The present study is to elucidate the mechanisms underlying Gleevec-induced apoptosis of chronic myeloid leukemia (CML) K562 cells in vitro. The apoptotic cell death and cell cycle distribution after Gleevec treatment and the effect of PDCD4 siRNA on Gleevec-induced apoptosis of K562 cells were analyzed by flow cytometry. The effect of Gleevec on p-Crkl, caspase-3, PARP and PDCD4 protein levels, and the knockdown efficacy of PDCD4 siRNA were detected by Western blotting. The results showed that Gleevec dramatically suppressed the phosphorylation level of Crkl in a dose-dependent manner and induced significant apoptosis and G0/G1 cell cycle arrest of K562 cells in time- and dose-dependent manners. In addition, Gleevec activated caspase-3 and its downstream substrates PARP, and the caspase pan inhibitor Z-VAD-FMK (50 micromol x L(-1)) markedly reduced Gleevec-induced apoptosis from 47.97% +/- 10.56% to 31.05% +/- 9.206% (P < 0.05). Moreover, Gleevec significantly increased the protein expression of programmed cell death 4 (PDCD4). PDCD4 knockdown by siRNA reduced Gleevec-induced apoptosis from 46.97% +/- 14.32% to 42.8% +/- 11.43%. In summary, Gleevec induced apoptosis in K562 cells via caspase-3 activation.
Amino Acid Chloromethyl Ketones ; Apoptosis ; drug effects ; Benzamides ; pharmacology ; Caspase 3 ; metabolism ; Cell Cycle ; drug effects ; Humans ; Imatinib Mesylate ; K562 Cells ; Phosphorylation ; Piperazines ; pharmacology ; Pyrimidines ; pharmacology

Objective:To investigate the influence of sCD80-Linker-sCD40L fusion protein on the unspecific anti- tumor immunity in vitro.Methods:Ovarian cancer SKOV3 cells were separately transfected with recombinant adenoviral vectors containing sCD80-Linker-sCD40L fusion gene,sCD80 gene,sCD40L gene or with control adenovirus.The expres- sion of the sCD80-Linker-sCD40L fusion protein,sCD80 protein and sCD40L protein in the supernatants of SKOV3 cells was determined by ELISA.Dendritic cells(DCs)were cultured with peripheral blood mononuclear cells from a patient with ovarian carcinoma.DCs and autologous T cells were co-cuhured and were exposed to different supernatants for 48 h. The allostimulatory effects of DCs on T cells were determined by mixed lymphocyte reaction(MLR).The unspecific kill- ing activities of induced T cells against SKOV3/K562 cells were measured by LDH-releasing assay.Results:ELISA assay showed that levels of the sCD80-Linker-sCD40L fusion protein,sCD80 protein and sCD40L protein in the supernatants of transfeced SKOV3 cells were 2.791 ng/ml,1.956 ng/ml and 1.407 ng/ml,respectively.The fusion protein-exposed DCs ([0.382?0.053]vs[0.167?0.028],P

BACKGROUNDThe cross-section of thoracolumbar vertebral body is kidney-shaped with depressed posterior boundary. The anterior wall of the vertebral canal is separated from the posterior wall of the vertebral body on the lateral X-ray image. This study was designed to determine the sagittal distance between the anterior border of the vertebral canal and the posterior border of the vertebral body (DBCV) and to analyze the potential role of DBCV in the estimation of cement leakage during percutaneous vertebroplasty (PVP) or percutaneous kyphoplasty (PKP).

METHODSWe retrospectively recruited 233 patients who had osteoporotic vertebral compression fractures and were treated with PVP or PKP. Computed tomography images of T11-L2 normal vertebrae were measured to obtain DBCV. The distance from cement to the posterior wall of the vertebral body (DCPW) of thoracolumbar vertebrae was measured from C-arm images. The selected vertebrae were divided into two groups according to DCPW, with the fracture levels, fracture grades and leakage rates of the two groups compared. A relative operating characteristic (ROC) curve was applied to determine whether the DCPW difference can be used to estimate the degree of cement leakage. The data were processed by statistical software SPSS version 21.0 using independent sample t-test and Chi-square tests.

RESULTSThe maximum DBCV was 6.40 mm and the average DBCV was 3.74 ± 0.95 mm. DBCV appeared to be longer in males than in females, but the difference was not statistically significant. The average DCPW of type-B leakage vertebrae (2.59 ± 1.20 mm) was shorter than that of other vertebrae (7.83 ± 2.38 mm, P < 0.001). The leakage rate of group DCPW ≤6.40 mm was lower than that of group DCPW >6.40 mm for type-C and type-S, but much higher for type-B. ROC curve revealed that DCPW only has a predictive value for type-B leakage (area under the curve: 0.98, 95% confidence interval: 0.95-0.99, P < 0.001), and when the cut-off value was 4.05 mm, the diagnostic sensitivity and the specificity were 94.87% and 93.02%, respectively.

CONCLUSIONSDepression of the thoracolumbar posterior vertebral body may be informative for the estimation of cement location on C-arm images. To reduce type-B leakage, DCPW should be made longer than DBCV on C-arm images for safety during PVP or PKP.

Aged ; Female ; Fractures, Compression ; surgery ; Humans ; Kyphoplasty ; methods ; Male ; Middle Aged ; Osteoporotic Fractures ; surgery ; Retrospective Studies ; Spinal Fractures ; surgery ; Vertebroplasty ; methods

OBJECTIVETo investigate the anti-tumor effects and mechanism of tumor vaccines and whether chemotherapeutic agents administered prior to immunotherapy could augment the efficacy of the vaccines.

METHODSC57/BL mice inoculated with Lewis lung cancer cells were used as tumor models. Granulocyte-macrophage colony-stimulating factor (GM-CSF) gene modified LA795 and Lewis lung cancer cell lines were administered as allogeneic and autologous tumor vaccines, respectively. After Lewis cells (1 x 10(7)) inoculation, the mice received irradiated GM-CSF secreting cancer vaccine solely or in combination with carboplatin. The survival of the mice was observed. The cytotoxicity of spleen cells or purified CD8(+) cells was analyzed by lactate dehydrogenase (LDH) assay. Serum level of IL-4 and IFN-gamma was detected using ELISA method.

RESULTSThe cytotoxicity of the spleen cells or purified CD8(+) T cells against Lewis cells in the mice immunized with cancer cell vaccine was significantly increased, relative to that of the control, untreated group (P < 0.05). Serum level of Th1-type cytokine IFN-gamma was increased after vaccination, whereas Th2-type cytokine IL-4 showed no significant change. The GM-CSF secreting cancer cell vaccine had no significant influence on the survival of the mice with established heavy tumor burden. The combination of chemotherapy and cancer vaccine could statistically prolong the survival time; whereas any method itself had no significant effect.

CONCLUSIONThe GM-CSF secreting cancer cell vaccine can induce immune responses. The chemotherapeutic agents may be beneficial to enhance the anti-tumor activity of cancer vaccine.

Animals ; Antineoplastic Agents ; therapeutic use ; Cancer Vaccines ; therapeutic use ; Carboplatin ; therapeutic use ; Carcinoma, Lewis Lung ; blood ; metabolism ; pathology ; therapy ; Cell Line, Tumor ; Female ; Granulocyte-Macrophage Colony-Stimulating Factor ; genetics ; metabolism ; Interferon-gamma ; blood ; Interleukin-4 ; blood ; Lung Neoplasms ; metabolism ; pathology ; Mice ; Mice, Inbred C57BL ; Transfection

This study was aimed to study the potential effects of alloreactive NK cells (allo-NKs) in therapy of relapsed lung cancer after haploidentical hematopoietic stem cell transplantation using donor lymphocyte infusion (DLI). The F1 donors derived-NK cells were purified with MACS magnetic separation system, in which the proportion of the alloreactive Ly49A(+) cells was detected by flowcytometry and alloreactivity was measured by LDH method. The relapse model of lung cancer after haploidentical-HSCT was established. The distribution kinetic of infused donor lymphocytes in vivo was analyzed. The inhibition of relapse tumor, infiltration of lymphocytes in situ and fluctuation of 22 kinds of cytokines in serum after DLI were compared among different groups. The results showed that the infused donor cells of allo-NK group were accumulated mostly in lung, spleen and kidney for more than 48 hours with considerable higher levels according to the distribution kinetic curve. The sizes of relapse tumors between chemotherapy + PBS group and chemotherapy + DLI group showed no difference. However, the relapsed tumors in allo-NK + DLI group were significantly smaller than that in chemotherapy + DLI group or allo-NK + PBS group, in which increased infiltration of lymphocytes were defined in situ. The levels of cytokines such as MCP-1, IL-17, IL-12 and MCP-5 in serum of allo-NK + DLI group ascended compared with control group, though the level of IL-10 declined simultaneously. It is concluded that allo-NKs prolong the survival time of infused donor lymphocytes in vivo, promote the secretion of inflammatory cytokines and Th1-type of cytokines, and further improve the antitumor effects of DLI against relapse after transplantation.
Animals ; Cytokines ; blood ; Hematopoietic Stem Cell Transplantation ; methods ; Killer Cells, Natural ; cytology ; Lung Neoplasms ; therapy ; Lymphocyte Transfusion ; methods ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C57BL ; Neoplasm Recurrence, Local ; therapy ; Transplantation Conditioning ; methods

OBJECTIVETo investigate the differences of psychological and behavioral development between children aged 1 to 3 years fostered by grandparents and those by parents.

METHODSPsychological and behavioral development of 443 children aged 1 to 3 years fostered by their grandparents and of aged-matched 443 children fostered by their parents were assessed with DST, an intellectual developmental screening test developed by Pediatric Hospital of Fudan University in Shanghai.

RESULTSThe abilities of social adaptation and intelligence development in children fostered by their grandparents were obviously retarded as compared with those in children fostered by their parents.

CONCLUSIONSThere are shortcomings in psychological and behavioral development in children aged 1 to 3 years fostered by grandparents.

Child Behavior ; Child Development ; Child Rearing ; Child, Preschool ; Female ; Humans ; Infant ; Intelligence ; Intergenerational Relations ; Male ; Parents ; Social Adjustment

OBJECTIVETo investigate whether dendritic cells pulsed with whole tumor lysates (WTL) could in vitro elicit antitumor T cell responses in patients with non-small-cell lung cancer (NSCLC).

METHODSMonocyte-derived immature DCs (imDCs) generated in the presence of human recombinant granulocyte-macrophage colony stimulating factor and interleukin-4 from peripheral blood mononuclear cell of NSCLC patients, and then were induced to mature by pulsing autologous WTL (DCs/WTL) or by the addition of TNF-alpha(TNF/DCs). FACS and MLR assay were used to monitor their phenotypic changes and capacity to stimulate allogeneic and autologous T cell proliferation. DCs/WTL activated with TNF-alpha (* DCs/WTL) were cocultured in vitro with autologous T cells for eliciting antitumor CTLs. T cell mediated antitumor responses were measured by IFN-gamma enzyme-linked immunospot (ELISPOT) assay for WTL-specific IFN-gamma releasing T cells and by lactate dehydrogenase release (LDH) assay for lysis of autologous tumor cells, respectively.

RESULTSWhen monocytes-derived imDCs from the patients with NSCLC (n = 10) were pulsed with autologous WTL for a day at 30 microg total protein of WTL per 10(6) DCs/ml, this led to up-regulation of CD1a, CD83 and CD86 as well as HLA-DR, and also led to marked stimulation of allogeneic T cell proliferating activity, which was comparable to that of TNF/DCs. However, their capacity of stimulating autologous T cell proliferation in vitro was significantly more potent than those of TNF/DCs (P < 0.05). The numbers of WTL-specific IFN-gamma releasing T cells in 1/3 cultures after one week exposure to * DCs/WTL was increased significantly compared with those pulsing with TNF/DCs plus IL-2 or IL-2 alone (P = 0.05). T cells derived by priming of non-adherent PBMCs with * DCs/WTL after 14 days in vitro stimulation were significantly more responsive to autologous tumor cells compared with LAK (n = 3, P < 0.05), but its cytotoxicity against K562 cells was also comparable to LAK cells.

CONCLUSIONMonocyte-derived DCs from NSCLC patients could serve as functional APC. The * DCs/WTL may effectively elicit T cell-mediated antitumor response in vitro and enhance NK killing activity.

Antigens, CD1 ; metabolism ; Carcinoma, Non-Small-Cell Lung ; immunology ; Cell Culture Techniques ; Cytotoxicity, Immunologic ; Dendritic Cells ; immunology ; HLA-DR Antigens ; metabolism ; Humans ; Interferon-gamma ; secretion ; K562 Cells ; Killer Cells, Lymphokine-Activated ; immunology ; Leukocytes, Mononuclear ; immunology ; pathology ; Lung Neoplasms ; immunology ; Lymphocyte Culture Test, Mixed ; T-Lymphocytes, Cytotoxic ; immunology ; pathology ; Tumor Necrosis Factor-alpha ; pharmacology

OBJECTIVETo establish a novel Myc gene transgenic mouse model for spontaneously forming B-lymphoma and assessing its tumorigenesis potential.

METHODSFreshly isolated hematopoietic progenitor cells served as the target for Myc gene transfer mediated by a retrovirus vector. These cells were engrafted into C57BL/6 mice with (60)Co-gamma ray radiation in advance. Tumor latency was measured and the tumor loaded mice were followed for survival time. Tumor was identified with histology and immunostaining. The exogenous Myc gene was detected by Western blot (in liver, spleen, tumor tissue) and flow cytometry (FCM) \[in bone marrow (BM)\].

RESULTSMice BM-infected with mutant Myc gene more readily gave rise to B-cell lymphomas than those infected with wild type Myc gene did Myc gene was expressed highly in BM and tumor tissues but not in liver and spleen.

CONCLUSIONOur model will be a tool in assessing the transforming potential of Myc mutants and in studying cooperation between Myc and other oncogenes. Mutant Myc is more effective than wild-type Myc in promoting B cell lymphomagenesis in mice.

Animals ; B-Lymphocytes ; Cell Transformation, Neoplastic ; Flow Cytometry ; Lymphoma ; Lymphoma, B-Cell ; Mice ; Mice, Inbred C57BL ; Mice, Transgenic ; Retroviridae Infections

BACKGROUNDS100A8 and S100A9 are two members of the S100 protein family characterized by the presence of two Ca2+-binding sites of the EF-hand type. Previous studies suggested that the whole S100 family displays significant functions in tumor growth, progression and invasion. This study aimed to determine the expression of the two indices of the family, S100A8 and S100A9, in lung cancer tissues and normal lung tissues and its correlation with clinical features.

METHODSA total of 60 cases with a variety of clinical data that were diagnosed with different histological subtypes of lung cancer were investigated. Semi-quantitative reverse transcriptase-PCR (Sq-Rt-PCR) and immunohistochemical staining of cancer, adjacent and peripheral lung tissues were executed to distinguish the expression patterns of S100A8 and S100A9 and to further clarify their correlation with clinical features.

RESULTSImmunohistochemical staining of both proteins showed a significant up-regulation in lung cancer tissue (S100A8, S100A9, P<0.0001), and PCR revealed that the levels of S100A8 and S100A9 expression were significantly higher in lung cancer tissues (S100A8 P=0.002/0.004; S100A9 P=0.022/0.026). The higher expression was found to be correlated with the clinical characteristics of adenocarcinoma, inflammation and stage IV lesion.

CONCLUSIONSS100A8, S100A9 up-regulation was found in the lung adenocarcinoma and end stage lung cancer tissue, the correlation of which with their higher expression in inflammatory lung tissues may indicate the collaborative effect of inflammation on the progression of cancer.

Adenocarcinoma ; genetics ; metabolism ; pathology ; Aged ; Calgranulin A ; genetics ; metabolism ; Calgranulin B ; genetics ; metabolism ; Female ; Humans ; Immunohistochemistry ; Inflammation ; genetics ; metabolism ; pathology ; Lung Neoplasms ; genetics ; metabolism ; pathology ; Male ; Middle Aged ; Reverse Transcriptase Polymerase Chain Reaction

OBJECTIVETo observe the effect of SSd on reversing the malignant phenotype of HepG2 cells and to investigate its mechanism in order to prove that SSd is a new choice to prevent and treat HCC.

METHODSHepG2 cells were cultured and treated by different concentration (0 mg/L, 2.5 mg/L, 5.0 mg/L, 10.0 mg/L and 20.0 mg/L) of SSd for 24 h, and treated by 10 mg/L of SSd for 0 h, 6 h, 12 h, 24 h, 48 h and 72h respectively. The cell inhibition rates were measured by MTT assay. Then cells were treated by 10 mg/L SSd for 48 hr in experimental group and treated by no SSd as a control, their morphological changes were observed by contrast phase microscope. The concentrations of ALB and AFP in clear supernatant liquid of cells were detected by radio-immunity and chemiluminescence. The cell migration rates were observed by transwell method, the relative expression levels of p27 mRNA were measured by RT-PCR.

RESULTSThe inhibitive effect of 10 mg/L SSd was the most significant among different concentrations ( F = 265.06, P less than 0.01). The shape of HepG2 from experimental group turned into small and round, and their volume ratios of nucleus to plasma decreased. ALB in supernatant liquid of HepG2 was higher ( t = 7.83, P less than 0.05, and its AFP was lower ( t = -10.72, P less than 0.01) as compared to control group. Cells migrated were fewer and p27 mRNA expression of HepG2 was higher in experimental group than that in control group (t = 22.00, P less than 0.05).

CONCLUSIONSSd could reverse the malignant phenotype of HepG2 cells. It was suggested that the up-regulation of p27 mRNA expression play an important role in the differentiation of HepG2 cells treated by SSd.

Carcinoma, Hepatocellular ; pathology ; Hep G2 Cells ; drug effects ; Humans ; Liver Neoplasms ; pathology ; Oleanolic Acid ; analogs & derivatives ; pharmacology ; RNA, Messenger ; genetics ; Saponins ; pharmacology