| Objective:TGF-β1 is closely associated with the occurrence, development and apoptosis of carcinogenesis. DNA methyltransferases (DNMTs) play an important role in the occurrence and drug-resistance of tumor with the expression of secreted protein acidic and rich in cysteine (SPARC) was decreased. Therefore, in order to explore the mechanism of TGF-β1 regulated the sensitivity of lung adenocarcinoma A549 cells to cisplatin through DNMT1, further to elaborate the mechanism of non-small cell lung cancer resistance to cisplatin and to make the foundation for illustrating the mechanism that lung adenocarcinoma resistance to cisplatin, and then to provide new gene targets for exploring lung cancer chemotherapy drug, exogenous TGF-β1 and its receptor inhibitors were explored to lung adenocarcinoma cells.Methods:Human adenocarcinoma A549 homology cells were cultured (DDP-A549 cells), which were resistanced to cisplatin. After treated with TGF-β1, the changes of TGF-β1 on DDP-A549 cell’s viability and sensitivity to cisplatin were evaluated, and the changes of DNMTs and SPARC in mRNA and proteins levels were respectively detected by RT-PCR and Western blot.Human lung adenocarcinoma A549 cells were cultured and treated with TGF-(31 receptor inhibitor SB431542 (10 μM), the changes of TGF-β1 on A549 cell’s viability and sensitivity to cisplatin were evaluated, and the changes of DNMTs and SPARC in mRNA and proteins levels were respectively detected by RT-PCR and Western blot.Results:It showed that with the treatment of 5 ng/mL and 10 ng/mL TGF-β1 to DDP-A549 cells for 24 h followed by the explosion to different concentrations of cisplatin for 24 h, the IC50 of 5 ng/mL and 10 ng/mL TGF-β1 groups were decreased greatly than that in negative control groups [(22.28±1.60) μmol/L vs (39.05±5.40) μmol/L, P< 0.05; (25.5±3.83) μmol/L vs (39.05±5.40) μmol/L, P< 0.01]; The clone formation number in 5 ng/mL and 10 ng/mL TGF-β1 groups were decreased greatly than that in negative control groups [(752.7±190.4) vs (1218±139.5), P< 0.01; (650.3±144.7) vs (1218±139.5), P< 0.01)] with 5 μmol/L cisplatin; The apoptosis fractions in 5 ng/mL and 10 ng/mL TGF-β1 groups were obviously higher than that in negative control groups [(42.01 ±3.24)% vs (35.5±1.94)%, P< 0.05; (43.76±1.54)% vs (35.5±1.94)%, P< 0.05)] with 15 μmol/L cisplatin for 24 h; The mRNA and proteins expressions of DNMT1 were reduced greatly, and SPARC were increased greatly.It showed that with the treatment in A549 cell with 10 μM SB431542 for 24 h:the IC50 in 10 μM SB431542 groups were increased obviously than that in negative control groups [(31.40±0.17) μmol/L vs (26.14±1.74) μmol/L, P< 0.05]; The clone formation number of 10 μM SB431542 groups were no obvious changed when compared with negative control groups [(405.7±18.15) vs (423.7±24.58), P> 0.05] with 0.5 μmol/L cisplatin; The apoptosis fractions of 10 μM SB431542 groups were no obvious changed when compared with negative control groups [(16.60±3.22)% vs (16.58±2.00)%, P> 0.05] with 10 μmol/L cisplatin for 24 h; The mRNA expressions of DNMT1 were no obviously changed while proteins were increased greatly, however, the mRNA and proteins expressions of SPARC were no obviously changed.It showed that when A549 cells was treated with 5 ng/mL TGF-β1 in the presence and absence of 10 μM SB431542 for 24h, the IC50 of 5 ng/mL TGF-(31 groups were decreased greatly than that in negative control groups and in the combined groups [(15.18±1.46) μmol/L vs (26.14±1.74) μmol/L, P< 0.01; (15.18±1.46) μmol/L vs (24.20±1.78)μmol/L, P< 0.05]; The clone formation number of 5 ng/mL TGF-β1 groups were decreased greatly than that in negative control groups and in the combined groups [(173.7±50.34) vs (423.7±24.58), P< 0.05; (173.7±50.34) vs (491.0±25.94), P< 0.01] with 0.5 μmol/L cisplatin; The apoptosis fractions of 5 ng/mL TGF-J31 groups were obviously higher than that in negative control groups and in the combined groups [(27.32±1.94)% vs (16.58±2.00)%, P< 0.05, (27.32±1.94)% vs (19.66±3.43)%, P< 0.05] with 10 μmol/L cisplatin for 24 h; The mRNA and proteins expressions of DNMT1 in 5 ng/mL TGF-β1 groups were decreased greatly than that in negative control groups and the combined, groups, and the mRNA and proteins expressions of SPARC in 5 ng/mL TGF-β1 groups were increased greatly than that in negative control groups and the combined groups.Conclusion:TGF-β1 can increase the sensitivity of lung adenocarcinoma A549 cells to cisplatin. TGF-β1→DNMT1→SPARC maybe one signal path of TGF-β1 regulate lung cancer susceptibility to cisplatin, it may provide a new idea for studying chemoresistance and help to further clarify the molecular mechanisms of lung cancer resistant to cisplatin. |
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