Effect Of EGCG On The Apoptosis And CUGBP1Protein Expression Of Human Lung Adenocarcinoma A549Cells Induced By LPS

ObjectivesTo study the effects of epigallocatechin-3-gallate (EGCG) on the apoptosis and CUGBP1expression of human lung adenocarcinoma A549cells stimulated by lipopolysaccharide (LPS).Materials and Methods1. MaterialsHuman lung adenocarcinoma A549cells (Medical Center of Affiliated Hospital of Qingdao University, Medical College)RPMI1640and fetal bovine serum (Hyclone)Ribbit anti human CUGBP1polyclone antibody (Abcam)Annexin V-FITC/PI Apoptosis Assay System (BD, America)Steady-Glo(?) Luciferase Assay System (Promega, America)2. MethodsThe A549cell in logarithmic phase were inoculated in96-well culture plate,5x103cell/each well, and treated with LPS (15ug/mL、25ug/mL、50ug/mL), EGCG(100ug/mL、200ug/mL、400ug/ml) and LPS(25ug/mL) plus EGCG (200ug/mL) for4and24hours respectively for the detection of A549cell’s proliferation by MTT assay. The A549cell in logarithmic phase were inoculated in6-well culture plate, randomly divided into4groups, the Control、25ug/ml LPS、200ug/ml EGCG、25ug/ml LPS+200ug/ml EGCG and incubation in37℃5%CO2incubator for4and24h respectively. All cells were collected, washed and stained by Annexin V/7-AAD for the detection of apoptosis ratio. The CUGBP1protein expression of A549cells in the4groups stimulated by LPS and EGCG were examined by immunocytochemistry (ICC).ResultsLPS markedly stimulated the proliferation of A549cells and increased CUGBP1protein expression in the nucleus and cytoplasm, compared with normal control (P<0.01); EGCG inhibited the proliferation and obviously induced the apoptosis of A549cells treated with LPS; EGCG also markedly antagonized the CUGBP1protein expression in nucleus of A549cells incubated with LPS (P<0.01); Quantitative analysis for the CUGBP1protein level showed that at4h or24h, EGCG significantly inhibited the CUGBP1expression of A549cells stimulated by LPS. The relative expression amount of CUGBP1protein stimulated by EGCG and LPS at24h (1210.565±3.46) was significantly higher than that at4h(67.344±3.68)(t=927.164, P<0.001).ConclusionsEGCG could reverse the promotional effects of LPS on the A549cell proliferation and CUGBP1expression and induce the apoptosis.