Objective To develop a method for quantitative real-time detection of perforin(PFR) using polymerase chain reaction(PCR).Methods Gene expression of PFR in peripheral blood mononuclear cell(PBMC) from 25 healthy individuals was detected by PCR.An endogenous"housekeeping" gene GAPDH was used to normalize the results.Results and Conclusion The spontaneous PFR mRNA expression in PBMC in a given sample could be measured by PCR with relativity correlation coefficient of r=1.

0.99.

Purpose:To detect the telomerase activity of lung cancer cells in peripheral blood by TRAP.Methods:The mononuclear cell fraction in peripheral blood was isolated by Ficoll Hypaque gradient centrifugation. Then the telomerase activity of cancer cells in peripheral blood of 49 pre chemotherapy patients was determined by PCR TRAP. Results:In the 49 cases, 65.3%(32/49) cases showed significantly increased telomerase activity, among these cases the telomerase activity was especially high in small cell lung cancer and advanced non small cell lung cancer patients, and telomerase activity in the primary chemotherapy patients was higher than in the recurrent chemotherapy patients. Conclusions:Telomerase activity may be a tumor marker of cancer cells in peripheral blood and can evaluate chemotherapy response in lung cancer.

Objective:To study anti tumor immunity against Lewis lung cancer of dendritic cells derived from proliferated bone marrow cells Methods:After immunization with MUT 1 peptide pulsed DCs (DC MUT 1) intravenously, C57BL/6 mice were challenged with tumor cells subcutaneously to test the immune protection effect Meanwhile, CTL assay was analyzed Results:DC MUT 1 could induce durable and specific antitumor immunity in mice, and the T cells from immunized mice with pulsed DCs showed strong CTL activity Conclusion: An antitumor cellular immunity and a specific immune protection could be induced by immunization with DC MUT 1

Objective:To analyze the drug sensitivity of human lung adenocarcinoma stem cells (LASC) to cisplatin (DDP) and carboplatin (CBP). Methods:Human lung adenocarcinomaic cells SPC-A1,AG,and CPA-Y2 were treated with DDP and CBP. The cell viability of cells was detected by CCK-8 assay. The phenotypic characteristics of drug surviving cells(DSCs)were determined by immunofluorescence staining. The LASC population was then separated by magnetic-activated cell sorting method. The LASC in DSCs was traced by using green fluorescence protein (GFP). The drug sensitivity of DSCs to DDP and CBP was analyzed.Results:The LASC exhibited the phenotypes of bronchioalveolar stem cells (BASC, OCT4~+CCSP~+SP-C~+). After mixture of CD221~+LASC with CD221~-lung adenocarcinoma differentiated cells, the DSC population showed OCT4~+BASC phenotypes. These DSCs were significantly resistant to DDP and CBP.Conclusion:LASC has a high resistance to DDP and CBP. This may be the reason for tumor recurrence after chemotherapy.

OBJECTIVETo evaluate the effect of vascular endothelia1 growth factor (VEGF) on the hematogenous metastasis of non-small cell lung cancer (NSCLC).

METHODSThe identification of lung cancer cells in the peripheral blood were carried out by cytological, immunohistocytologica1 and immunofluorecent stains respectively, following isolation of cytokeratin-expressing cells with magnetic activated cell sorting. The quantification of cancer cells in the blood was performed according to the established flow cytometric assay. The plasma VEGF was measured by commercially available ELISA kit.

RESULTSThe lung cancer cells in the blood, showing a remarkable nuclear polymorphism, expressed the epithelial marker cytokeratin and telomerase reverse transcriptase (hTERT). These cells were stained positive by an NSCLC-specific monoclonal antibody S5Al0-2, but negative by antibodies against CD34 and CD45 antigens. Using the flow cytometric assay, 44 cases (28.6%) of l54 NSCLC patients were found to have cancer cells in their blood, with the incidence of positive cases correlated with the stage of disease. The plasma VEGF level was significantly increased in NSCLC patients in comparison with healthy individuals and patients with benign pulmonary diseases. This level was correlated with the stages of disease in patients with adenocarcinoma. In patiens with cancer cells in their blood, a higher level of plasma VEGF was related with an increased number of cancer cells.

CONCLUSIONThe plasma VEGF level is increased in NSCLC patients with approximate1y one fourth to have cancer cells in the peripheral blood. In these patients, increased VEGF level promotes hematogenous tumor metastasis, as indicated by a much higher number of cancer cells in the blood.

Adult ; Aged ; Aged, 80 and over ; Carcinoma, Non-Small-Cell Lung ; blood ; pathology ; DNA-Binding Proteins ; Endothelial Growth Factors ; blood ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Immunohistochemistry ; Keratins ; analysis ; Lung Neoplasms ; blood ; pathology ; Lymphokines ; blood ; Male ; Middle Aged ; Neoplasm Metastasis ; Neoplastic Cells, Circulating ; chemistry ; pathology ; Telomerase ; analysis ; Vascular Endothelial Growth Factor A ; Vascular Endothelial Growth Factors

BACKGROUNDTo detect dendritic cells (DC)in the peripheral blood and plasma concentration of vascular endothelial growth factor (VEGF) of patients with non-small cell lung cancer (NSCLC) and to evaluate their relationship.

METHODSThe quantitation of DC in the blood was performed in 55 patients with NSCLC, 13 patients with pulmonary benign diseases, and 12 healthy volunteers by a novel flow cytometric assay. The concentration of VEGF in the plasma was measured by ELISA kit.

RESULTSNo significant difference was found in the levels of DC and VEGF between the patients with pulmonary benign diseases and healthy volunteers (P>0.05). In comparison with subjects of healthy volunteers and pulmonary benign diseases, the level of DC was significantly decreased, while that of VEGF was significantly increased in the patients with NSCLC(P < 0.05 to 0.01). The levels of DC and VEGF in the peripheral blood of NSCLC were closely associated with TNM stages and lymph node metastasis. However, no correlation was found among the levels of DC and VEGF and age, gender, cell differentiation and histologic classification. There was a negative correlation between the VEGF concentration and the DC counts.

CONCLUSIONSThe decline of DC count in peripheral blood and the enhancement of plasma VEGF are remarkably related to the malignancy of NSCLC. And VEGF overexpression may be one of mechanisms of DC maturation and differentiation inhibition in patients with NSCLC.

BACKGROUNDDendritic cell (DC)-based immunotherapy is a new approach and effective for some malignant tumors. The aim of this study is to observe the efficacy and toxicity of immunotherapy with carcinoembryonic antigen (CEA) peptide-pulsed DCs in patients with refractory advanced lung cancer.

METHODSLung cancer patients with high CEA expression were enrolled into this project. Autologous DCs were generated from patients' plastic-adherent peripheral blood mononuclear cells and loaded with CEA 5 days later. Cytokine-induced killer cells (CIK) were cultured from non-adherent peripheral blood mononuclear cells. DCs and CIK were transfused to patients. Responses and toxicities were observed.

RESULTSA total of 22 patients with lung cancer received DCs immunotherapy. DCs doses were 2.5×10⁶-9.6×10⁷ (5.03×10⁶). CIK doses were 3.4×10⁸-46×10⁸. CD3, CD8, NK and IFN-γ levels obviously increased after treatment (P < 0.05). The 1-year survival rate was 68.2% (15/22). Main toxicities were fever and rash.

CONCLUSIONSDCs-based immunotherapy is feasible and safe to patients with lung cancer.