The purpose of the present study was to explore the seizure-induced changes in Bad (Bcl-2-associated death protein), 14-3-3, phosphoBad, Bcl-2 and Bcl-XL expression in the rat model of focal limbic seizure. Unilateral intra-amygdaloid injection of kainic acid (KA) was made to induce seizure. Electroencephalogram (EEG) and regional cerebral flow (r-CBF) were monitored continuously. Diazepam (30 mg/kg) was administered to terminate the seizure. The apoptotic and surviving neurons in the hippocampus were observed by terminal deoxynucleotidyl transferrase-mediated dUTP nick end labeling (TUNEL) and cresyl violet staining, the expression of Bad, 14-3-3, phosphoBad, Bcl-2 and Bcl-XL were detected with immunofluorescence, Western blot and immunoprecipitation. The results showed that TUNEL-positive neurons appeared at 8 h and reached maximum at 24 h following seizure cessation within the ipsilateral CA3 subfield of the hippocampus. Seizure induced the dephosphorylation of Bad and the dissociation of Bad from its chaperone protein 14-3-3 and subsequent dimerization of Bad with Bcl-XL. The expression of phosphoBad decreased and Bcl-2 increased. There was little change in r-CBF after the seizure. These results suggest that seizure leads to a dephosphorylation of Bad and an upregulation of Bcl-2. Dephosphorylation of Bad may be injurious while the upregulation of Bcl-2 may be protective to the brain damage induced by seizures, but not related with r-CBF.
Amygdala ; physiology ; Animals ; Epilepsies, Partial ; chemically induced ; metabolism ; Hippocampus ; metabolism ; Kainic Acid ; Male ; Microinjections ; Phosphorylation ; Proto-Oncogene Proteins c-bcl-2 ; biosynthesis ; genetics ; Rats ; Up-Regulation ; bcl-Associated Death Protein ; metabolism

OBJECTIVETo evaluate the relationship between combined multigene detection and response to adjuvant chemotherapy and prognosis in early-stage non-small cell lung cancer (NSCLC).

METHODSTissue microarray was prepared from samples of 86 cases of early-stage NSCLC who received adjuvant chemotherapy after radical surgery. The expressions of caspase-3, Fas, bax, bcl-2, survivin, PCNA, Ki67, MGMT, p53, p63, p73, p16, p27, VEGF, nm23, P-gp, MRP, LRP, GST-pi, Topo II, c-myc, cyclin-D1, Her-2, Cox-2, Ku70, Ku80, DNA-PKcs, ERCC1, MSH2, BCRP proteins were detected using immunohistochemical two-step method.

RESULTSThe positive rate of the 30 genes in lung cancer tissue were 27.9% - 91.9%, respectively. By univariate analysis, the expression of 8 genes was shown to be related with SCLC adjuvant chemotherapy. The cases with higher expression of survivin, P-gp, LRP, Ki67, p53, ERCC1 and lower expression of bax,VEGF had worse prognosis. By logistic regression analysis, the ERCC1, survivin, bax and VEGF were a marker group. Multivariate analysis showed the predict value of the response to adjuvant chemotherapy in early-stage NSCLC was 96.5%.

CONCLUSIONSurvivin, ERCC1, bax and VEGF are an ideal marker group to predict the effect of adjuvant chemotherapy in early-stage NSCLC.

Adult ; Aged ; Carcinoma, Non-Small-Cell Lung ; drug therapy ; metabolism ; pathology ; surgery ; Chemotherapy, Adjuvant ; DNA-Binding Proteins ; metabolism ; Endonucleases ; metabolism ; Female ; Follow-Up Studies ; Humans ; Inhibitor of Apoptosis Proteins ; Logistic Models ; Lung Neoplasms ; drug therapy ; metabolism ; pathology ; surgery ; Male ; Microtubule-Associated Proteins ; metabolism ; Middle Aged ; Neoplasm Staging ; Survival Rate ; Tissue Array Analysis ; Vascular Endothelial Growth Factor A ; metabolism ; bcl-2-Associated X Protein ; metabolism

OBJECTIVETo isolate and characterize the side population cells (SP cells) in the lung adenocarcinomas cell line A549.

METHODSThe protein expression of ABCG2 in human lung adenocarcinoma cell line A549 was detected by immunohistochemistry. SP and NSP cells in the cell line A549 were isolated by FACS, and their differentiation was analysed. ABCG2 expression in the two cell subsets was detected by RT-PCR. The cell growth curves, cell division indexes, cell cycles, plate clone formation tests, migration and invasion assays, chemotherapeutic susceptibility tests, tests of the intracellular drug levels, and the tumor cell implantation experiments on nude mice were applied to study the biological properties of the two cell subsets. The expression of ABCG2 in the transplanted tumor in nude mice was detected by immunohistochemistry and RT-PCR.

RESULTSThe positive rate of ABCG2 expression in the A549 cells by immunohistochemistry was 2.13%. SP and NSP cells were isolated by FACS. The SP cells could produce both SP and NSP cells, while NSP cells only produced NSP cells. SP cells expressed ABCG2, but NSP cells did not. The proliferation and migration abilities of the two cell subsets were similar, but the invasion and tumorigenic ability of SP cells was significantly higher than that of NSP cells. The susceptibilities to DDP and its intracellular levels of the two cell subsets were similar, but the susceptibilities to 5-FU, VP16, NVB and GEM and their intracellular levels of NSP cells were significantly higher than those of the SP cells.

CONCLUSIONSSP cells in the human lung adenocarcinomas cell line A549 is enriched with tumor stem cells. An effective way to get lung adenocarcinomas stem cells is to isolate SP cells by FACS.

ATP-Binding Cassette Transporters ; metabolism ; Adenocarcinoma ; pathology ; Animals ; Cell Cycle ; Cell Differentiation ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Cell Transformation, Neoplastic ; metabolism ; Fluorouracil ; metabolism ; Humans ; Lung Neoplasms ; pathology ; Mice, Nude ; Neoplasm Proteins ; metabolism ; Neoplasm Transplantation ; Neoplastic Stem Cells ; drug effects ; Side-Population Cells

Objective To analyze the effects of S100A6 on Wnt/β-catenin signaling pathway and its molecular mechanism. Methods The expression of GST-hS10OA6 was induced with IPTG in Escherichia coli BL21, and the fusion protein was purified with glutathione-sepharose 4B beads. β-catenin level of human colon cancer cell line MG63 and human osteosarcoma cell line HCTl16 cells infected with AdS10OA6 was measured by Western blot. Luciferase activity assay was applied to analyze the effect of S100A6 on the β-catenin/TCF4 activity. The interactions between S100A6 and β-catenin/GSK-3β/Dvl/Axin were detected by GST-pulldown/Western blot. Results The β-catenin level in AdS100A6-infected MG63 and HCT116 cells was significantly increased in comparison with that in the AdGFP control group (P<0.01). The luciferase activity in human embryonic renal cell line 293 cells transfected with pTOP-Luc and followed by GST-hS100A6 treatment was increased by 20. 2-fold in comparison with that in the GST control group (P<0.01). The interaction between GST-hS100A6 and Axin was not found. Conclusion S100A6 up- regulates the Wnt/β-catenin signaling pathway, and this may be attributed to the interaction between S100A6 and β-catenin/GSK-3β/Dvl.

Objective To analyze the effects of S100A6 on Wnt/β-catenin signaling pathway and its molecular mechanism. Methods The expression of GST-hS10OA6 was induced with IPTG in Escherichia coli BL21, and the fusion protein was purified with glutathione-sepharose 4B beads. β-catenin level of human colon cancer cell line MG63 and human osteosarcoma cell line HCTl16 cells infected with AdS10OA6 was measured by Western blot. Luciferase activity assay was applied to analyze the effect of S100A6 on the β-catenin/TCF4 activity. The interactions between S100A6 and β-catenin/GSK-3β/Dvl/Axin were detected by GST-pulldown/Western blot. Results The β-catenin level in AdS100A6-infected MG63 and HCT116 cells was significantly increased in comparison with that in the AdGFP control group (P<0.01). The luciferase activity in human embryonic renal cell line 293 cells transfected with pTOP-Luc and followed by GST-hS100A6 treatment was increased by 20. 2-fold in comparison with that in the GST control group (P<0.01). The interaction between GST-hS100A6 and Axin was not found. Conclusion S100A6 up- regulates the Wnt/β-catenin signaling pathway, and this may be attributed to the interaction between S100A6 and β-catenin/GSK-3β/Dvl.

Talents are the main force for the development of traditional Chinese medicine（TCM）， and the construction of TCM talents and the reformation of talent evaluation system are essential to promote the inheritance and innovation of TCM. At present， we are still exploring and developing in the fields of the formulation， implementation and evaluation indicators of TCM talent evaluation system. However， there are shortcomings and difficulties. For instance， insufficient stratification in the evaluation， excessive emphasis on the quantity of achievements， neglecting the quality of the achievements and the actual contribution， imperfect assessment indicators， and the weak characteristics of TCM. Therefore， national ministries and commissions have jointly issued a document requesting to break the four only and set a new standard， in order to promote the construction of a scientific and technological talent evaluation system oriented by innovation value， ability and contribution. For the evaluation of TCM clinical talents， China Association for Science and Technology commissioned China Association of Chinese Medicine to build the China Clinical Cases Library of TCM（CCCL-TCM）， which aims at collecting the most authoritative and representative TCM clinical cases and exploring the advantages of applying clinical cases as masterpiece of achievement in TCM clinical talents evaluation. CCCL-TCM can promote the construction of a talent evaluation system that is more in line with the development characteristics of TCM industry， and to carry out relevant pilot in TCM colleges and institutions across the country in order to promote the reformation of TCM talent evaluation system.