Experimental Study on Inhibition Effects of the XAF1 Gene against Lung Cancer Cell Proliferation


Objective: To investigate the effect of high expression of XAF1 in vivo or in vitro on lung cancer cell growthand apoptosis.
Methods: 1. The A549 human lung cancer cell line was transfected with Ad5/F35 - XAF1, or Ad5/F35 – Null at the same multiplicity of infection (MOI); (hereinafter referred to as transient transfected cell strain);XAF1 gene mRNA and protein expression was detected by reverse transcription polymerase chain reaction (RTPCR)and Western blotting respectively. 2. Methyl thiazolyl tetrazolium (MTT) and annexin V-FITC/PI doublestaining were used to detect cell proliferation and apoptosis before and after infection of Ad5/F35 - XAF1 withWestern blotting for apoptosis related proteins, caspase 3, caspase - 8 and PARP. 3. After the XAF1 gene wastransfected into lung cancer A549 cells by lentiviral vectors, and selected by screening with Blasticidin, reversetranscription polymerase chain reaction (RT-PCR) and Western blotting were applied to detect mRNA andprotein expression, to establish a line with a stable high expression of XAF1 (hereinafter referred to as stableexpression cell strain). Twenty nude mice were randomly divided into groups A and B, 10 in each group: A549/XAF1 stable expression cell strain was subcutaneously injected in group A, and A549/Ctrl stable cell line stableexpression cell strain in group B (control group), to observe transplanted tumor growth in nude mice.
Results:The mRNA and protein expression of XAF1 in A549 cells transfected by Ad5/F35 - XAF1 was significantly higherthan in the control group. XAF1 mediated by adenovirus vector demonstrated a dose dependent inhibition oflung cancer cell proliferation and induction of apoptosis. This was accompanied by cleavage of caspase -3, -8, -9and PARP, suggesting activation of intrinsic or extrinsic apoptotic pathways. A cell strain of lung cancer highlyexpressing XAF1 was established, and this demonstrated delayed tumor growth after transplantation in vivo.
Conclusion: Adenovirus mediated XAF1 gene expression could inhibit proliferation and induce apoptosis in lungcancer cells in vitro; highly stable expression of XAF1 could also significantly inhibit the growth of transplantedtumors in nude mouse, with no obvious adverse reactions observed. Therefore, the XAF1 gene could become anew target for lung cancer treatment.