Objective: To investigate the phenotype and molecular mechanism of DCA on MDS cell model, and to study the response of chemotherapeutic medicines to MDS cells through multiple dimensions, such as cell proliferation, invasion, migration and apoptosis, thus revealing the molecular mechanism of DCA treatment of MDS and its relationship with SHP-1 gene methylation.
Methods: MTT assay was used to determine the survival rate of MDS cells after treated by different concentrations of DCA. The effect of DCA on the invasion and migration of MDS cells was detected by Transwell assay method. Apoplexin V-FITCPI was used to detect apoptosis, the MDS treatment on the mechanism of DCA was investigated by Western blot and Real-time PCR experiment.
Results: According to the experiment, it was found that tumor proliferation could be inhibited when MDS skm-1 cells was treated by DCA, and the absorbance was lower and the inhibitory effect was more obvious in the 2.0, 5.0 μmol/L DCA group than in the 0.5 μmol/L DCA group and the negative control group. Compared with the control group, the number of MDS skm-1 cells crossing through the transwell upper chamber was significantly decreased after DCA application. After treated with 0.5, 2.0 and 5.0 μmol/L DCA, the apoptosis rate of MDS cells was 4.54%, 9.31% and 16.58% respectively, while the apoptosis rate of the control group was 3.20%, which shows the apoptosis rate increased significantly with the concentration of DCA. After treatment of MDS cell lines with different concentration of DCA, the methylation status of SHP-1 gene was decreased with the increase of drug concentration, the expression of SHP-1 was increased, the expression of STAT3 was decreased and the level of phosphorylation was decreased.
Conclusion: By analyzing the phenotypic response of DCA treatment on MDS cells, it was found that interfere with MDS can be performed by inhibiting proliferation, metastasis, and inducing apoptosis in a dose-dependent way. It revealed that the molecular mechanism by DCA treatment can improve the methylation of SHP-1 gene and inhibit the expression of p-STAT3.
题目: 地西他滨抑制SHP-1基因甲基化对MDS细胞株Skm-1增殖及凋亡影响.
目的: 探讨DCA作用于MDS细胞模型后的表型及分子机制，通过细胞增殖、侵袭迁移及细胞凋亡等多重维度研究化疗药物作用于MDS细胞后的反应，揭示DCA治疗MDS的分子机理及其与SHP-1基因甲基化的关系.
方法: 采用MTT法测定不同浓度DCA处理后的MDS细胞存活率，采用Transwell实验检测DCA对MDS细胞侵袭和迁移能力的影响，应用AnnexinV-FITC/PI法检测细胞凋亡情况，Western blot免疫蛋白印迹和real time PCR实验研究DCA治疗MDS的作用机制.
结果: 实验发现，DCA作用于MDS skm-1细胞能抑制肿瘤增殖，且2.0及5.0 μmol/L DCA组相对0.5 μmol/L DCA组与阴性对照组相比吸光度降低、抑制效应更加明显。与对照组比较，应用DCA后MDS skm-1细胞穿过Transwell上室底膜的数量显著下降。在0.5、2.0及5.0 μmol/L DCA处理后，MDS细胞凋亡率依次为4.54%、9.31%及16.58%，对照组细胞凋亡率为3.20%。这表明，细胞凋亡率随DCA浓度显著增加。在经不同浓度DCA处理的MDS细胞系中，SHP-1基因甲基化状态随着药物浓度升高而减弱，SHP-1表达量升高，STAT3表达量减少，而STAT3磷酸化水平则受到明显抑制.
结论: 通过对DCA作用于MDS细胞后的表型反应分析，发现其主要通过抑制增殖和转移并诱导凋亡来干预MDS，并具有药物剂量依赖性，这揭示了DCA治疗能够改善SHP-1基因的甲基化进而抑制p-STAT3表达的分子机制.