| Lung cancer in the incidence and mortality of malignant tumors is in the first place, of which non small cell lung cancer accounted for new cases of lung cancer 80%-85%, about 1/3 lung cancer patients diagnosed with locally advanced. Effective rate of traditional chemotherapy and survival is not satisfactory, and most of the chemotherapy drμgs in the gastrointestinal tract and blood system toxicity is very large, often drμg application dose limiting toxic side effects, including most of patients on chemotherapy prohibitive. The pathogenesis of cancer is extremely complex, can not rely on a single drμg or treatment and achieved satisfactory effect. Therefore, in the case of no increase in patients with toxic and side effects, to find a drμg that can increase the effect of chemotherapy, improve the effective rate of chemotherapy and prolong the survival time of patients become the research focus of cancer doctors.In recent years, paclitaxel(PTX) is widely used in the treatment of non small cell lung cancer and the anticancer mechanism: paclitaxel can act on microtubules and tubulin, inhibition of microtubule depolymerization, the microtubule bundle arrangement abnormalities, leading to loss of normal function of spindle, induce cell death. Paclitaxel combined with platinum is one of the most commonly used chemotherapy regimens in the treatment of non small cell lung cancer. Compared with cisplatin and other non small cell lung cancer, paclitaxel has the advantages of significant anticancer effect and low side effect, which has been widely used in clinical practice.With the further development of epigenetic regulation and control agents, the clinical research of the comprehensive treatment plan including HDAC inhibitors is of practical significance, and VPA is the new and high safety HDAC inhibitor. Sodium valproate(VPA) belong to the short chain fatty acids of HDAC inhibitors, scholars at home and abroad application of VPA on a wide variety of hematologic and solid malignancies for basic research, demonstrate the obvious inhibition of tumor. Sodium valproate compared with other drμgs, has low toxicity, and good efficiency Aim:Our in vitro, sodium valproate monotherapy and in combination with paclitaxel on lung cancer A549 cells. By MTT method and flow cytometry technology observation of sodium valproate combined effects of single drμg and drμg on A549 cell cycle arrest and apoptosis, using fluorescence quantitative PCR and Western blot detection of the two drμgs on the proliferation of lung cancer cell line A549 Bcl-2, Bax and Caspase3 gene expression and protein expression, whether VPA monotherapy and in combination with paclitaxel can increase lung cancer cell apoptosis and its possible mechanism, explore new ideas for lung cancer therapy and for the lung cancer clinical antitumor treatment and provide a theoretical basis. Method:1 Cell source: Lung cancer A549 cells presented by the research foundation of Chengde Medical College2 Experimental groups: Combination group, blank control group, paclitaxel group, sodium valproate group, paclitaxel and valproate3 Effect of sodium valproate MTT detection method on proliferation of lung cancer A549 cells4 Effect of flow cytometry to detect the apoptosis of A549 cells and the cell cycle5 The determination of differences in expression of Bcl-2, Bax cells and Caspase3 gene by fluorescence quantitative PCR method6 Determination of expression of Bax and Caspase3 protein in Bcl-2 cells were Western, blot7 Statistical analysis: each experiment repeated at least three times, using the statistical data of SPSS19.0, respectively, with the mean and standard deviation(s) data representation. Comparison between groups using one-way ANOVA, SNK method comparison between the two groups, P<0.05 said the difference was statistically significant. Result:1 Effect of MTT assay for the detection of sodium valproate on the proliferation of lung cancer A549 cells: screening out the drμgs for 24 h ic20, respectively: sodium valproate for 5.80mmol/L, paclitaxel 0.05μg/mL, and taking them as the drμg concentration of subsequent experiments in each intervention group. MTT assay with different concentrations of sodium valproate in A549 cells for 24 h, 48 h and 72 h after inhibition rate, results are shown, at the same time, with the increase of concentration of valproic acid, inhibition rate is gradually increased, differences were statistically significant(P < 0.05); the same concentration of valproic acid, with the extension of time, inhibition rate is gradually increased, the difference is statistical significance(P < 0.05), indicating that VPA on the proliferation of lung cancer A549 cells with inhibitory effect and dose dependence and time dependence.2 Flow cytometric detection of sodium valproate combined with paclitaxel on apoptosis and cell cycle in lung cancer A549 cells.: rabbits were divided into four groups, blank control group, paclitaxel group(0.05μg/ml), valproate group 5.80mmol/L, combined medication group(PTX 0.05 μg/ml+VPA 5.80 mmol/L), the group for 24 h flow cytometry detection confirmed sodium valproate and paclitaxel combined with promoting apoptosis of lung cancer cells, four groups of total apoptosis rate assumes the trend of escalation and combined treatment group the total apoptosis rate the highest, and the difference was statistically significant(P < 0.05). Compared with the blank control group, paclitaxel group G2 / M phase cells increased significantly, sodium valproate in G0 / G1 phase cells increased significantly(P < 0.05), and the combination group of cells in G0 / G1 phase percentage increased significantly and compared with sodium valproate group is more obvious(P < 0.05). Compared with the traditional.3 Fluorescence quantitative PCR and Western blot to detect the two drμgs combined effects on the expression of Bcl-2, Bax and Caspase3 gene and protein in A549 lung cancer cells: experimental group: ibid, 24 h, and blank compared to the control group, the expression of bcl-2 gene and protein in each treatment group A549 cells decreased in and combination group of lowest of each treatment group A549 cells of Caspase3 and Bax gene and protein expression increased expression of and combined medication group was the highest(P < 0.05. The difference between the groups was statistically significant). With the combination of drμgs, the expression of Bcl-2 gradually decreased, and the expression of Caspase3 and Bax increased gradually. Conclusion:Sodium valproate can inhibit the proliferation of A549 cells and showed a dose dependence and time dependence, to A549 cell growth arrest at G0 / G1 phase and sodium valproate and paclitaxel combined application of to inhibit the proliferation of lung cancer cells have a synergistic effect. The mechanism may be related to the expression of apoptosis factor Bax and Caspase3 up-regulated in A549 cells, down-regulation of Bcl-2 expression in A549 cells is related to. |
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