Effects of silencing LSD1 and NDRG1 on migration of SKOV3 ovarian cancer cells

SHAO Genbao; WANG Ranran; WEI Ye; JIN Jie; ZHANG Liuping

doi:10.3969/j.issn.1002-266X.2017.38.006

Abstract：Objective To observe the effects of silencing lysine specific demetylase 1 (LSD1) and N-myc down-stream regulated gene 1 (NDRG1) on cell migration of ovarian cancer cells SKOV3 and to investigate the correlation be-tween them. Methods Stable shRNA knockdown technique was employed to generate an inducible and stable SKOV3 cell line expressing tetracycline-regulated shRNA against LSD1 (LSD1-shRNA-SKOV3). The LSD1-shRNA-SKOV3 cells were divided into the control group,Dox group,transfection group,and co-treatment group. Cells in the control group were trea-ted with water,cells in the Dox group were treated with 100 ng/ mL Dox,cells in the transfection group were transfected with NDRG1 siRNA,and cells in the co-treatment group were transfected with NDRG1 siRNA and were added with 100 ng/mL Dox at the same time. The mRNA and protein levels of LSD1 and NDRG1 genes in these cells were detected by real-time fluorescent quantitative PCR and Western blotting,respectively. The histone H3 lysine 4 dimethylation (H3K4me2) levels in the promoter region of NDRG1 gene were measured by chromatin immunoprecipitation (ChIP). The cell migration rate was calculated by the Transwell chamber assay. Results The mRNA levels of LSD1 were 0. 407 ± 0. 029,0. 936 ± 0. 024,0. 413 ± 0. 018,and 0. 941 ± 0. 035,and its protein levels were 0. 306 ± 0. 013,0. 879 ± 0. 036,0. 341 ± 0. 057,and 0. 893 ± 0. 052 in the Dox group,transfection group,co-treatment group,and control group,respectively. Compared with the control group,the mRNA and protein levels of LSD1 decreased in the Dox and co-treatment groups (all P <0. 05). The mRNA levels of NDRG1 were 0. 791 ± 0. 045,0. 107 ± 0. 016,0. 165 ± 0. 021,and 0. 239 ± 0. 027,and its protein levels were 0. 907 ± 0. 005,0. 130 ± 0. 006,0. 216 ± 0. 019,and 0. 358 ± 0. 062 in the Dox group,transfection group,co-treatment group,and control group,respectively. Compared with the control group,NDRG1 mRNA and protein levels significantly changed in the Dox group,transfection group and co-treatment group (all P < 0. 05). The levels of H3K4me2 in the promoter of NDRG1 gene were up-regulated in the Dox group as compared with that of the control group (3. 32 ± 0. 41 vs. 0. 83 ± 0. 17;P < 0. 01). Compared with the control group (68. 91% ± 3. 167%),the migration rate decreased in the Dox group (21. 75% ± 1. 816%)and co-treatment group (40. 13% ± 2. 039%),but increased in the transfection group (79. 13% ± 2. 561%,all P < 0. 05). Furthermore,compared with the Dox group and transfection group,the migration rate significantly changed in the co-treatment group (all P < 0. 05). Conclusions LSD1 gene silen-cing decreases the migration ability of human ovarian cancer SKOV3 cells,and the NDRG1 gene silencing increases the mi-gration ability of human ovarian cancer SKOV3 cells. LSD1 can mediate the decrease of H3K4me2 levels in the promoter of NDRG1 gene,down-regulate the expression levels of NDRG1,and consequently promote the migration of ovarian cancer SKOV3 cells.