The Inhibitory Effect Of LITAF Gene On Human Lung Cancer A549Cell In Vitro

Objectives：The relationship between LITAF (lipopolyaccharide-induced tumor necrosisfactor alpha factor) and tumor has now just gained attention. Recent studies havesuggested that LITAF exerts tumor suppressive function. The objectives of thepresent study are to examine the tumor suppressive function of LITAF and associatedchanges in malignant behavior in lung cancer cells where LITAF expression issilenced, and to explore the underlying mechanism.Methods：1ShRNAs for LITAF and control were constructed in retroviral vector andstable cells were made in A549cell lines infected retrovirus and selected withpuromycin.2The biological behavior of the two stable cell lines (A549-LITAF shRNA andA549-shRNA-C) was examined by MTT, anchorage independent growth and woundhealing method.3Microarray, qRT-PCR and Western blot were used to identify downstreamtargets of LITAF.Results：1Greater than80%efficiency of LITAF knockdown was achieved as evidencedby Western blot analysis.2Knockdown of LITAF significantly increased cell proliferation, migrationandanchorage-independent growth of A549cells on soft agars.3A microarray assay has identified many miRNA targets that are upregulated ordownregulated accompanied with silencing LITAF. Among them, miR-15a/128/192/194were reported to have tumor suppressive function and thus further chosen forvalidation using qRT-PCR. Our data revealed the upregulation of of LITAF can increase the abundance of these miRNAs in A549cells, which is blunted byknockdown of LITAF. As Bmi-1, an important regulator for cancer cell renewal,contains taget sequence in its mRNA, assessed if the expression of Bmi-1is regulatedby LITAF. Interestingly, we found that knockdown of LITAF results in upregulationof Bmi-1.Conclusion:Collective, our studies support the notion that LITAF acts as a tumor suppressorand may prosecute such a function by regulating expression of miRNA and Bmi-1.