(-1)) and EAEEP VIII (57.44 mg x L(-1)), which showed the strongest activity, was not the highest, while EAEEP III (132.25 mg x L(-1)) displayed the highest total mass. In the combination with the "component structure" theory, the analysis showed a significant difference in the mass structure among phenolic acid, coumaric grass ether and flavonoid in EAEEP VIII and EAEEP VIII, which were 1.0: 1. 0: 0.5 and 1.0: 1.9: 0.8, respectively. The results suggested a specific optimal "component structure" relationship may exist in EAEEP, which could provide reference for the material base study and quality control.
Bronchi ; cytology ; drug effects ; Drugs, Chinese Herbal ; chemistry ; isolation & purification ; pharmacology ; Eclipta ; chemistry ; Epithelial Cells ; cytology ; drug effects ; Humans ; Protective Agents ; chemistry ; isolation & purification ; pharmacology ; Tobacco Smoke Pollution ; adverse effects

Objective To study the characteristics of Genetic typing and the antibiotic susceptibility testing of strains from Pseudomonas aeruginosa keratitis patients.Design Experimental study,Participants 23 eyes of 23 patients of Pseudomonas aeruginosa keratitis.Methods The genomic was extracted and amplified with PCR.The PCR products were purified and sequenced.The results were registered in MIST web Antibiotic susceptibility testing were performed in theses strains,Main Outcome Measures Sequence types and antibiotic susceptibility.Results The isolates were resolved into 20 STs.Two lineages were identified.MIC test showed that strains were more susceptible to aminoglycosides,The activity of quinolones and cephalosporin were higher than that of aminoglycosides.Conclusion MIST can determine homology of the strains from Pseudomonas aeruginosa keratitis by clustering results. There is no finding about relationship between Genetic typing and drug resistance.

Traditional Chinese medicine(TCM) is a complex system, featured with integrity and characteristics. Structural component TCM is a well-organized integrity of traditional Chinese medicine, reflecting multi-component integration effect of TCM. It gives us a new view on the material basis of TCM. Currently, conventional researching strategies are not enough to deal with the relationship between material basis and efficacy, multi-composition, multi-targets, and multi-section mechanism. Post-genome area gives a birth to bioinformatics, which involves systematic biology, different levels of omics, corresponding mathematics and computer techniques. It increasingly becomes a powerful tool to understand complicated system and life essential laws. Research ideas, methods. and knowledge of data mining technology of bioinformatics combined with the theory of structural components of Chinese medicine bring a new opportunity for developing structural components of Chinese medicine, systematically exploring the essence of TCM and promoting the modernization of TCM.
Animals ; Biomedical Research ; Computational Biology ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Humans

Hydrolytic amino acids were extracted by acid hydrolysis method, then derivatized with phenyl isothiocyanate (PITC). And the samples were analysed by HPLC on an Ultimate Prime C18 (4.6 mm x 250 mm, 5 microm) column with gradient elution of 0.1 mol x L(-1) sodium acetate buffer solution (adjusted to pH 6. 5)-acetonitrile (93:7) (A) and acetonitrile-water (8:2) (B) at a flow rate of 1.0 mL x min(-1). Column temperature was 40 degrees C and the detected wavelength was 254 nm. Amino acids derivative solution remained stable in 36 hours. The response was linear for 16 amino acids with a correlation coefficient r > 0.999 5. The average recoveries were 98.01% -101.8%. The method is reliable with good accuracy and repeatability, which is useful for the determination of amino acids in Galli Gigerii Endothelium Corneum.
Amino Acids ; analysis ; Animals ; Chickens ; Chromatography, High Pressure Liquid ; Chromatography, Reverse-Phase ; Endothelium ; chemistry ; Gizzard, Avian ; chemistry

Objective To investigate the effect of adenovirus-bone morphogenic protein 9 ( Ad-BMP9 ) on osteogenic differentiation of immortalized calvarial mesenchymal progenitor cells ( iCALs ) .Methods iCALs were infected with adenoviral vectors encoding BMP-9 or green fluorescent protein ( GFP) and the early osteogenic differentiation was assessed by detecting alkaline phosphatase (ALP) activity after being cultured for 3, 5 and 7 days.14 days after infection, alizarin red S staining was performed to study the formation of osteogenic calcium nodules .The expression of osteogenic marker genes Runx2 and OCN was assessed by quantitative real-time ( RT )-PCR and Western blotting .Results Significant increases in ALP activity and in the expressions of Runx 2 and OCN were detected in BMP-9 treated iCALs compared with GFP-treated cells(P<0.05).Meanwhile, alizarin red S staining showed that more mineralized nodules were found in the BMP-9 induced group .Conclusion BMP-9 can promote the osteogenic differentiation of iCALs .

Objective To obtain scientific data for control of iodine deficiency disorders(IDD) by reviewing the surveillance information of school children intelligence quotient(IQ) after the implementation of universal salt iodization. Methods One thousand five hundred and eighty children were selected from 11 primary school in Dalian city of Liaoning province during 2006 to 2009. IQ was measured by Combined Raven Test-C2(CRT-C2) in China. Groups of IQ were classified as outstanding(≥ 130), excellent (120- 129), above average (110- 119), average (90 - 109), below average(80 - 89), margin(70 - 79), low(≤69). Urinary samples of children were collected randomly. Urinary iodine were determined by As-Ce catalytic spectrophotometry. The growth characteristics of IQ were analyzed according to surveillance year and born year. Results The average IQ of children aged 8-10 were 110.4 ± 14.0,112.5 ± 12.4,117.2 ± 11.4,116.2 ± 12.6, respectively, increased year by year from 2006 to 2009. Its average annual increase from 2007 to 2009 were 2.1,3.4,1,9 compared with the IQ in 2006 respectively. The medians of urinary iodine were 224.7,266.7,222.1 μg/L from the year 2007 to 2009, respectively, which were all between 200 - 300 μg/L and can be classified as more than adequate level. The average IQ of children born during the year of 1994 to 2000 were 106.7 ± 13.0,108.1 ± 13.9,108.5 ± 13.4,111.3 ± 14.3,113.6 ± 12.5,115.3 ± 12.3,119.8 ± 11.2, respectively. Its average annual increase from 1995 - 2000 were 1.4,0.9,1.5,1.7,1.7,2.2 compared with the IQ born in 1994 respectively. The ratio of IQ in margin group and low group were all below 2% ; the ratio was increasing in excellent group and outstanding group and decreasing in average group and below average group significantly year after year(x2 = 52.471,34.329,66.483,11.148, all P<0.01). Conclusions Universal salt iodization improves IQ scores. IQ index should be brought into the surveillance project and put in use in IDD control.

The aim of this study was to investigate the role of Delta-like 1 (Dll1) in differentiation and antigen pre-sensation of mouse bone marrow-derived dendritic cells (DCs). In the presence of granulocyte macrophage colony stimulating factor (GM-CSF) and interleukin 4 (IL-4), mouse bone marrow cells were co-cultured with OP9-Dll1 and OP9-GFP cell lines respectively. After 8 days, the immature DCs were stimulated with tumor antigen. The surface molecules of the activated DCs including MHC II, CD80 and CD86 were analyzed by flow cytometry. Levels of IL-12 and IL-10 in the culture supernatant were detected by ELISA. In addition, the proliferation of T-cells co-cultured with DCs was analyzed by FACS through mixed T-lymphocyte reaction. The results showed that compared with OP9-GFP, the bone marrow cells co-cultured with OP9-Dll1 produced significantly more CD11c(+) DCs (p < 0.05), and possessed higher levels of surface molecule expression including MHC II, CD80 and CD86 after tumor antigen stimulation. The DCs secreted higher level of IL-12 (p < 0.05) and less IL-10 (p < 0.01). They also resulted in significantly stronger T-cell proliferation response. It is concluded that Dll1 can promote the differentiation of DCs from mouse bone marrow cells and enhance their antigen presentation capacity.
Animals ; Antigen Presentation ; immunology ; Bone Marrow Cells ; cytology ; Cell Differentiation ; immunology ; Cells, Cultured ; Dendritic Cells ; cytology ; immunology ; Granulocyte-Macrophage Colony-Stimulating Factor ; pharmacology ; Intercellular Signaling Peptides and Proteins ; metabolism ; Interleukin-4 ; pharmacology ; Male ; Mice ; Mice, Inbred C57BL