Growth Suppression And Chemosensitivity Of Lung Cancer Cells Induced By Recombinant Adenovirus Expressing ING4 And HIL-24 In Vitro

Objective:To study the growth suppression and chemosensitivity of lung cancer cells (A549) induced by recombinant adenovirus expressing ING4 and hIL-24 (Ad-ING4-IRES- hIL-24) in vitro.Methods: Recombinant replication-incompetent adenovirus expressing ING4 and hIL-24 (Ad-ING4-IRES-hIL-24) was constructed and transfected into QBI-293A cells.The adenovirus completed the amplification in QBI-293A cells and the titration was also detected. The expression of ING4 and hIL-24 in QBI-293A cells was identified by RT-PCR. After the collection of Ad-ING4-IRES-hIL-24,we use it in order to infect the human pulmonary carcinoma cell line A549 and human embryonic lung fibroblast cell line WI-38 in vitro.The transcription of objective gene was detected by RT-PCR.The influences of two kinds of cell growth which made by Ad-ING4-IRES-hIL-24 were tested by MTT essay.The morphological changes before and after the infection were observed by using light microscope and fluorescence microscope.The apoptotic effect were evaluated by flow cytometry (FCM) and laser scanning confocal microscopy (LSCM). Caspase3,Bcl-2 and Bax were explored by means of immunocytochemistry for the purpose of understanding the main mechanism of apoptosis induced by Ad-ING4-IRES-hIL-24. We also use Ad-ING4-IRES-hIL-24 combined with cisplain in order to observe the chemosensitivity of lung cancer cells. The inhibition of cell growth and the apoptotic effect were tested by MTT essay and flow cytometry.Laser scanning confocal microscopy (LSCM) was used to detect the impact on the apoptosis of A549 cells.Results: After 48h of the amplification of Ad-ING4-IRES-IL-24 in QBI-293A cells,we could see the cells becoming round.The cells gathered like grapes under the light microscope.We could observe the green light under fluorescence microscope which means high efficiency of infection.ING4 and hIL-24 were proved to be successfully transcribed in QBI-293A cells and A549 cells.The titration was 108 pfu/mL.The growth of A549 cells was suppressed obviously after transfection of Ad-ING4-IRES-hIL-24 and it showed time dependence and dose dependence. However we could not find the same phenomenon on WI-38 cells. Ad-ING4-IRES-hIL-24 enhanced the apoptosis and growth inhibition of A549 than single gene therapy.Cell apoptosis factors such as Caspase3 and Bax/Bcl-2 expression were significantly elevated.It may be the main mechanism of apoptosis. Ad-ING4-IRES- hIL-24 combined with cisplain could obviously inhibit the growth and induce the cell apoptosis than any single group.It showed the chemosensitivity on lung cancer cells.Conclusion: Recombinant adenovirus expressing ING4 and hIL-24 (Ad-ING4- IRES-hIL-24) induces growth suppression and apoptosis in A549 cells and can remarkably increase the drug sensibility of the lung cancer cells.It is a potential effective approach in the treatment of pulmonary carcinoma.