长链非编码RNA FLG-AS1对宫颈癌细胞侵袭、迁移和STAT3/VEGF通路的影响

Effects of long non-coding RNA FLG-AS1 on invasion,migration and STAT3/VEGF pathway of cervical cancer cells

目的探讨长非编码RNA丝聚蛋白反义RNA 1(FLG-AS1)对宫颈癌细胞侵袭、迁移和信号转导和转录激活因子3(STAT3)/血管内皮生长因子(VEGF)通路的影响。方法采用实时荧光定量PCR(qPCR)检测宫颈上皮细胞End1/E6E7及宫颈癌细胞(CaSki、C-33A、HeLa和SiHa)的FLG-AS1水平,脂质体法向SiHa细胞转染FLG-AS1过表达载体(pcDNA3.1-FLG-AS1组)和空载体(pcDNA3.1组),并设仅经脂质体处理的对照组,MTT比色法评估细胞增殖活力,划痕实验和Transwell小室实验检测划痕愈合率和穿膜细胞数,qPCR和Western blotting检测FLG-AS1、STAT3、磷酸化STAT3(p-STAT3)和VEGF水平。结果宫颈癌细胞的FLG-AS1水平均低于End1/E6E7细胞(P<0.05)。与对照组(1.009±0.121)和pcDNA3.1组(0.994±0.123)的FLG-AS1水平相比,pcDNA3.1-FLG-AS1组SiHa细胞的FLG-AS1水平明显升高(44.634±5.108),差异有统计学意义(P<0.05)。与对照组和pcDNA3.1组相比,pcDNA3.1-FLG-AS1组SiHa细胞转染48、72 h的增殖活力降低(P<0.05)。pcDNA3.1-FLG-AS1组的划痕愈合率和穿膜细胞数分别为(39.817±1.385)%和(160.574±16.492)个,低于对照组的(82.601±4.383)%和(327.106±28.539)个及pcDNA3.1组的(79.518±3.526)%和(348.558±19.861)个(P<0.05)。与对照组和pcDNA3.1组相比,pcDNA3.1-FLG-AS1组的p-STAT3和VEGF水平均降低(P<0.05),而STAT3变化的差异无统计学意义(P<0.05);对照组和pcDNA3.1组上述指标的差异无统计学意义(P>0.05)。结论FLG-AS1在宫颈癌细胞中低表达,过表达其水平可抑制宫颈癌细胞的增殖、迁移和侵袭,可能通过STAT3/VEGF信号途径来发挥抑癌作用。

Objective To investigate the effect of FLG antisense RNA 1(FLG-AS1)on invasion,migration and the signal transducer and activator of transcription 3(STAT3)/vascular endothelial growth factor(VEGF)pathway of cervical cancer cells.Methods FLG-AS1 levels of cervical epithelial cells End1/E6E7 and cervical cancer cells including CaSki,C-33A,HeLa and SiHa cells were detected by real-time quantitative PCR(qPCR).SiHa cells were transfected with FLG-AS1 overexpression vector(pcDNA3.1-FLG-AS1 group)and empty vector(pcDNA3.1 group)by liposome method,and cells only treated with liposome was used as the control group.MTT colorimetric assay was used to evaluate the cell proliferation.Scratch test and Transwell chamber test were used to detect the wound healing rate and number of cells penetrating the membrane.QPCR and Western blotting were used to detect levels of FLG-AS1,STAT3,phosphorylated STAT3(p-STAT3)and VEGF.Results FLG-AS1 levels in cervical cancer cells were lower than that in end1/E6E7 cells(P<0.05).Compared with control group(1.009±0.121)and pcDNA3.1 group(0.994±0.123),FLG-AS1 level of SiHa cells in pcDNA3.1-FLG-AS1 group increased to(44.634±5.1080)folds(P<0.05).Compared with control group and pcDNA3.1 group,the proliferation activity of SiHa cells in pcDNA3.1-FLG-AS1 group was decreased after 48 h and 72 h transfection(P<0.05).The wound healing rate and number of penetrating cells in pcDNA3.1-FLG-AS1 group were(39.817±1.385)%and 160.574±16.492,lower than(82.601±4.383)%and 327.106±28.539 of control group and(79.518±3.526)%and 348.558±19.861 of pcDNA3.1 group(P<0.05).Compared with control group and pcDNA3.1 group,levels of p-STAT3 and VEGF in pcDNA3.1-FLG-AS1 group were decreased(P<0.05),but there was no significant difference in STAT3 changes(P<0.05).There was no significant difference between control group and pcDNA3.1 group(P>0.05).Conclusion FLG-AS1 is low expressed in cervical cancer cells.Overexpression of FLG-AS1 can inhibit the proliferation,migration and invasion of cervical cancer cells,which may play an anti-cancer role through STAT3/VEGF signal pathway.