Fu Q, Cheng J, Zhang J D, Zhang Y L, Chen X B, Xie J G, Luo S X
Gastrointestinal Surgery Center, Henan Cancer Hospital, the Affiliated Cancer Hospital of Zhengzhou University, Zhengzhou 450002, China.
Department of Oncology, Zhengzhou Central Hospital, Zhengzhou 450007, China.
Zhonghua Zhong Liu Za Zhi. 2020 May 23;42(5):369-375. doi: 10.3760/cma.j.cn112152-112152-20190118-00017.
To investigate the effects and the mechanism of FoxO6 on the proliferation and invasion of colorectal cancer cells. FoxO6 siRNA was transfected into colorectal cancer cell HCT116 and SW480. The overexpression vector pcDNA.3.1-c-Myc was constructed and co-transfected into HCT116 and SW480 cells with FoxO6 siRNA. Real-time fluorescent quantitative PCR (RT-qPCR) and western blot were used to detect the mRNA and protein expressions of FoxO6, c-Myc, and p21 in HCT116 and SW480 cells. Bromodeoxyuridine (BrdU) was used to detect cell proliferation and Transwell assay was performed to detect the invasion ability of these cells. SW480 cells transfected with FoxO6 shRNA lentivirus (LV-FoxO6) and were injected into the right armpit of BAL b/c nude mice to construct a tumor-bearing mode and the tumor volumes were measured on the days of 10, 13, 16, 19, 22, and 25 after injection. The FoxO6 mRNA were 0.91±0.04, 1.72±0.07, and 2.03±0.06, and protein expression were 0.70±0.04, 1.35±0.08, and 1.56±0.07 in normal colon cell FHC, colorectal cancer cells HT116 and SW480, respectively. The protein and mRNA levels of FoxO6 in HCT116 and SW480 were significantly higher than those in FHC (both <0.05). Knockdown of FoxO6 in HCT116 and SW480 cells decreased the mRNA and protein expressions of FoxO6 (both <0.05), the cell proliferation ability (absorbances were 0.26±0.07 and 0.27±0.06, both <0.05), cell invasion ability (the invaded cell numbers were 42.3±3.3 and 45.7±4.1, both <0.05), and the mRNA and protein expressions of c-Myc, while increased the mRNA and protein expressions of p21 (both <0.01). Overexpression of Myc in FoxO6 silenced HCT116 and SW480 cells decreased the expression of p21, while increased the cell proliferation ability (absorbances were 0.54±0.09 and 0.58±0.07, both <0.01) and invasion ability (the invaded cell numbers were 79.2±5.9 and 80.5±6.4, both <0.01). On the 25th day after cell inoculation in nude mice, the tumor volume of LV-FoxO6 group was (190.6±36.2) mm(3), significantly lower than (437.8.6±69.2) mm(3) of LV-NC group (<0.05). FoxO6 promotes the proliferation and invasion of colorectal cancer cells through facilitating c-Myc mediated p21 expression inhibition.
为研究FoxO6对大肠癌细胞增殖和侵袭的影响及其机制。将FoxO6 siRNA转染至大肠癌细胞HCT116和SW480。构建过表达载体pcDNA.3.1-c-Myc，并与FoxO6 siRNA共转染至HCT116和SW480细胞。采用实时荧光定量PCR（RT-qPCR）和蛋白质印迹法检测HCT116和SW480细胞中FoxO6、c-Myc和p21的mRNA及蛋白表达。用溴脱氧尿苷（BrdU）检测细胞增殖，并进行Transwell实验检测这些细胞的侵袭能力。将转染了FoxO6 shRNA慢病毒（LV-FoxO6）的SW480细胞注射到BAL b/c裸鼠右腋窝构建荷瘤模型，并在注射后第10、13、16、19、22和25天测量肿瘤体积。正常结肠细胞FHC、大肠癌细胞HT116和SW480中FoxO6的mRNA分别为0.91±0.04、1.72±0.07和2.03±0.06，蛋白表达分别为0.70±0.04、1.35±0.08和1.56±0.07。HCT116和SW480中FoxO6的蛋白和mRNA水平显著高于FHC（均P<0.05）。敲低HCT116和SW480细胞中的FoxO6可降低FoxO6的mRNA和蛋白表达（均P<0.05）、细胞增殖能力（吸光度分别为0.26±0.07和0.27±0.06，均P<0.05）、细胞侵袭能力（侵袭细胞数分别为42.3±3.3和45.7±4.1，均P<0.05）以及c-Myc的mRNA和蛋白表达，同时增加p21的mRNA和蛋白表达（均P<0.01）。在FoxO6沉默的HCT116和SW480细胞中过表达Myc可降低p21的表达，同时增加细胞增殖能力（吸光度分别为0.54±0.09和0.58±0.07，均P<0.01）和侵袭能力（侵袭细胞数分别为79.2±5.9和80.5±6.4，均P<0.01）。在裸鼠细胞接种后第25天，LV-FoxO6组的肿瘤体积为（190.6±36.2）mm³，显著低于LV-NC组的（437.8±69.2）mm³（P<0.05）。FoxO6通过促进c-Myc介导的p21表达抑制来促进大肠癌细胞的增殖和侵袭。
