| Objective:To evaluate lung Aden carcinoma patients with pleural effusion syndrome type distribution; through isolated and cultured tumor cells in pleural effusions, observe its biological characteristics, such as cell cycle, apoptosis; pleural effusion and study of vascular endothelial growth factor (VEGF) levels, pleural tumor cell surface of cancer stem cell marker CD133expression level of disease progression and clinical relevance.Methods:(1) An Experimental Study:Collection Guang An Men Hospital of Traditional Chinese Academy of Oncology27cases of lung cancer patients with pleural effusion, sterile closed drainage in patients with pleural effusion collected by density gradient centrifugation on tumor cells in pleural fluid separation collected and suspension culture, observe the growth and the resulting cells were identified with clinical diagnostic information via HE staining; using Annexin V-FITC/PI double staining were detected by flow cytometry on pleural fluid suspension culture tumor cell suspension cultures of human lung Aden carcinoma A549cells were cultured adherent cell apoptosis in human lung Aden carcinoma A549cells; using PI staining flow cytometry on adherent cultured human lung Aden carcinoma A549cells and suspension cultures pleural tumor cell cycle.(2) Clinical study:60patients with lung Aden carcinoma Guang An Men Hospital oncology patients with pleural effusion cases observation and questionnaire investigation TCM table, analyze the distribution of syndromes patients;27patients received no bacteria closed drainage of pleural fluid extraction, the supernatant was collected in patients with pleural effusion, detected by ELISA experiments which27cases of pleural fluid concentrations of vascular endothelial growth factor (VEGF), and to study the relationship between VEGF concentration of TCM; by flow cytometry on16cases of pleural tumor cells in patients with tumor stem cell surface marker CD133detection.Results:(1) Can be isolated by density gradient centrifugation of pleural effusion tumor cells, tumor cells are able to continue the resulting suspension culture can subculture, but grow more slowly. HE staining of cell morphology was observed under the microscope, we can see larger cell volume, cell shape heterogeneity, large nuclei, there are giant nuclear, nuclear-cytoplasm ratio, nuclear morphology varied, showing dual-core and multi-core, pathologically identified as cancer; combined with patient clinic pathological diagnosis,27patients were primary lung Aden carcinoma, which determine the suspension culture cells of lung Aden carcinoma cells.(2)Apoptosis experiments, A549ce11sadherentgroup, the total rate of apoptosis (6.12±0.19%) was the lowest of the three groups; pleural tumor cell apoptosis rate of the total group (11.87±0.11%)> A549adherent cells group, the re was statistically significant(P<0.05); A549cell suspension group, the total rate of apoptosis(27.17±0.15%)was significantly higher than the other two groups was statistically significant (P<0.05).(3) The cell cycle experiment, two groups of cells in the presence of significant differences in cell content G0-G1phase, A549cells <pleural tumor cells, was statistically significant (P<0.05); two groups of cells in G2phase of the cell contents exist differences, A549cells> pleural tumor cells, with statistical significance (P<0.05); two cells in the S phase of the cell contents differ, A549cells> pleural tumor cells, was statistically significant (P<0.05).(4)60patients with TCM survey, the document appears in the maximum frequency qi deficiency syndrome (43cases,35.2%), followed by the order of the documents appear more blood stasis (24cases,19.7%), sputum Wet-type (21cases,17.2%), Yin type (17cases,13.9%). Most patients are complex syndrome, which mainly have Qi Deficiency Syndromes (15cases,16.9%), QDBS (11cases12.4%), Qi phlegm (13cases,14.6%).(5)Detected by ELISAtest27cases of pleural effusion in patients with lung Aden carcinoma pleural effusion concentrations of vascular endothelial growth factor (VEGF), and it concentration fluctuated with in the range of4.72pg/ml1812.09pg/ml pleural effusion in different patients, the meanto730.31pg/ml.Pleural effusion in patients with stage III VEGF concentration of higher thanIV period, but the difference was not statistically significant(P>0.05).QDBS patients with pleural effusion VEGF mean1425.83pg/ml, significantly higher total compares with the average of27patients, P<0.05,a statistically significant difference.VEGF concentrations of each of the other syndromes with no significant difference between the mean total, P>0.05, not statistically significant.(6) By flow cytometrysurfacemarkerCD133express ion levels of cancer stem cells in16cases of pleural tumor cell lung cancer patients with pleural effusion, and it sex pression level in the range ofl.39%-34.67%. Which pleural tumor cell surface expression of CD133only one case the amount of34.67%, and lexpression was4.50%, the other14cases were between1%to2%, no significant expression.Conclusion:(1) Density gradient centrifugation able to isolate tumor cells in pleural fluid, and can continue to develop, in order to maintain the biological characteristics of pleural tumor cell suspension culture using the method, but the growth rate is relatively slow.(2) Suspension culture of pleural tumor cell apoptosis rates of around10%, in suspension culture can provide suitable conditions for the growth of tumor cells in pleural fluid to simulate the environment in pleural effusions, indicating pleural tumor cell apoptosis with anti-shedding character is tics.(3)About90%of pleural tumor cells quiescent, indicating that most of the cell sin suspension cell cycle arrest, reducing proliferation activities. Because of this characteristic of pleural tumor cells, it has the ability to chemo the rape tic drug resistance.(4)Lung cancer patients with pleural effusion syndrome sin clouding Qi Deficiency, Qi phlegm, blood stasis syndrome, most patients syndromes based virtual reality, the actual situation of inclusions, which advanced with the disease patients was Ted righteousness, evil lag related.(5)Clinical stage and no significant correlation between the concentration of VEGF. QDBS meanl425.83pg/ml, with a total of27patients were significantly higher mean comparison, P<0.05, a statistically significant difference.(6)16cases of lung Aden carcinoma pleural surface of cancer stem cell marker CD133effusion pleural tumor cells, where CD133expression of only one case of pleural tumor cell surface was34.67%, and lexpression was4.50%,the other14patients werebetweenl%to2%, no significant expression. The number of cases detected in this study less, did not accurately reflect the over all relationship between the amount of pleural tumor cells expressing CD133and clinical information between patients, pending further expand the number of cases in the future further research and analysis. |
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