Meng B, Han F, Gao B, Zhuang H, Zhang X Z, Wang Y J, Zhang M
Hepatobiliary and Pancreatic Surgery, Cancer Hospital, Zhengzhou University, Zhengzhou 450000, China.
Zhonghua Zhong Liu Za Zhi. 2021 Nov 23;43(11):1148-1155. doi: 10.3760/cma.j.cn112152-20200222-00116.
To investigate the effects of lncRNA LINC00839 on the proliferation, migration and invasion of hepatocellular carcinoma cells and its mechanism. Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the expression of LINC00839 and miR-3666 in hepatocellular carcinoma tissues and adjacent tissues. Pearson correlation was used to analyze the correlation between LINC00839 and miR-3666 expression in liver cancer tissues. Hepatocellular carcinoma cells MHCC97H were cultured in vitro and divided into si-NC group, si-LINC00839 group, miR-NC group, miR-3666 group, si-LINC00839+ anti-miR-NC group, and si-LINC00839+ anti-miR-3666 group. Methylthiazoletrazolium (MTT) method and clone formation experiment were used to detect cell proliferation. Transwell array was used to detect the cell migration and invasion. Western blot was used to detect the protein expressions of p21, E-cadherin and MMP-2. The double luciferase reporter gene experiment was used to verify the regulatory relationship between LINC00839 and miR-3666. Compared with adjacent tissues, the expression level of LINC00839 in hepatocellular carcinoma tissues increased (2.82±0.27 vs. 0.96±0.10, <0.001), but the expression level of miR-3666 decreased (0.23±0.02 vs. 1.01±0.10, <0.001). The expression levels of LINC00839 and miR-3666 in liver cancer tissue were negatively correlated (r=-0.658, <0.001). The survival rate of MHCC97H cells in the si-LINC00839 group [(53.91±5.41)% vs. (100.53±10.22)%], the number of clones formed (92.0±8.0 vs. 164.0±14.3), the number of migration (131.0±12.7 vs. 247.0±22.4), the number of invasion (66.0±6.4 vs. 120.0±11.6) and the protein level of MMP-2 (0.20±0.02 vs. 0.67±0.06) were lower than those in the si-NC group (<0.001). However, the protein levels of p21 (0.76±0.07 vs. 0.25±0.02) and E-cadherin (0.78±0.08 vs. 0.14±0.01) were higher than those in the si-NC group (<0.001). LINC00839 targeted and negatively regulated the expression of miR-3666. The survival rate of MHCC97-H cells in the miR-3666 group [(47.93±4.86)% vs. (100.11±10.21)%], the number of clone formation (78.0±7.7 vs. 166.0±15.9), the number of migration (117.0±12.1 vs. 250.0±25.0), the number of invasion (57.0±5.7 vs. 121.0±12.3) and the protein level of MMP-2 (0.16±0.01 vs. 0.69±0.07) were lower than those in the miR-NC group (all <0.001). However, the protein levels of p21 (0.83±0.08 vs. 0.24±0.02) and E-cadherin (0.87±0.09 vs. 0.13±0.01)were higher than those in the miR-NC group (all <0.001). The survival rate of MHCC97-H cells in the si-LINC00839+ anti-miR-3666 group [(89.94±9.05)% vs. (54.12±5.39)%], the number of clones (143.0±13.8 vs. 94.0±9.4), the number of migration (208.0±19.8 vs. 129.0±12.6), the number of invasion (108.0±10.1 vs. 65.0±6.4) and the protein level of MMP-2 (0.31±0.03 vs 0.66±0.06) were higher than those in the si-LINC00839+ anti-miR-NC group (<0.001). However, the protein levels of p21 (0.31±0.03 vs. 0.74±0.07) and E-cadherin (0.28±0.03 vs. 0.80±0.08) were lower than those int the si-LINC00839+ anti-miR-NC group (<0.001). Inhibition of LINC00839 expression may inhibit the proliferation, migration and invasion of hepatocellular carcinoma cells by targeting up-regulation of miR-3666 expression.
探讨长链非编码RNA LINC00839对肝癌细胞增殖、迁移和侵袭的影响及其机制。采用实时定量聚合酶链反应(RT-qPCR)检测肝癌组织及癌旁组织中LINC00839和miR-3666的表达。采用Pearson相关性分析肝癌组织中LINC00839与miR-3666表达的相关性。体外培养肝癌细胞MHCC97H,分为si-NC组、si-LINC00839组、miR-NC组、miR-3666组、si-LINC00839+anti-miR-NC组和si-LINC00839+anti-miR-3666组。采用甲基噻唑基四唑(MTT)法和克隆形成实验检测细胞增殖。采用Transwell小室检测细胞迁移和侵袭。采用蛋白质免疫印迹法检测p21、E-钙黏蛋白和基质金属蛋白酶-2(MMP-2)的蛋白表达。采用双荧光素酶报告基因实验验证LINC00839与miR-3666之间的调控关系。与癌旁组织相比,肝癌组织中LINC00839的表达水平升高(2.82±0.27 vs. 0.96±0.10,<0.001),但miR-3666的表达水平降低(0.23±0.02 vs. 1.01±0.10,<0.001)。肝癌组织中LINC00839与miR-3666的表达水平呈负相关(r=-0.658,<0.001)。si-LINC00839组MHCC97H细胞的存活率[(53.91±5.41)% vs. (100.53±10.22)%]、克隆形成数(92.0±8.0 vs. 164.0±14.3)、迁移数(131.0±12.7 vs. 247.0±22.4)、侵袭数(66.0±6.4 vs. 120.0±11.6)及MMP-2蛋白水平(0.20±0.02 vs. 0.67±0.06)均低于si-NC组(<0.001)。然而,p21(0.76±0.07 vs. 0.25±0.02)和E-钙黏蛋白(0.78±0.08 vs. 0.14±0.01)的蛋白水平高于si-NC组(<0.001)。LINC00839靶向并负调控miR-3666的表达。miR-3666组MHCC97-H细胞的存活率[(47.93±4.86)% vs. (100.11±10.21)%]、克隆形成数(78.0±7.7 vs. 166.0±15.9)、迁移数(117.0±12.1 vs. 250.0±25.0)、侵袭数(57.0±5.7 vs. 121.0±12.3)及MMP-2蛋白水平(0.16±0.01 vs. 0.69±0.07)均低于miR-NC组(均<0.001)。然而,p21(0.83±0.08 vs. 0.24±0.02)和E-钙黏蛋白(0.87±0.09 vs.
0.13±0.01)的蛋白水平高于miR-NC组(均<0.001)。si-LINC00839+anti-miR-3666组MHCC97-H细胞的存活率[(89.94±9.05)% vs. (54.12±5.39)%]、克隆数(143.0±13.8 vs. 94.0±9.4)、迁移数(208.0±19.8 vs. 129.0±12.6)、侵袭数(108.0±10.1 vs. 65.0±6.4)及MMP-2蛋白水平(0.31±0.03 vs 0.66±0.06)均高于si-LINC
00839+anti-miR-NC组(<0.001)。然而,p21(0.31±0.03 vs. 0.74±0.07)和E-钙黏蛋白(0.28±0.03 vs. 0.80±0.08)的蛋白水平低于si-LINC00839+anti-miR-NC组(<0.001)。抑制LINC00839表达可能通过靶向上调miR-3666表达来抑制肝癌细胞的增殖、迁移和侵袭。