Effect Of N-( 3-trifluoromethyl-phenyl) Retinamide On Differentiation,apoptosis And Expression Of Osteopontin In Human Pulmonary Adenocarcinoma A549 Cells And Their Mechanism

Objective To investigate the effect of novel synthetized [N-(3-trifluoromethyl-phenyl) retinamide, NTFMP] on the proliferation, differentiation, apoptosis and the expression of osteopontin(OPN) of the human lung adenocarcinoma A549 cells in vitro and their mechanism.Methods A549 cells were treated with ATRA and NTFMP, respectively. The proliferation of A549 cells was observed by MTT assay. The morphology of A549 was observed by light microscope and the secretory volume of carcinoembryonic antigen(CEA) was detected by kinetics cytometry. The related protein expression was analyzed by western blot. A549 cells were treated with NTFMP for 3 days, then added with PMA and PD98059 and took the pictures for observing the cell morphous at 0min, 5min, 10min, 45min and 2hour respectively. The expression of apoptosis-related proteins, OPN, P-ERK_1/2 and ERK_1/2 in A549 cells treated with NTFMP were detected by western blot.Results The NTFMP could inhibit the proliferation of A549 cells in a dose-dependent manner. The CEA secretory volume of A549 cells decreased evidently in NTFMP group at 10mg/L(P<0.05). Compared with control group, A549 cells in NTFMP group became lesser, more irregular, non-overlapping and monolayer. The results of western blot indicated that NTFMP induced apoptosis with the decreased expression of Bcl-2, NF-κB (P<0.05)and increased expression of Bax, without significant alteration of expression of Bak, P53, Caspase-3 and Caspase-9. It also indicated that the phosphorylation of ERK_1/2 and the expression of OPN down-regulation significantly in A549 cells treated with 7.5mg/L of NTFMP for 3 days(P<0.05), however, no effect was observed with ERK_1/2. Then we added in stimulator of ERK_1/2 pathway (PMA) , we found that the phosphorylation of ERK_1/2 and the expression of OPN inceasing again. But application of PD98059 caused a dramatic reduction of OPN expression which was induced by PMA until 45min, especially at 10min. Then with the prolong of time, the effect of PMA becomes to weaken while that of PD98059 to enhance.Conclusion NTFMP could inhibit the A549 cells proliferation and induce differentiation and apoptosis, the mechanism of which possibly is to decrease the expression of OPN by inhibiting the phosphorylation of ERK_1/2.