[Effect of Ad-Gax transfection on apoptosis of human lung adenocarcinoma A549 cells and its mechanism].
BACKGROUND
Apoptosis is closely related to development of lung cancer. It is a strategy of lung cancer therapy to induce apoptosis. The aim of this study is to explore the effects of growth arrest-specific homeobox (Gax) transfection on apoptosis and expression of Bcl-2 and Bax proteins of human lung adenocarcinoma A549 cells.


METHODS
A549 cells were transfected with Gax gene by a replication-deficient adenovirus expressing the hemagglutinin-tagged Gax cDNA (Ad-Gax). Apoptosis of A549 cells was observed by transmission electronic microscope and terminal-deoxynucleotidyl transferase mediated nick end labeling (TUNEL) positive staining. Apoptotic rate of A549 cells was evaluated by flow cytometry. Expressions of Bcl-2 and Bax proteins in A549 cells were detected by immunocytochemistry.


RESULTS
Before Ad-Gax transfection, none or few of TUNEL-positive A549 cells were detected. After Ad-Gax transfection, a marked increase in TUNEL-positive staining occurred, especially at 24 h later. The ratio of apoptosis of A549 cellsin non-transfection group and transfection groups at 12 h, 24 h, 48 h were 0.25%, 12.57%, 17.29%, 15.03%, respectively. Compared with non-transfection group, the apoptotic rates of transfection groups increased significantly (Chi-square value was 7.357, 11.126 and 9.943 respectively, P < 0.01). The average optical density (AOD) of Bcl-2 protein in A549 cells in non-transfection group and transfection groups at 12 h, 24 h, 48 h were 2.02±0.07, 1.79±0.02, 1.25±0.51 and 1.21±0.24 respectively. Compared with non-transfection group, AOD of Bcl-2 protein in A549 cells in transfection groups decreased significantly (t value was 6.651, 7.089 and 7.438 respectively, P < 0.01). On the other hand, Bax protein expression in transfection groups increased, the AODs of Bax were 4.49±0.61, 4.24±0.37 and 3.95±0.43, respectively. Compared with non-transfection group (3.12±0.42), AOD of Bax protein in A549 celle in transfection groups increased significantly (t value was 7.469, 7.287 and 6.473 respectively, P < 0.01). In the Ad-Gax transfection groups the lower Bcl-2/Bax ratio was, the higher the apoptotic rate of A549 cells was (r=-0.49, P < 0.01).


CONCLUSIONS
Ad-Gax transfection can induce A549 cells apoptosis. Possible mechanism is that Gax can downregulate Bcl-2 protein expression and upregulate Bax protein expression, and A549 cells apoptosis is related to the Bcl-2/Bax ratio.
Laboratory of Molecular Biology of Institute of Integrated Traditional Chinese and Western Medicine, Huashan Hospital, Fudan University, Shanghai 200040, P.R.China.
Shijin
S
Xia
Xiantao
X
Tai
Jingcheng
J
Dong
Guisheng
G
Qian
Mingdong
M
Hu