Effect of methotrexate packaged by tumor derived microparticles combined with radiotherapy on proliferation of cancer stem cells in lung adenocarcinoma in vitro
10.3760/cma.j.issn.1006-9801.2016.10.002
- VernacularTitle:含甲氨蝶呤的肿瘤源性微粒联合放疗体外对肺腺癌干细胞增殖的影响
- Author:
Xuesong ZHENG
;
Ying WANG
;
Yiliang MENG
;
Youke XIE
- Publication Type:Journal Article
- Keywords:
Lung neoplasms;
Methotrexate;
Radiotherapy;
Tumor cell-derived microparticles;
Cell cycle;
Neoplastic stem cells
- From:
Cancer Research and Clinic
2016;28(10):654-658,663
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the effect of methotrexate packaged by tumor derived microparticles (T-MP MTX) combined with radiotherapy on lung cancer stem cell (CSC) in vitro. Methods T-MP MTX was prepared from non-small cell lung cancer A549 cells. Proliferative changes of A549 cells, bronchial epithelial cells H460 and 16HBE cells treated by T-MP MTX were assayed by MTT method. Cell cycles of A549 cells in blank group and T-MP MYX group were examined by fluorescence activated cell sorting (FACS). The effect of T-MP MTX combined with radiotherapy on CSCs was assessed by tumor sphere formation experiment and animal experiment. The expressions of stemness relative genes (such as β-catenin, Nanog, SOX-2 and KLF4) were measured by Western blot. Results T-MP MTX dose-dependently inhibited the cell growth in A549 cells, but didn't in H460 cells and 16HBE cells. The S cycle ratio of A549 cells in blank group and T-MP MYX group measured by FACS were (15.83±3.14)%and (47.47±6.69)%, respectively. S cycle ratio of T-MP MYX group was notably higher compared with that of blank group (t=7.411, P=0.002). Further study revealed that the number of tumor sphere in blank group, control group, 2 Gy group, 4 Gy group and 6 Gy group was (268.9±22.4), (172.4±18.7), (48.3±5.1), (16.3±3.5) and (5.1±3.1), respectively. The number of tumor sphere in other groups was decreased compared with that in blank group (F=228.291, P=0.000). The numbers of tumor sphere in 2 Gy group, 4 Gy group and 6 Gy groups was also reduced compared with that in control group. Importantly, the number of tumor sphere in these groups were decreased dramatically as the dose of radiotherapy increased (F=95.142, P=0.000). The results of tumor sphere volume were similar with the number of tumor sphere. Western blot experiment showed that T-MP MTX treatment in A549 cells decreased the expression of stemness relative genes (β-catenin, Nanog, SOX-2 and KLF4), and its role was reinforced when radiotherapy was combined. Animal experiment implied that activity of luciferase in T-MP MTX group was decreased compared with that in blank group (P=0.000), and the activity of luciferase in T-MP MTX plus 2 Gy group was reduced significantly (t=6.887, P=0.002). Conclusions T-MP MTX has a potential to sensibilize radiotherapy, and it will synergistically inhibit the proliferation of CSCs when combined with radiotherapy. Moreover, its mechanism may be related with T-MP MTX activating CSCs from hypometabolism state and blocking process of cell cycle.
