抑制lncR-HOXA-AS3表达的人前列腺癌细胞增殖、迁移、侵袭能力及EMT相关蛋白表达变化

Proliferation,migration,invasion abilities and expression changes of EMT-associated proteins of human prostate cancer cell line LNCaP with inhibition of lncR-HOXA-AS3 expression

目的观察抑制长链非编码RNA(long non-coding RNA,lncR)HOXA-AS3表达的人前列腺癌(prostate cancer,PCa)细胞系LNCaP增殖、迁移、侵袭能力及上皮间质转化(epithelial-mesenchymal transition,EMT)相关蛋白的表达变化。方法本研究受试细胞为人PCa细胞系LNCaP。将对数生长期LNCaP细胞分为2组,分别转染lncRHOXA-AS3沉默质粒si-HOXA-AS3(观察组)和阴性对照质粒si-NC(对照组),转染48 h时采用qRT-PCR法检测lncR-HOXA-AS3,分别于转染24、48、72 h时采用CCK8实验测算细胞增殖活性(光密度OD值),转染48 h时采用细胞划痕实验观察细胞迁移能力,转染48 h时采用Transwell实验观察细胞侵袭能力,转染48 h时采用WESTERN Blotting法检测两组EMT相关蛋白E-钙黏蛋白(E-cadherin)、波形蛋白(Vimentin)、纤维连接蛋白(Fibronectin)。结果与RWPE-1相比,DU-145、LNCaP、C4-2B中lncR-HOXA-AS3相对表达量均升高(P均<0.05)。与对照组相比,转染48 h时观察组细胞lncR-HOXA-AS3相对表达量降低(P<0.05)。与对照组比较,转染24、48、72 h时观察组细胞OD值降低(P均<0.05)。与对照组比较,转染48 h时观察组划痕愈合率小、侵袭穿膜细胞数目少(P均<0.05)。与对照组比较,转染48 h时观察组LNCaP细胞E-cadherin蛋白相对表达量升高,Vimentin、Fibronectin蛋白相对表达量表达降低(P均<0.05)。结论沉默lncR-HOXA-AS3表达能抑制LNCaP细胞的增殖、迁移及侵袭,其机制可能为lncRHOXA-AS3促进LNCaP细胞E-cadherin蛋白表达、抑制Vimentin及Fibronectin蛋白表达。

Objective To observe the proliferation,migration,invasion abilities and expression changes of epitheli⁃al-mesenchymal transformation(EMT)-related proteins in human prostate cancer(PCa)cell line LNCaP with inhibition of expression of long non-coding RNA(lncR)HOXA-AS3.Methods The cells in this study were human PCa cell line LN⁃CaP.LNCaP cells in the logarithmic growth phase were divided into two groups:observation group(transfected with lncRHOXA-AS3 silencing plasmid si-HOXA-AS3)and control group(transfected with negative control plasmid si-NC).At 48 h of transfection,lncR-HOXA-AS3 was detected by qRT-PCR.At 24,48 and 72 h of transfection,the cell prolifera⁃tion activity(optical density OD value)was measured by CCK-8 assay.At 48 h of transfection,the cell migration ability was observed by cell Scratch test.At 48 h of transfection,the cell invasion ability was observed by Transwell test.Western blotting was used to detect the EMT-associated proteins E-cadherin,Vimentin,and Fibronectin in the two groups at 48 h after transfection.Results Compared with RWPE-1,the relative expression levels of lncR-HOXA-AS3 in DU-145,LN⁃CaP and C4-2B increased(all P<0.05).Compared with the control group,the relative expression of lncR-HOXA-AS3 in the observation group decreased at 48 h after transfection(P<0.05).Compared with the control group,the OD values of the observation group decreased at 24,48 and 72 h after transfection(all P<0.05).Compared with the control group,the scar healing rate and the number of invasive transmembrane cells in the observation group were lower at 48 h after transfection(all P<0.05).Compared with the control group,the relative expression of E-cadherin protein increased and the rela⁃tive expression levels of Vimentin and Fibronectin proteins decreased at 48 h after transfection in LNCaP cells of the obser⁃vation group(all P<0.05).Conclusion Silencing the expression of lncR-HOXA-AS3 can inhibit the proliferation,mi⁃gration and invasion of LNCaP cells.The mechanism may be that lncR-HOXA-AS3 promotes the expression of E-cadherin protein in LNCaP cells and inhibits the expression levels of Vimentin and Fibronectin proteins.