Objective:To explore stem cells phenotype of human epidermal growth factor receptor 2(HER2)-overexpressing breast cancer cells,MDA-MB-453 by isolation of CD44+CD24-/low and side population(SP).Analyze the relationship between SP cells and the gene amplification levels of the key proteins,?-catenin and Notch-1 in Wnt and Notch signal pathways.And study the correlation of SP cells and the gene amplification of HER2 and human epidermal growth factor receptor 3(HER3).Methods:CD44+CD24-/low and SP phenotype were studied in MDA-MB-453(HER2-overexpressing cell line)and MCF7(HER2 negative cell line)respectively by flow cytometry.The gene amplification of ?-catenin and Notch-1 in Wnt and Notch pathways were also studied by reverse transcriptase polymerase chain reaction(RT-PCR).The gene amplification levels of the proteins mentioned,HER2 and HER3 were roughly compared in MDA-MB-453 cells and their SP by RT-PCR.Results:There was no CD44+CD24-/low subpopulation in MDA-MB-453 cells,However,SP could be observed in MDA-MB-453.The gene amplification levels of ?-catenin and Notch-1 in Wnt and Notch pathways were both observed in MDA-MB-453 and MCF7.Their gene amplification levels were more evident in SP of MDA-MB-453 than in MDA-MB-453 cells.However,the similar gene amplification levels of HER2 and HER3 were observed.Conclusion:SP can be enriched in breast cancer stem cells of MDA-MB-453.Wnt and Notch pathways are activated in MDA-MB-453,which may be related to maintenance and activity of breast cancer stem cells of MDA-MB-453.

Objective To evaluate the value of color Doppler flow imaging(CDFI) in the diagnosis of congenital coronary artery fistula (CAF).Methods Thirteen consecutive patients ranging in age from 8 to 68 years underwent conventional transthoracic echocardiography,which depicts their origin, course, and drainage site. Results Right CAFs were detected in 8 patients: four of them had drainages to the left ventricle, two to the main pulmonary artery, one to the right ventricle(RV) and the other to the right atrium(RA). CAFs arising from the left coronary artery were noted in five patients: three of them had drainages to the main pulmonary, and the other two to the RA and RV respectively. The origin and the course were precisely defined and the drainage site was clearly depicted in all thinteen patients. The diagnosis of the thirteen patients was confirmed by coronary angiography, and eight of them received surgical treatment. Conclusions CDFI offers a simple, useful and reliable technique in diagnosis of CAF.

Bupleurum chinense used in the study were cultivated in the experimental fields of Gansu agricultural University for three years. The seeds of B. chinense were collected every 3 days 10 d after the blossom. The result showed that the 1 000-grain fresh weight reached the maximum 43 d after the blossom and then decreased rapidly, at the mature period the fresh weight of seeds were falling to the same level of the dry weight. The dynamic change of the grain dry matter accumulation showed as an S-shape curve, the rapid increase stage was 25-34 d following the flower, and the grain filling was ended 46 d after blossom. Grain filling rate was under the law "fast-slow-fast-slow". And there were two peaks of grain filling rate appeared, after reached the second peak 28 d after the flower the filling rate decreased rapidly and stayed steadily 43 d after flowering. The dehydration rate was also measured at its maximum 43 d following flower. The indexes of seeds all reached the top 52 days following the blossom, when the germination rate reached the peak (34.33%) and water content of seeds was near 10%. The rate of germination and the 1 000-graid weight of seed showed significant positive correlation, while the water content of seeds was found significant negatively correlation with germination percentage. So the best time for harvest should be 52 d after flowering (9 month), the seeds collected at that time showed both high quality and germination rate.
Bupleurum ; chemistry ; growth & development ; Flowers ; growth & development ; Germination ; Seeds ; chemistry ; growth & development ; Water ; analysis

[ ABSTRACT] AIM:To investigate the effects of BARF1 down-regulation on EBV-positive gastric carcinoma cell apoptosis, and the molecular mechanisms by BARF1 silencing-mediated apoptosis.METHODS: After NUGC3 and SNU719 cells were transfected with NCsiRNA and siRNA, respectively, the protein levels of BARF1, Bcl-2, Bax, cyto-chrome C, caspase 3 and capase 9 were detected by Western blot, and the mRNA expression of BARF1, Bcl-2 and Bax was determined by RT-PCR.The cell viability was measured by the method of Trypan blue exclusion and the cell apoptosis was analyzed by flow cytometry analysis with Annexin V-FITC/PI staining.The expression of the apoptosis-related proteins in the cells transfected with siRNA and NCsiRNA was examined by human apoptosis antibody arrays.Mitochondrial mem-brane potential was determined by flow cytometry.The interaction between Apaf-1 and caspase 9 was confirmed by immuno-precipitation.RESULTS: Compared with untreated and NCsiRNA groups, BARF1 gene silencing significantly inhibited the cell viability, induced apoptosis, and reduced the mitochondrial membrane potential in the NUGC3 and SNU719 cells transfected with siRNA.BARF1 gene silencing up-regulated the expression of pro-apoptotic proteins and down-regulated the expression of anti-apoptotic proteins, and the Bcl-2/Bax ratio was significantly decreased.In BARF1 gene silencing cells, the caspase inhibitor z-VAD-fmk inhibited BARF1 silencing-mediated apoptosis, and significantly increased the levels of cleaved caspase 3 and caspase 9.The concentration of cytochrome C significantly increased as compared with NCsiRNA group, and Apaf-1 interacted with caspase 9 in the cytoplasm.CONCLUSION:BARF1 silencing induces apoptosis via the mitochondrial pathway through regulating the expression of Bcl-2 and Bax proteins in a caspase-dependent manner in the NUGC3 and SNU719 cells.

Objective To evaluate the effectiveness of the suture anchor in the treatment of acute closed distal achilles tendon rupture. Methods A retrospective review of clinical data in 25 patients with acute achilles tendon rupture was performed in the department of orthopedic trauma of Tianjin Hospital between October 2012 to January 2014. The operation was performed by standard approach, with the suture anchors to reconstruct the achilles tendon insertion, and the Krachow technique was used to suture the achilles tendon. All patients were followed up for 19~35 months. The situation of the wound healing, heel pain, and achilles tendon re-rupture were observed and analyzed. According to the passive range of motion of the bilateral ankle joint, the achilles tendon total rupture score (ATRS), the American orthopaedic foot and ankle society (AOFAS) score and functional recovery were assessed one year after operation. Results The operation was successfully completed in this group. The operation time was 25-35 min, the average time was (30.4 ± 4.8) min. No major vascular and nerve injury occurred in the operation. All patients were followed up. The wound healed well. No infection and skin necrosis, no heel pain and achilles tendon re-rupture were found during follow-up. There was no significant difference in mean passive plantar flexion between injured ankles and uninjured ankles one year after operation (44.36° ± 3.33° vs. 46.40°±4.53°, P>0.05). But mean passive dorsiflexion was lower in injured ankles than that of uninjured ankles (16.88°± 4.10° vs. 20.12°±3.21°,P<0.05). The mean ATRS score was 88.7±6.1 (range 79-97), and the mean AOFAS score was 92.4± 5.6 (range 76-100) at first postoperative year. According to the AOFAS scale, the results were excellent in 20 cases and good in 5 cases. Conclusion It was successful to repair the acute closed distal achilles tendon injuries with the suture anchors.

ObjectiveTo detect the mutations of GJB3 and GJB4 genes in two sporadic cases of erythrokeratodermia variabilis(EKV).MethodsGenomic DNA was extracted from two sporadic patients with EKV,their family members,and 100 normal human controls.All the exons and adjacent splice sites of GJB3 and GJB4 genes were amplified by PCR.Mutation scanning was carried out via direct bidirectional DNA sequencing.ResultsA G134C mutation was found at the GJB3 gene in patient 1,which caused a substitution of glycine by alanine at codon 45 (G45A).No mutation was found in the GJB4 gene in case 1 or GJB3 and GJB4 genes in case 2.ConclusionA missence mutation G45A in GJB3 gene is found in a patient with EKV.

Objective To detect the relationship between apical rotation and apical myocardial structure.Methods To observe the change of apical rotation,14 mongrel dogs were heated at apexes,and half and three quarter of apical wall thicknesses were destroyed according to their corresponding heating time.Ultrasonic tissue Doppler imaging was used to record the velocities of apical rotation.Computer programming was applied to calculate apical rotation according to the formulas of apical rotation calculation.Apical rotations were compared between before apical heating,half and three quarter of apical wall thicknesses destruction,respectively.Results Compared with the apical rotation before heating,apical rotation after half of apical wall destroyed decreased [(5.2±3.1)°vs (12.4±2.9)°,P＜0.001];but after three quarter of apical wall destroyed,the apical rotation increased [(7.3±3.6)°vs (5.2±3.1)°P＜0.01].Conclusions Outer part of apical myocardium was outer layer myocardium of which the damage leads to the decrease of apical rotation;inner part of apical myocardium was inner layer myocardium of which the damage leads to the increase of apical rotation to some extent.

Objective:To evaluate the clinical efficacy of maintenance chemotherapy for patients who had local advanced non-small cell lung cancer (NSCLC) and was responsive to primary radiotherapy and chemotherapy. Methods:One hundred and twenty patients with stage ⅢA or ⅢB NSCLC received 4 cycles of chemotherapy combined with radiotherapy. The 63 patients who achieved certain remission were randomly divided into maintenance chemotherapy group(n=33) and control group(n=30). The patients in maintenance chemotherapy group (treatment group) received vinorelbine (20 mg/m2, d 1 and d 8, per 28 d a cycle) and those in control group were not given maintenance chemotherapy. The clinical efficacy, survival rate and adverse reaction of the two groups were evaluated. Results:There are a longer median time to progression(TTP) in treatment group compared with control group (8.5 month vs 5.0 month, P<0.05). The 1-and 2-year survival rates were 66.7% and 36.4% in the treatment group and 60.7% and 32.1% in the control group, respectively. The difference between the survival rates of two groups was not significant (P>0.05). Conclusion:Maintenance vinorelbine-based chemotherapy prolonged the median time to progression but had no effect on survival time in patients with local advanced NSCLC who responded to induction chemotherapy.

Objective To explore the immune function of the whole body and the tumor site of human nasopharyngeal carcinoma (NPC) patients as well as its correlation with CCR4. Methods The ratios of CD4~+CD25~+T cells and CCR4+cells to lymphocytes were measured by flow cytometry in tissues (25 cases) and peripheral blood (35 cases) from nasopharyngeal carcinoma patients, and then statistical analysis was made. Results The ratios of CD4~+CD25~+T cells and CCR4+ cells in tissue and peripheral blood of NPC were higher than those in the control group (P<0.05). In NPC the ratios of the two cells in tissue were higher than in peripheral blood respectively (P<0.05), but there was no such difference between tissue and peripheral blood in the control group (P>0.05). The ratio of CD4~+CD25~+T cells in tissue at stage Ⅲ+Ⅳ was higher than that at stage Ⅰ+Ⅱ of NPC (P<0.05), but there was no such difference between the two stages in peripheral blood in NPC. There was a positive correlation between CD4~+CD25~+T cells and CCR4+ cells in tissue and peripheral blood of NPC (P<0.05). Conclusion NPC patients have different immunosuppression, and there is even more severe immunosuppression in the tumor site. The ratio of CD4~+CD25~+T cells is correlated with the stage of NPC. Therefore, as NPC progresses to later stages, the percentage of CD4~+CD25~+T cells is increased, which is correlated with poor prognosis. CCR4 plays an important role in reactant of CD4~+CD25~+T cells to tumor sites, and is resistant to immunosurveillance.

Objective: To examine expression patterns of focal adhesion kinase (FAK) and its activated form, phosphorylated FAK (pFAK),in human osteosarcoma and to investigate the correlation of FAK expression with clinicopathological parameters and prognosis. Functional consequence of manipulating FAK protein levels was also investigated in human osteosarcoma cell lines. Methods: Immunohistochemical staining was used to detect FAK and pFAK levels in pathologically archived materials from 113 patients with primary osteosarcoma. Kaplan-Meier survival and Cox regression analyses were used to evaluate prognoses. The role of FAK in cytological behavior of MG63 and 143B human osteosarcoma cell lines was studied via the FAK protein knockdown with siRNA. Cell proliferation, migration, invasiveness, and apoptosis were assessed using cell counting kit-8, Transwell, and Annexin V/PI staining methods. Results: Both FAK and pFAK were overexpressed in osteosarcoma patients. Tumor cells exhibited cytoplasmicity and occasional membranous immunoreactivity for FAK. A total of 42 cases (37.17%) mainly showed expressed pFAK in cytoplasm of osteosarcoma cells. No overexpression staining of anti-FAK and anti-pFAK antibodies was observed in normal cancellous bone tissues or negative controls. Significant differences were observed in overall survival between FAK-/pFAK- and FAK+/pFAK- groups (P=0.016), FAK+/pFAK- and FAK+/pFAK+ groups (P=0.012), and FAK-/pFAK- and FAK+/pFAK+ groups (P<0.001). All groups showed similar metastasis-free survival. Cox proportional hazard analysis showed that FAK expression profile is an independent indicator of both overall andmetastasis-free survival. siRNA-based knockdown of FAK significantly reducedmigration and invasion of MG63 and 143B cells and affected proliferation and apoptosis in osteosarcoma cells. Conclusion: Osteosarcoma malignancies in vitro and in vivo were correlated with overexpression and phosphorylation of FAK. These findings suggest that FAK plays an important biological role in osteosarcoma carcinogenesis. This study provides a better understanding of diagnostic and prognostic relevance of FAK overexpression and phosphorylation in osteosarcoma patients. Therefore, FAK and pFAK can be used as independent predictors of overall and metastasis-free survival in osteosarcoma patients.