Mechanism of Poly(I∶C)-induced apoptosis in lung adenocarcinoma A549 cells
10.3760/cma.j.issn.1673-422X.2017.05.001
- VernacularTitle:多聚肌苷酸-多聚胞苷酸诱导肺腺癌A549细胞凋亡的机制
- Author:
Liang YI
;
Dan SUN
;
Qian HAN
;
Xiaoyu CHAI
;
Xinmin LIU
- Keywords:
Lung neoplasms;
Apoptosis;
Poly(I∶C)
- From:
Journal of International Oncology
2017;44(5):321-326
- CountryChina
- Language:Chinese
-
Abstract:
Objective To study the effects of Poly(I:C) on lung adenocarcinoma A549 cells viability and illuminate the mechanism of Poly (I:C)-induced apoptosis in A549 cells.Methods A549 cells were transfected with the complex of Poly(I:C) and lipofectamine 3000.The viability of A549 cells was tested by methyl thiazolyl tetrazolium (MTF) method.The apoptotic cells were tested by flow cytometry.The caspase proteins were tested by Western blotting and the expressions of interferon-β (IFN-β) and CXCL-10 were assayed by real-time PCR.After employing the pan-caspase inhibitor Z-VAD-FMK and caspase-8 inhibitor Z-IETD-FMK,the variation of Poly (I:C) proapoptosis in A549 cells was observed.RNA interfering experiments were employed to knock down melanoma differentiation related antigen 5 (MDA5) or retinoic acidinduced gene Ⅰ (RIG-Ⅰ),and the above indexes were tested.Results The viability of A549 cells was significantly reduced to 74.92％ ±--6.24％ after 200 ng/ml Poly (I:C) transfection compared with that before transfection (95.32％ ± 3.05％,t =2.883,P =0.041).The apoptotic rates induced by 100,200,400 ng/ml Poly(I:C) were 9.97％-± 0.88％,23.63％-± 1.41％,32.57％-± 2.39％,respectively.All of them were higher than that in the control group (0.74％-± 0.15％),with significant differences (t =4.489,P =0.002;t =11.616,P =0.000;t =16.932,P =0.000).Besides,the death receptor pathway proteins such as TNF-related apoptosis inducing ligand (TRAIL),cleaved-caspase-8 and cleaved-caspase-3 increased obviously.MDA5/RIG-Ⅰ pathway was also activated dramatically and the expressions of IFN-β,CXCL-10 were significantly up-regulated.The apoptotic rates reduced to 3.17％ ± 0.66％,5.35％ ± 0.64％ with pancaspase inhibitor Z-VAD-FMK and caspase-8 inhibitor Z-IETD-FMK pretreatment,compared with the control group (15.87％ ±0.93％),and the differences were statistically significant (t =8.643,P =0.001;t =6.824,P =0.002).Moreover,the expressions of TRAIL,IFN-β and CXCL-10 induced by Poly (I:C) were inhibited with MDA5 or RIG-Ⅰ depletion.Conclusion Poly(I:C) can reduce the survival rate of A549 cells and promote the apoptosis mainly by activating the death-receptor pathway mediated by MDA5/RIG-Ⅰ probably,which may involve in IFN-β,CXCL-10.
