lncRNA SNHG3靶向调控miR-340对卵巢癌细胞上皮间质转化的影响

The effect of lncRNA SNHG3 targeted regulation of miR-340 on epithelial-mesenchymal transition of ovarian cancer cells

目的探究长链非编码RNA小核仁RNA宿主基因3(lncRNA SNHG3)对卵巢癌细胞恶性生物学行为的调控机制。方法体外培养人卵巢癌细胞株(SKOV3、A2780、SW626和OVCAR3)和正常卵巢上皮细胞(HOSEpiC),将对数生长期的OVCAR3细胞分为对照组、si-NC组、SNHG3沉默组、miR-NC组、miR-340-5p过表达组、SNHG3沉默+anti-miR-NC组、SNHG3沉默+anti-miR-340-5p组。RT-qPCR检测细胞中lncRNA SNHG3、miR-340-5p表达;双荧光素酶实验验证SNHG3和miR-340-5p的调控作用;CCK-8法、划痕实验和Transwell小室实验检测各组细胞增殖、迁移、侵袭能力;Western blot检测各组细胞中上皮间质转化(epithelial mesenchymal transformation,EMT)相关蛋白[E-钙粘蛋白(E-cadherin)、N-钙粘蛋白(N-cadherin)、波形蛋白(VIM)]表达。裸鼠成瘤实验检测沉默SNHG3的OVCAR3细胞体内生长情况;免疫组织化学法和RT-qPCR检测移植瘤组织中E-cadherin、VIM和lncRNA SNHG3、miR-340-5p表达。结果与HOSEpiC细胞相比,卵巢癌细胞系中lncRNA SNHG3表达增加,miR-340-5p表达降低(P<0.05)。双荧光素酶实验结果证实,miR-340-5p是SNHG3的靶基因。敲低SNHG3或过表达miR-340-5p可降低OVCAR3细胞的增殖活性和迁移、侵袭能力以及N-cadherin、VIM水平,升高E-cadherin水平(P<0.05);下调miR-340-5p可激活EMT,减弱SNHG3沉默对卵巢癌细胞增殖、侵袭和迁移的影响。体内实验结果显示,敲低SNHG3可抑制裸鼠移植瘤生长,并升高肿瘤组织中E-cadherin、miR-340-5p表达,降低VIM表达(P<0.05)。结论敲低SNHG3可能通过上调miR-340-5p,抑制EMT,进而抑制卵巢癌细胞的迁移和侵袭。

Objective To explore the regulatory mechanism of long-chain non-coding RNA small nucleolar RNA host gene 3(lncRNA SNHG3)on the malignant biological behavior of ovarian cancer cells.Methods Human ovarian cancer cell lines(SKOV3,A2780,SW626 and OVCAR3)and normal ovarian epithelial cells(HOSEpiC)were cultured in vitro,and OVCAR3 cells in the logarithmic growth phase were divided into control group,si-NC group,SNHG3 silence group,miR-NC group,miR-340-5p overexpression group,SNHG3 silence+anti-miR-NC group,and SNHG3 silence+anti-miR-340-5p group.RT-qPCR was used to detect the expression of lncRNA SNHG3 and miR-340-5p of cells;Double luciferase experiments was used to verify the regulatory role of SNHG3 and miR-340-5p;CCK-8 method,scratch test and Transwell chamber test were used to detect cell proliferation,migration and invasion abilities of each group;Western blot was used to detect the expression of epithelial-mesenchymal transition(EMT)-related proteins[E-cadherin,N-cadherin and vimentin(VIM)]of cells in each group.Nude mouse tumorigenesis experiment was used to detect the growth of OVCAR3 cells silencing SNHG3 in vivo;Immunohistochemical method and RT-qPCR were used to detect the expression of E-cadherin,VIM,lncRNA SNHG3 and miR-340-5p in transplanted tumor tissues.Results Compared with HOSEpiC cells,the expression of lncRNA SNHG3 in ovarian cancer cell was increased,and the expression of miR-340-5p was decreased(P<0.05).The results of dual luciferase experiments confirmed that miR-340-5p is the target gene of SNHG3.Knock down SNHG3 or overexpression of miR-340-5p could reduce the proliferation activity,migration and invasion capabilities of OVCAR3 cells,as well as the levels of N-cadherin and VIM,and increase the level of E-cadherin(P<0.05).Turn down miR-340-5p to activate EMT and attenuates the effects of SNHG3 silencing on ovarian cancer cell proliferation,invasion and migration.The results of in vivo experiments showed that knocking down SNHG3 could inhibit the growth of transplanted tumors in nude mice,increase the expression of E-cadherin and miR-340-5p in tumor tissues,and reduce the expression of VIM(P<0.05).Conclusion Knock down SNHG3 may inhibit EMT by up regulating miR-340-5p,thereby inhibiting migration and invasion of ovarian cancer cells.