ABSTRACT
AIM: To investigate the inhibitory effect of Yiqi Huoxue decoction on the malignant biological behavior of lung cancer cells and its mechanism. METHODS: Human lung cancer A549 cells were treated with different doses of Yiqi Huoxue Decoction serum (5%, 10%, 15%). CCK-8 assay, transwell chamber experiment, flow cytometry, Western blot and qRT-PCR method were used to study the effect of different doses of Yiqi Huoxue Decoction-containing serum to cell proliferation, cell migration and invasion, cell apoptosis, PTEN protein and miR-21 expression. RESULTS:Compared with the drug-free serum group, survival rate, migration and invasion ability of A549 cells decreased after treatment with different doses of drug-containing serum. The apoptosis rate of A549 cells increased, PTEN mRNA and the expression of its protein increased, the expression of miR-21 decreased, and the medium-dose (10%) drug-containing serum group had the best effect. After the transfection of miR-21 mimics, miR-21 expression was up-regulated, while PTEN protein expression was down-regulated in cells. PTEN protein expression was up-regulated after treatment with medium-dose (10%) drug-containing serum. CONCLUSION: Yiqi Huoxue Decoction can effectively inhibit the malignant cell biological behavior of human lung cancer A549 cells and may be related to the regulation of the miR-21/PTEN signaling pathway.
ABSTRACT
OBJECTIVE： To investigate the mechanism of calycosin （CA） inhibiting the proliferation and migration of lung adenocarcinoma cells by regulating miR-21/PTEN signaling pathway. METHODS： Using lung adenocarcinoma SPC-A1 cells as objects， cell proliferation was detected by MTT method after treated with different doses of CA （5， 15， 25， 50， 75， 100 μg/mL） for 12， 24， 48， 72 h. Cell survival rate， 30％ cell growth inhibition concentration （IC30） and half inhibition concentration （IC50） were calculated. Transwell migration test was used to detect the migration of cells after treated with low-dose， medium-dose and high-dose of CA （50， 75， 100 μg/mL） for 24 h. The number of stained cells was recorded and inhibition rate of cell migration were calculated. Western blotting assay and real-time PCR were used to detect the expression of miR-21 as well as the proteins and their mRNAs expression of PTEN， VEGF， MMP-9 after treated with low-dose， medium-dose and high-dose of CA （50， 75， 100 μg/mL） for 24 h. After transfected with miR-21 mimics and miR-21 inhibitor， the effects of CA （75 μg/mL） on the expression of miR-21 and the protein expression of PETN， VEGF and MMP-9 were detected. RESULTS： After treated with 50， 75， 100 μg/mL CA for 12， 24， 48 h， 25， 50， 75， 100 μg/mL CA for 72 h， cell survival rate was decreased significantly （P＜0.05 or P＜0.01）. IC30 of CA were 82.24， 50.45， 46.34， 31.81 μg/mL ； IC50 of CA were 108.06， 73.35， 70.08， 49.89 μg/mL during 12-72 h. Compared with normal control group， the number of stained cells in CA groups， protein expression of VEGF in CA low-dose group， expression of miR-21 as well as proteins and their mRNAs expression of VEGF， MMP-9 in CA medium-dose and high-dose groups were decreased significantly； the medium-dose and high-dose groups were significantly less or lower than low-dose group； the high-dose group was significantly less or lower than medium-dose group （P＜0.05 or P＜0.01）. Cell migration rate of CA groups as well as protein and its mRNA expression of PTEN in CA medium-dose and high-dose groups were increased significantly； the medium-dose and high-dose groups were significantly higher than the low-dose group； the high-dose group was significantly higher than the medium-dose groups （P＜0.05 or P＜0.01）. After transfected with miR-21 mimics， expression of miR-21 as well as protein expression of VEGF and MMP-9 were increased significantly in miR-21 mimic group， compared with normal control group； protein expression of PTEN was decreased significantly （P＜0.01）. After intervened by CA， expression of miR-21 as well as protein expression of VEGF and MMP-9 in cells were decreased significantly， compared with miR-21 mimic group； protein expression of PTEN was increased significantly （P＜0.05 or P＜0.01）. After transfected with miR-21 inhibitor， expression of miR-21 as well as protein expression of VEGF and MMP-9 were decreased significantly in miR-21 inhibitor group， compared with normal control group； protein expression of PTEN was increased significantly （P＜0.05 or P＜0.01）. After intervened by CA， the expression of miR-21 and above protein had no significant change in cells， compared with miR-21 inhibitor group （P＞0.05）. CONCLUSIONS： CA can inhibit the proliferation and migration of lung adenocarcinoma SPC-A1 cells in a dose-dependent manner， which may be associated with the regulation of miR-21/PTEN signaling pathway.
