BACKGROUND:Breast cancer stem cel s have a greater impact on the occurrence and metastasis of breast cancer. Under simulated tumor microenvironment, we can better analyze the proliferation and differentiation of breast cancer stem cel s. OBJECTIVE:To explore the tumor microenvironment effect on the differentiation of breast cancer stem cel s. METHODS:Breast cancer cel s and MCF-7 cel s were primarily cultured in fibroblast supernatant and serum-free PCM-2 medium, and formation of breast cancer cel s microspheres was observed. Proliferative ability of breast cancer cel s was detected using MTT colorimetry, and the surface markers of breast cancer stem cel s and epithelial-mesenchymal transition markers were measured using immunocytochemistry and RT-PCR methods. RESULTS AND CONCLUSION:The diameter of primary cel microspheres was larger in the serum-free PCM-2 medium than in the fibroblast supernatant, but the culture speed was faster in the fibroblast supernatant than the serum-free PCM-2 medium. At 3 days of primary culture, the expression of ALDH1 in primary cel s was greatly higher in the serum-free PCM-2 medium than in the fibroblast supernatant. However, the expressions of E-cadherin and vimentin were up-regulated in the fibroblast supernatant than in the serum-free PCM-2 medium. In addition, the expressions of E-cadherin and vimentin in MCF-7 cel s cultured in the fibroblast supernatant were up-regulated, while the expressions of ALDH1 and Oct-4 were downregulated. These findings indicate that the tumor environment has some certain effects on the growth and differentiation of breast cancer stem cel s, and some cytokines secreted from fibroblast supernatant can promote the proliferation and differentiation of breast cancer stem cel microspheres to some extent.

Objective To assess the role of E-cadherin (CDH1) promoter methylation in bladder carcinogenesis by meta-analysis. Methods The relevant database were searched by the retrieval strategy of Cochrane network. All included studies were collected following data:the first author’s surname, publication year of article, country, language of publication, design of study, sample size, ethnicity, histological subtypes, methylation detection method and genotype frequencies etc. This meta-analysis was performed using the STATA 12.0 software. The crude odds ratio (OR) with 95%confidence interval (CI) was calculated. Results Ten case-control studies were included in this meta-analysis. The methylation frequency of CDH1 was detected in 620 bladder cancer tissues and 341 normal or cancerous tissues. Results showed that the methylation frequency of CDH1 was significantly higher in bladder cancer tissue than that of normal or cancerous tissue (OR=3.09, 95%CI:1.13~8.50, P=0.029). Furthermore, the ethnicity-stratified analysis revealed that the methylation frequency of CDH1 was significantly higher in bladder cancer tissue of Asian populations than that of normal or cancerous tissue (OR=3.85, 95%CI:1.46~10.14, P=0.006), but no such association was found in Caucasian populations(OR=2.22, 95%CI:0.38-12.91, P=0.375). The subgroup analysis based on the detection methods revealed that there was a statistically significant difference in the methylation frequency of CDH1 between bladder cancer tissue and adjacent tissues and normal tissues under the MSP subgroup (P<0.001), while such association was not observed under the Q-MSP subgroup (P=0.818). Conclusion Pro?moter methylation of CDH1 gene may be involved in the occurrence and development of bladder cancer, which may serve as a biomarker for diagnosis and prognosis of bladder cancer.

Objective To describe the genetic epidemiologic features of alopecia areata (AA) patients in China and to presume the possible genetic mo del of AA.Methods A case-controlled study of 1032 AA patients was performed to analyze the effect of genetic factors on the liability to AA.Complex segreg ation and heritability analysis were performed using Falconer's method and SAGE-REGTL programs.Results The mean age of onset was 28.98 ? 13.43 years.The d ifference in the mean age of onset was not significant between males and females.A total of 82.6 percent of patients experienced their first episode of AA befo re the fourth decades of life.A positive family history of AA was obtained in 8 7 patients (8.43%).The prevalences of AA were 1.58%,0.19% and 0.03% in the firs t-,second-and third-degree relatives of the probands respectively,which were significantly higher than those in the controls(P

Objective To study the phylogenetic evolution and genetic variations of gag gene among the prevalent human immunodeficiency virus (HIV )‐1 strains in Guangxi Zhuang Autonomous Region . Methods Plasma samples of 158 HIV‐1 infected patients in Guangxi area were collected during October 2011 to March 2012 .The gag gene fragments of HIV‐1 were amplified by reverse transcription/nested‐polymerase chain reaction and then sequenced .MEGA 5 .03 was utilized to construct phylogenetic tree and to calculate the genetic distances and selection pressures (globle ω) of gag gene and its coding regions . The comparisons between two groups were tested by Student′s t test ,and the comparisons of multiple groups were tested by one‐way ANOVA .Results A total of 140 amplification products of gag gene were obtained from 158 samples .Four subtypes of HIV‐1 were found ,including CRF01_AE (80 ,57 .1% ) , CRF08_BC (46 ,32 .9% ) ,CRF07_BC (10 ,7 .1% ) ,and subtype B (B′) (4 ,2 .9% ) .The genetic distances of gag gene of the above subtypes were 0 .036 ± 0 .001 ,0 .031 ± 0 .002 ,0 .043 ± 0 .003 and 0 .102 ± 0 .006 ,respectively ,with statistical significance (F=220 .62 ,P<0 .01) .The p17 and p24 coding regions suffered negative selection pressure (globleω<1) .Neither the globle ω in p17 region nor that in p24 region had significant differences among different subtypes (F=0 .761 ,P=0 .469 and F=0 .037 ,P=0 .964 , respectively ) . Conclusion CRF01_AE is the major subtypes of HIV‐1 in Guangxi Zhuang Autonomous Region .The coding regions of gag gene are relatively conserved during evolution .Changes of HIV‐1 prevalence ,however ,may affect the genetic variation of gag gene ,which should be continuously monitored .

Objective: To construct a polycistron tandem repeated Echistatin (Ecs) gene. Methods: Three Ecs genes with independent initiation and termination codon were ligated tandem through restriction enzyme sites after amplified with 3 pairs of primers using pMD18T-Ecs as template. The polycistron Ecs gene was inserted into pET30a and expressed in E.coli BL21(DE3) with IPTG induction. The expression results were identified by 18% SDS-PAGE and Western blot. Results: The expression of Ecs polycistron was accomplished with 18% expression level of total protein determined by SDS-PAGE and Western blot. Conclusion: The successful expression of Ecs polycistron provided a new method for the preparation of low molecular weight protein.

The potential of Rhizobium sp. N613 to produce the exopolysaccharide (REPS) was studied in this paper. Using an orthogonal design in a flask-shaker culture system, the fermentation medium and conditions of synthesizing REPS were optimized. Based on these results, the fermentation kinetic parameters were obtained in the batch fermentation with a 10L fermentor. The REPS yield of 11.31g/L was achieved by metabolic regulation during 40 h fed-batch fermentation. Transplanted tumor models of sarcoma 180 in mice were used to evaluate the anti-tumor effect. The result of anti-tumor activities showed that inhibition rate was 53.40%, when dose of REPS was 5mg/kg. These results indicate that REPS has the following properties: the short duration of fermentation, the high yield, the low cost, the effective immunocompetence and thickening. Thus, REPS has the value of development and application.

AIM:To observe related biological parameters of 3 minutes dark-room provocative test in patients with laser peripheral iridectomy(LPI) in the fellow eyes of acute primary angle-closure (APAC) by ultrasound biomicroscopy (UBM).To explore the risk factors in primary angle closure suspect(PACS) patients with progressive angle closure after LPI.METHODS: Seventy-eight eyes of APAC patients without peripheral anterior synechia were selected.Each eye underwent 3 minutes dark-room provocative test after LPI.Anterior segment parameters, including anterior chamber depth (ACD), anterior chamber angle open distance500 (AOD500), peripheral iris thickness (PIT), iris convex (IC), the position of iris insertion and trabecular-ciliary process distance (TCPD), and the number of positional angle closure(NPAC) were observed and analyzed by statistic methods.RESULTS:Patients with APAC were examined by UBM after LPI and 26 eyes(33%) occurs at least one positional angle closure,19 eyes(24%)were positive in 3 minutes dark-room provocative test among them.It occurs a positive relationship between the elevation intraocular pressure and the number of positional angle closure in dark-room provocative test(r=0.84, P<0.01).AOD500, IT and IC were significantly changed from normal light to darkroom between positional angle closure positive group and positional angle closure negative group(all P<0.01).In single factor analysis, AOD500(P=0.003), IT(P=0.012), IC(P=0.043), TPCD(P=0.015), the position of iris insertion(P=0.024) were correlative factors of positive results.In multiple-factor analysis, only IT(P=0.011), TPCD(P=0.009), iris root attachment points(P=0.02) were independent risk factors of positive results.CONCLUSION:A certain proportion of patients with PACS after LPI appeared positional angle closure in a dark room.Peripheral iris hypertrophy, anterior displacement of the ciliary body and iris root attachment points are vital risk factors.Long-term follow-up study and intervention treatment are required in these patients after LPI.

0.05).Conclusion There is no association between IL-2 levels in NS and RSV bronchitis.The IL-2 levels show a heterogenous behavior.

A chiral high-performance liquid chromatography method was developed for the simultaneous determination of ibuprofen enantiomers in dog plasma. It was used to study the pharmacokinetics in the Beagle dog after intravenous administration of racemic-ibuprofen, S-ibuprofen and R-ibuprofen. Ketoprofen was chosen as the internal standard. After a simple precipitation using methanol as the precipitating solvent, both analytes and IS were separated on a Kromasil 100-5CHI-TBB chiral column (250 mm x4.6 mm, 5 μm) with isocratic elution using acetonitrile - 20 mmol x L(-1) phosphate buffer (pH 3.0, containing 5% methanol) (6 : 4) as the mobile phase. The detection wavelength was 220 nm. Liner calibration curves for both of the ibuprofen enantiomers were over the concentration range from 0.5 to 50 μg x mL(-1) with a lower limit of quantification of 0.5 μg x mL(-1), the accuracies were all in standard ranges. The intra- and inter- assay precisions were all below 7%. The recovery rate was 93.1% to 100.4%. The experiments proved that the method was simple, rapid and sensitive. It can be used in the quantitative determination of ibuprofen enantiomers in dog plasma. The method was used to determine the concentration of ibuprofen enantiomers in Beagle dog plasma after a single intravenous administration of racemic-ibuprofen, S-ibuprofen and R-ibuprofen (9 mg x kg(-1)) and the pharmacokinetics parameters were calculated based on the concentration-time curves. The C(max) of S-ibuprofen in Beagle dog plasma after a single intravenous administration of racemic-ibuprofen, S-ibuprofen and R-ibuprofen were 30.8 ± 4.7, 46.1 ± 5.9 and 20.0 ± 2.6 μg x mL(-1), respectively. In terms of the exposure of active ingredient, it revealed a significant difference between the administration of S-ibuprofen and the other two groups. The systematical R- to S- chiral inversion was discussed. Comparing the pharmacokinetic parameters at different doses, chiral inversion were 70.1% ± 36.6% and 76.4% ± 36.2%, respectively, after intravenous administration of racemic- and R-ibuprofen. This study provides a theoretical basis for the safety of ibuprofen formula of injection drug.
Animals ; Chromatography, High Pressure Liquid ; Dogs ; Ibuprofen ; blood ; pharmacokinetics ; Stereoisomerism

Objective To investigate the potential application of targeting at vascular endothelial growh factor (VEGF) induced apoptosis in leukemic cell lines by combined use of Bevacizumab and chemotherapeutic drug. Methods Leukemic cells were treated with several drugs at different concentrations in culture. The effect of VEGF, Bevacizumab and co-treated with Ara-C on leukemic cells proliferation were evaluated by CCK-8 and apoptosis and cell cycle were detected by flow cytometry (FCM). Results VEGF could enhance the proliferation of leukemic cells and caused a dose-dependent manner on U937 cell. It also increased the percentage of cells in S phase, tested by, and Bevacizumab group was decreased. Apoptotic rate of cells treated with Bevacizumab or co-treated with Bevacizumab and Ara-C for 48 h were significantly higher when compared with control or Ara-C group, respectively (P<0.05), but the apoptotic rate of VEGF group or VEGF and Ara-C group was lower (P>0.05). There was no significant difference in apoptotic rate between control and combined use of VEGF, Bevacizumab and Ara-C group(P>0.05). Conclusion VEGF could enhance the proliferation of some leukemic cells, and may contribute to leukemic cells survival and a resultant resistance to chemotherapy-triggered cell death. The study also showed that leukemic cells growth was significantly inhibited by Bevacizumab through directly against VEGF, and the sensitivity of leukemic cells for chemotherapeutic drug was increased.