hsa_circ_0000520通过调控miR-556-5p/SLC38A2促进乳腺癌的发生和转移

目的:探究hsa_circ_0000520促进乳腺癌发生和转移的作用机制。方法:双荧光素酶报告基因实验、RNA pull-down实验验证miR-556-5p与hsa_circ_0000520、溶质载体家族38成员2（SLC38A2）的靶向关系。MCF7细胞分为sh-NC组、sh-hsa_circ_0000520组、sh-hsa_circ_0000520+anti-NC组、sh-hsa_circ_0000520+anti-miR-556-5p组，qRT-PCR或Western blot检测细胞中hsa_circ_0000520、miR-556-5p、SLC38A2表达水平；MTT检测、平板克隆形成实验评估细胞增殖能力；划痕愈合实验、Transwell实验评估细胞迁移、侵袭能力。通过裸鼠成瘤实验评估hsa_circ_0000520对miR-556-5p/SLC38A2的调控作用及对移植瘤生长的影响。结果:经验证，MCF7细胞中miR-556-5p与hsa_circ_0000520、SLC38A2均存在靶向关系。与sh-NC组比较，sh-hsa_circ_0000520组可降低MCF7细胞中hsa_circ_0000520、SLC38A2 mRNA和蛋白表达水平、细胞活力、克隆形成数目、划痕愈合率及迁移、侵袭细胞数（P＜0.05），升高miR-556-5p表达水平（P＜0.05）；与sh-hsa_circ_0000520+anti-NC组比较，sh-hsa_circ_0000520+anti-miR-556-5p组可降低MCF7细胞中miR-556-5p表达水平（P＜0.05），升高SLC38A2 mRNA和蛋白表达水平、细胞活力、克隆形成数目、划痕愈合率及迁移、侵袭细胞数（P＜0.05），而对hsa_circ_0000520表达无显著影响（P＞0.05）。裸鼠成瘤实验结果表明，敲低移植瘤中hsa_circ_0000520的表达可升高miR-556-5p表达水平并降低SLC38A2 mRNA和蛋白表达水平，同时降低肿瘤体积和肿瘤重量（P＜0.05）。结论:hsa_circ_0000520可能通过靶向调控miR-556-5p/SLC38A2促进乳腺癌的发生和转移。

hsa_circ_0000520 promotes occurrence and metastasis of breast cancer by regulating miR-556-5p/SLC38A2

Objective:To explore the mechanism of hsa_circ_0000520 in promoting the occurrence and metastasis of breast cancer.Methods:Dual-luciferase reporter gene experiments and RNA pull-down experiments verified the targeting relationship of miR-556-5p with hsa_circ_0000520 and solute carrier family 38 member 2 (SLC38A2).MCF7 cells were separated into sh-NC group,sh-hsa_circ_0000520 group,sh-hsa_circ_0000520+anti-NC group,and sh-hsa_circ_0000520+anti-miR-556-5p group.qRT-PCR or Western blot were applied to detect the expression levels of hsa_circ_0000520,miR-556-5p and SLC38A2 in cells.MTT assay and plate colony formation assay were used to evaluate cell proliferation.Scratch-healing assay and Transwell assay were used to evaluate cell migration and invasion abilities.The nude mice tumorigenesis experiments were applied to evaluate the regulatory effect of hsa_circ_0000520 on miR-556-5p/SLC38A2 and its effect on the growth of xenografts.Results:It had been verified that miR-556-5p had a targeting relationship with hsa_circ_0000520 and SLC38A2 in MCF7 cells.Compared with the sh-NC group,the sh-hsa_circ_0000520 group could reduce the expression levels of hsa_circ_0000520 and SLC38A2 mRNA and protein,cell viability,number of colonies,wound healing rate,and the numbers of migrating and invasive cells in MCF7 cells (P<0.05),increase the expression level of miR-556-5p(P<0.05).Compared with sh-hsa_circ_0000520+anti-NC group,sh-hsa_circ_0000520+anti-miR-556-5p group could reduce the expression level of miR-556-5p in MCF7 cells (P<0.05),increase the expression levels of SLC38A2 mRNA and protein,cell viability,number of colonies,wound healing rate,and numbers of migrating and invasive cells (P<0.05),however,it had no obvious effect on the expression of hsa_circ_0000520 (P>0.05).The results of tumorigenesis experiments in nude mice showed that knocking down the expression of hsa_circ_0000520 in xenograft tumors could increase the expression level of miR-556-5p and decrease the expression levels of SLC38A2 mRNA and protein,and reduce tumor volume and tumor weight (P<0.05).Conclusion:hsa_circ_0000520 may promote the occurrence and metastasis of breast cancer by targeting miR-556-5p/SLC38A2.