lncRNA AFAP1-AS1在视网膜母细胞瘤组织中的表达及其临床意义

Expression and clinical significance of long non-coding RNA AFAP1-AS1 in retinoblastoma tissues

目的:探讨长链非编码RNA(lncRNA)肌动蛋白纤维相关蛋白1-反义RNA1(AFAP1-AS1)在视网膜母细胞瘤(RB)组织中表达的临床意义及相关机制。方法:收集2015年12月至2019年12月于本院行眼球摘除术的RB患儿36例,取患儿RB组织以及癌旁组织作为研究样本,实时荧光定量PCR(qRT-PCR)法检测组织中AFAP1-AS1表达情况。体外培养人视网膜母细胞瘤株HXO-RB44,分为对照组(不进行转染)、si-NC组(转染siRNA-NC)和si-AFAP1-AS1组(转染siRNA-AFAP1-AS1),qRT-PCR法检测HXO-RB44细胞中AFAP1-AS1表达情况,Transwell法检测HXO-RB44细胞迁移、侵袭情况,蛋白免疫印迹(Western blot)法检测HXO-RB44细胞中肌动蛋白纤维相关蛋白1(AFAP1)、基质金属蛋白酶(MMP)-2、MMP-9蛋白表达水平。结果:与癌旁组织相比,RB组织中AFAP1-AS1表达水平显著升高(P<0.05);RB组织中AFAP1-AS1表达水平与视神经是否浸润、分化程度、有无淋巴结转移有关(P<0.05)。对照组、si-NC组HXO-RB44细胞迁移细胞数、侵袭细胞数及细胞中AFAP1-AS1表达水平、AFAP1、MMP-2、MMP-9蛋白表达水平差异无统计学意义(P>0.05);与si-NC组相比,si-AFAP1-AS1组HXO-RB44细胞迁移细胞数、侵袭细胞数及细胞中AFAP1-AS1表达水平、AFAP1、MMP-2、MMP-9蛋白表达水平显著降低(P<0.05)。结论:AFAP1-AS1在RB组织中高表达,与视神经浸润、分化程度、淋巴结转移相关,抑制其表达可降低AFAP1、MMP-2、MMP-9蛋白表达水平,进而抑制RB细胞HXO-RB44的迁移、侵袭过程,可为减少RB细胞转移提供治疗靶点。

Objective:To investigate the clinical significance and mechanism of long non-coding RNA(lncRNA)actin filament-associated protein 1-antisense RNA1(AFAP1-AS1)expression in retinoblastoma(RB)tissues.Methods:From December 2015 to December 2019,36 children with RB underwent enucleation in our hospital,RB tissues and adjacent tissues were taken as research samples.The expression of AFAP1-AS1 was detected by real-time fluorescence quantitative PCR(qRT-PCR).Human retinoblastoma cell line HXO-RB44 were cultured in vitro and divided into control group(without transfection),si-NC group(transfected with siRNA-NC)and si-AFAP1-AS1 group(transfected with siRNA-AFAP1-AS1).The expression of AFAP1-AS1 in HXO-RB44 cells was detected by qRT-PCR.The migration and invasion of HXO-RB44 cells were detected by Transwell method.Western blot was used to detect the expression levels of actin filament-associated protein 1(AFAP1),matrix metalloproteinase(MMP)-2 and MMP-9 in HXO-RB44 cells.Results:The expression level of AFAP1-AS1 in RB tissues was significantly higher than that in adjacent tissues(P<0.05).The expression level of AFAP1-AS1 in RB tissues was related to optic nerve invasion,differentiation and lymph node metastasis(P<0.05).There was no significant difference between the control group and the si-NC group in the number of migration cells,the number of invasive cells,the expression level of AFAP1-AS1,the protein expression levels of AFAP1,MMP-2 and MMP-9 between the control group and the si-NC group(P>0.05).Compared with those in si-NC group,the number of migration cells,the number of invasive cells and the expression level of AFAP1-AS1,the protein expression levels of AFAP1,MMP-2 and MMP-9 were significantly lower than those in si-AFAP1-AS1 group(P<0.05).Conclusion:AFAP1-AS1 is highly expressed in RB tissues,which is related to optic nerve invasion,differentiation and lymph node metastasis.Inhibition of AFAP1-AS1 can reduce the expression levels of AFAP1,MMP-2 and MMP-9,and then inhibit the migration and invasion of RB cells HXO-RB44,which can provide therapeutic targets for reducing RB cell metastasis.