Objective To establish a method for the isolation of Zika virus and to gather experi-ences for viral isolation. Methods Suckling mice at age 1-3 days were inoculated with serum samples posi-tive for Zika virus through intracranial injection. All mice were sacrificed 6 days after the injection. Viral nu-cleic acids were extracted from brain, heart, liver, spleen, lung, kidney, muscle, skin and intestine tissue samples and analyzed by real-time RT-PCR. The supernatants of brain tissues positive for Zika virus were used for subculturing. Nested PCR was performed to amplify the NS5 gene of the isolated virus. The se-quences of NS5 gene were analyzed by using MEGA6. 0 software. Results All of the tissue samples were positive for Zika virus. Higher viral loads were detected in heart and brain tissue samples with cycle thresh-old (Ct) values of 24. 4 and 25. 3, respectively. The second generation of Zika virus was identified in suck-ling mice brain tissues 2 days after infection by using real-time RT-PCR. The amplified product of nested PCR was 972 bp in length. Sequencing analysis showed that the isolated Zika virus ( GDZ16002 strain) be-longed to the Asian lineage. Conclusion A strain of Zika virus was successfully isolated in China by using intracranial injection via a suckling mouse model. The isolated Zika virus belonged to the Asian lineage.

Objective To screen protective antigen genes and construct the T7 phage display library from adult worms of Schistosoma japonicum.Methods Total RNA was extracted from adult worms of S.japonicum by Trizol reagent and mRNA was isolated from the total RNA.The ds cDNA was synthesized by reverse transcription using random primer.Directional EcoRⅠ/HindⅢ linkers were ligated into the ends of ds cDNA and the ds cDNA was digested with EcoRⅠand HindⅢ,which resulted in ds cDNA with EcoRⅠand HindⅢ adhering ends.The digested ds cDNA fragments longer than 300 bp in length were fractionated and ligated into T7 Select 10-3b vector.After packaging in vitro,the T7 Select 10-3b vector was transformed into BLT5403 to construct the T7 phage display cDNA library.Plaque assay and PCR were used to evaluate the library.Seven known objective genes of S.japonicum were screened by PCR to detect the representation of the library.Result Primary library capacity was 4.98?106 pfu,and the titer of amplified library was 3.85?1011 pfu/mL.The PCR identification result of 96 clones picked at random showed that recombination rate was 93.8%,in which 95.6% inserted cDNA fragments were longer than 300 bp in length.All the seven known objective genes of S.japonicum were amplified from the library.Conclusion The T7 phage display library from adult worms of Schistosoma japonicum was constructed.

Objective To look for the genes of Schistosoma japonicum related to the Schistosoma-resistance of Microtus fortis.Methods The fresh sera of Microtus fortis were used to screen a T7 phage display cDNA library from worms of Schistosoma japonicum established in our lab.The positive clones were sequenced and functionally analysed through bioinformatics.Results The specific phages binding to the sera of Microtus fortis were enriched 857-fold after three rounds of biopanning,and 58 positive clones picked at random were sequenced and 10 ESTs were obtained.BLASTn results showed that 7 ESTs had 99%-100% similarity to the genes of Shistosoma japonicum reported in GenBank and 1 EST had 82% similarity to a zinc finger protein encoden gene from Pan troglodytes.The results of these ESTs function prediction indicated most of them were involved in the regulation of gene expresion of Schistosoma japonicum.Conclusions Several target genes of Schistosoma japonicum related to the Schistosoma-resistance of Microtus fortis are obtained and those would lay foundation to expatiate the native resistance mechnism of Microtus fortis to Schistosoma japonicum.

OBJECTIVETo study the anticancer effect in vitro and in vivo and mechanism of DHA compound.

METHODSCervical cancer cell line HeLa cells, glioma cell line U251 cells and mouse hepatoma H(22) tumor were used in this study. Transmission electron microscopy and fluorescence microscopy were used to observe the morphological changes of cell apoptosis. Western blot was used to detect the expression of caspase-3. RT-PCR was used to determine the effect on Bcl-2 and Bax mRNA transcription in U251.

RESULTSAntitumor effect was observed in vivo and in vitro. Typical morphological changes were seen in cancer cells. The level of caspase-3 was significantly increased and the content of Bcl-2 mRNA was decreased significantly, while the content of Bax mRNA was significantly increased in the U251 cells after treatment with DHA compound.

CONCLUSIONDHA compound can inhibit the growth of some types of tumors and the increase of caspase-3 and Bax mRNA and decrease of Bcl-2 mRNA may be involved in its mechanism of action.

Animals ; Antineoplastic Agents ; pharmacology ; Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Docosahexaenoic Acids ; pharmacology ; Glioma ; pathology ; HeLa Cells ; Humans ; Liver Neoplasms, Experimental ; metabolism ; pathology ; Male ; Mice ; Neoplasm Transplantation ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism ; RNA, Messenger ; metabolism ; bcl-2-Associated X Protein ; genetics ; metabolism

Objective To construct a T7 phage display cDNA library from the lung of Microtus fortis for further screening the schistosomiasis-resistence-related genes of Microtus fortis. Methods mRNA was isolated from total RNA extracted from the lungs of Microtus fortis by TRIzol reagent, and was used to synthesize double strain cDNA by the reverse transcription. Then the double strain cDNA was given with EcoRⅠ and Hind Ⅲ adhering ends by ligation with the directional EcoRⅠ/Hind Ⅲ linkers and digestion with EcoRⅠ and Hind Ⅲ. The double strain cDNA fragments longer than 300 bp in length were fractionated by the Mini Column, and ligated into the T7 Select 10-3b vector with EcoRⅠ and Hind Ⅲ adhering ends. After packaging in vitro, the recombinant T7 Select 10-3b was transformed into BLT5403 to construct a T7 phage display cDNA library. Results The library constructed here contained 1.5?106 clones and the titer of the amplied library was 1.1?1012 pfu/ml. The PCR identification results of 100 clones picked at random showed that 91% clones were recombinant and 90% of recombinant clones contained cDNA fragments longer than 300 bp in length. Conclusion A T7 phage display cDNA library from the lung of Microtus fortis is successfully constructed.

OBJECTIVETo explore the feasibility of multi-dimensional and comprehensive evaluation on the quality of life among rural elderly population in Anhui province.

METHODS5652 rural elderly people aged above 65 in Anhui province were selected by cluster sampling method and were studied by cross-sectional study through a questionnaire on health information. The quality of life was evaluated by comprehensive evaluation method.

RESULTSThe total score of satisfactory quality of life in the studied rural elderly people was 0.1432 +/- 0.5170, while not satisfied was -0.2521 +/- 0.6081, with significant difference between the two groups (F = 666.221, P < 0.0001). There was positive correlation between subjective satisfaction and total score of quality of life, with r(s) = 0.345 (P < 0.0001). The results of logistic regression analysis between comprehensive index of quality of life and subjective satisfaction indicated that filial piety, income, sleeping condition, chronic disease, nutrition status, economic dominance in the family, amusement activities etc. were important factors influencing the quality of life.

CONCLUSIONIt was feasible to evaluation on the quality of life by comprehensive evaluation method.

Activities of Daily Living ; Aged ; Aged, 80 and over ; China ; Evaluation Studies as Topic ; Female ; Health Status ; Humans ; Male ; Personal Satisfaction ; Quality of Life ; Regression Analysis ; Rural Health ; standards ; statistics & numerical data ; Socioeconomic Factors ; Surveys and Questionnaires

OBJECTIVETo investigate the chemical constituents in leaves of Ilex centrochinensis and their antitumor bioactivity.

METHODVarious chromatography techniques such as column chromatography on silica gel, Sephadex LH-20 and preparative HPLC were used to isolate and purify the compounds and their structures were identified by spectral data and physicochemical properties. Their antitumor effect was tested by MTT method.

RESULTTen compounds were isolated and identified as 1,4-benzenediol (1), (2S)-5,4'-dihydroxy-7,3'-dimethoxyflavan(2), (2S)-5,4'-dihydroxy-7-methoxyflavan (3), kaempferol (4), quercetin (5), naringenin (6), ursolic acid (7), uvaol (8), oleanolic acid (9) and beta-sitosterols (10).

CONCLUSIONCompounds 1-5, 7, 8 were isolated from the species for the first time, among which compounds 1-3 were isolated from the Ilex genus for the first time. Compounds 2 and 3 showed strong cytotoxic activity against Huh7 cell lines with IC50 values of 8.98, 13.04 mg x L(-1), respectively. Compounds 7-9 exhibited weak cytotoxic activity against Caco-2 cell lines with IC50 values of 28.52, 38.28, 33.04 mg x L(-1), respectively.

Antineoplastic Agents ; chemistry ; pharmacology ; Caco-2 Cells ; Cell Line, Tumor ; Cell Survival ; drug effects ; Dose-Response Relationship, Drug ; Humans ; Ilex ; chemistry ; Inhibitory Concentration 50 ; Plant Extracts ; chemistry ; pharmacology ; Plant Leaves ; chemistry ; Plants, Medicinal ; chemistry

OBJECTIVE: To develop the visual uroflow scale (VUS), analyze the relationship of VUS score and index of free uroflowmetry, assess urination function preliminarily and improve the work efficiency in the clinic. METHODS: Male lower urinary tract symptoms (LUTS) patients, who attended the Department of Urology in Peking University People's Hospital from March 2016 to March 2017, were assessed for their urination function according to the Visual Uroflow Scale without help from clinicians before undertaking a free uroflowmetry test. And afterwards, a free uroflowmetry was undertaken, and variables including maximal flow rate (Qmax), the average flow rate (Qave) and voiding volume (VV) was obtained. During the study, 124 cases were collected and 53 cases met the inclusion and exclusion criteria and were included in the study cohort. The Spearman correlation analysis was used for analyzing the correlation of VUS scores with free uroflowmetry variables and age. The validity of VUS was evaluated. RESULTS: Most of the patients could choose the very figure matched with self-condition by first instinct without any help from the clinician. The data were analyzed by Spearman correlation analysis. In the present study, voiding time was positively correlated with the VUS score (correlation coefficient, 0.62, P < 0.05). In the present cohort, the patients chose the third and fourth figures to take longer time to urinate, implying worse LUTS situation. Flow time and VUS scores were positively correlated (correlation coefficient, 0.61, P < 0.05). The patients with higher VUS scores would spend more time on urinate, no matter how long urinary hesitation was. Both Qmax and Qave were negatively correlated with the VUS score (correlation coefficient －0.54, －0.62, P < 0.05). The study illustrated that the VUS score suggested that the Qmax basically and further reflected the urination function. And its relationship to age revealed the decreased urination function of aging male, which had reached a consensus. CONCLUSION Development of VUS has helped the clinician assess the urination function preliminarily at the first time. Patients are assessed for a VUS score before getting surgery or receiving the drug for treatment, and can be re-assessed after. The VUS score can provide an objective quantitative basis to evaluate the treatment efficacy. In addition, considering that it is convenient, timesaving and easy to understand, the VUS is available for follow-up.
Cohort Studies ; Humans ; Lower Urinary Tract Symptoms ; Male ; Urination ; Urodynamics

OBJECTIVETo evaluate the correlation of 64-multidetector-row CT (64MDCT) perfusion imaging with microvessel density(MVD) and vascular endothelial growth factor(VEGF) in colorectal carcinoma.

METHODS64MDCT perfusion imaging was performed in 33 patients with pathologically verified colorectal carcinoma. Time-density curves (TDC) were created from the region of interest (ROI) drawn over the tumor, target artery and vein by 64MDCT with perfusion functional software. The individual perfusion maps generated were for blood flow (BF), blood volume (BV), mean transit time (MTT) and permeability-surface area product (PS). MVD and VEGF expression of surgical specimens were examined by immunohistochemical staining with anti-CD34, anti-VEGF monoclonal antibody. MVD and VEGF were compared among the different types of TDC in colorectal carcinoma. The correlation of CT perfusion parameters with MVD and VEGF was also examined.

RESULTSTDC of colorectal carcinoma was divided into five types according to their shapes. MVD in the colorectal carcinoma was 22.61+/-9.01. VEGF staining was found in 25 of 29 tumors (86.2%). The score of VEGF expression was 4.15+/-1.09. No significant differences of MVD and VEGF expression among TDC types were found (F=2.59, 1.11, P>0.05). There were also no correlations of MVD and VEGF expression with any dynamic CT parameters (P>0.05).

CONCLUSION64MDCT perfusion imaging, MVD and VEGF may reflect angiogenic activity, but no significant correlations are found among them.

Adult ; Aged ; Colorectal Neoplasms ; blood supply ; diagnostic imaging ; Female ; Humans ; Male ; Microvessels ; Middle Aged ; Neovascularization, Pathologic ; Tomography, Spiral Computed ; methods ; Vascular Endothelial Growth Factor A ; metabolism ; Young Adult

Monitoring the long-term radiotherapy-associated molecular changes in low-grade gliomas (LGGs) facilitates the understanding of LGG response to radiotherapy. In this study, we used immunohistochemistry to analyze the expression of Ki-67, tumor protein P53 (TP53), P21, and P27 in 8 paired WHO grade II astrocytoma samples. The interval between radiotherapy (RT) and the second surgery was more than 3 months in all cases. The average Ki-67 labeling index (LI) was 5.3% in pre-RT samples and 11.54% in post-RT samples. Ki-67 LI was higher in the primary tumors that underwent malignant transformation observed at the second surgery after radiation. Post-RT Ki-67 LI decreased in 2 cases with an interval of less than 12 months between RT and the second surgery. TP53 expression was found in 3 out of 4 pre-RT samples with malignant transformation and in 1 out of 4 pre-RT samples without malignant transformation. Post-RT TP53 increased in 2 cases in which increased expression of P21 or P27 was also observed. Our study suggests that radiotherapy can inhibit WHO grade II astrocytoma proliferation as reflected by Ki-67 LI, but the effect attenuates with time. In addition, there is a tendency of malignant transformation for WHO grade II astrocytomas with a high Ki-67 level or TP53 expression in initial samples.
Adult ; Astrocytoma ; metabolism ; pathology ; radiotherapy ; surgery ; Brain Neoplasms ; metabolism ; pathology ; radiotherapy ; surgery ; Cell Proliferation ; radiation effects ; Cell Transformation, Neoplastic ; radiation effects ; Cyclin-Dependent Kinase Inhibitor p21 ; metabolism ; Cyclin-Dependent Kinase Inhibitor p27 ; metabolism ; Female ; Humans ; Immunohistochemistry ; Ki-67 Antigen ; metabolism ; Male ; Middle Aged ; Neoplasm Grading ; Tumor Suppressor Protein p53 ; metabolism