沉默lncRNA PSMB8-AS1通过调控miR-144-5p表达对甲状腺癌细胞恶性生物学行为的影响

Silencing lncRNA PSMB8-AS1 affects malignant biological behaviors of thyroid cancer cells regulating miR-144-5p expression

目的探讨长链非编码RNA(lncRNA)PSMB8-AS1通过靶向miR-144-5p对甲状腺癌细胞增殖和迁移的影响。方法研究时间为2023年3月至2024年5月,地点:青岛大学附属医院科研中心。采用GeneCards数据库分析PSMB8-AS1在甲状腺癌组织中的表达。采用实时定量聚合酶链反应(qPCR)检测PSMB8-AS1在人正常甲状腺上皮细胞HTori-3以及甲状腺癌细胞系(8505C、FTC-133、KTC-1、TPC-1)中的表达。采用LncRBase和Cancer LncRNA Census数据库预测PSMB8-AS1的靶基因,用双荧光素酶基因报告实验验证。将FTC-133细胞根据转染物的不同分为si-NC组和si-PSMB8-AS1组,CCK-8法和划痕实验分别检测低表达PSMB8-AS1对FTC-133细胞增殖和迁移的影响,PSMB8-AS1与miR-144-5p的靶向关系。qPCR检测低表达PSMB8-AS1对FTC-133细胞中miR-144-5p和ITGA3 mRNA表达的影响。Western blot法检测低表达PSMB8-AS1对FTC-133细胞中ITGA3、Cyclin A、CDK2、FOXC2、Hedgehog蛋白表达的影响。结果PSMB8-AS1在甲状腺癌组织中表达高于癌旁组织(P<0.01)。PSMB8-AS1在甲状腺癌细胞质8505C、FTC-133、KTC-1、TPC-1中表达均高于HTori-3细胞(均P<0.01)。PSMB8-AS1能够靶向结合miR-144-5p(P<0.01)。si-PSMB8-AS1组FTC-133细胞中PSMB8-AS1表达低于si-NC组[(1.07±0.54)比(6.69±1.16),t=8.78,P<0.01]。低表达PSMB8-AS1后,si-PSMB8-AS1组FTC-133细胞的增殖水平均低于si-NC组(均P<0.01)。si-PSMB8-AS1组FTC-133细胞的迁移率低于si-NC组[(27.89±6.01)%比(68.08±6.16)%,t=4.67,P<0.01]。si-PSMB8-AS1组细胞中miR-144-5p表达高于si-NC组,ITGA3 mRNA表达低于si-NC组(t=7.45、3.13,均P<0.01)。低表达PSMB8-AS1能够抑制FTC-133细胞中ITGA3、Cyclin A、CDK2、FOXC2、Hedgehog蛋白表达(均P<0.01)。结论PSMB8-AS1在甲状腺癌组织和细胞系中表达显著上调,低表达PSMB8-AS1可通过靶向调控miR-144-5p抑制FTC-133细胞的增殖和迁移。

Objective To investigate the effects of long noncoding RNA(lncRNA)PSMB8-AS1 on the proliferation and migration of thyroid cancer cells by targeting miR-144-5p.Methods The study was conducted from March 2023 to May 2024 at the Scientific Research Center of the Affiliated Hospital of Qingdao University.The GeneCards database was used to analyze the expression of PSMB8-AS1 in thyroid cancer tissue.The real-time quantitative polymerase chain reaction(qPCR)was used to detect the expressions of PSMB8-AS1 in human normal thyroid epithelial cells HTori-3 and thyroid cancer cell lines(8505C,FTC-133,KTC-1,and TPC-1).The target gene of PSMB8-AS1 was predicted using the LncRBase and Cancer LncRNA Census databases and verified by the dual luciferase gene reporter assay.The FTC-133 cells were divided into a si-NC group and a si-PSMB8-AS1 group according to the different transfectants.The CCK-8 method and scratch assay were used to detect the effects of low-expression PSMB8-AS1 on the proliferation and migration of the FTC-133 cells.qPCR was used to detect the effects of low-expression PSMB8-AS1 on the expression of miR-144-5p and ITGA3 mRNA in the FTC-133 cells.Western blot was used to detect the effect of low-expression PSMB8-AS1 on the expression of ITGA3,Cyclin A,CDK2,FOXC2,and Hedgehog proteins in the FTC-133 cells.Results The expression of PSMB8-AS1 in the thyroid cancer tissue was higher than that in the adjacent tissue(P<0.01).The expressions of PSMB8-AS1 in the 8505C,FTC-133,KTC-1,and TPC-1 cells were higher than that in the HTori-3 cells(all P<0.01).PSMB8-AS1 could target and bind to miR-144-5p(P<0.01).The expression of PSMB8-AS1 in the FTC-133 cells in the si-PSMB8-AS1 group was lower than that in the si-NC group[(1.07±0.54)vs.(6.69±1.16);t=8.78;P<0.01].The migration rate of FTC-133 cells in the si-PSMB8-AS1 group was lower than that in the si-NC group[(27.89±6.01)%vs.(68.08±6.16)%;t=4.67;P<0.01].The expression of miR-144-5p in the FTC-133 cells in the si-PSMB8-AS1 group was higher than that in the si-NC group,and the expression of ITGA3 mRNA was lower than that in the si-NC group(t=7.45 and 3.13;both P<0.01).Low-expression PSMB8-AS1 could inhibit the expressions of ITGA3,Cyclin A,CDK2,FOXC2,and Hedgehog proteins in the FTC-133 cells(all P<0.01).Conclusions PSMB8-AS1 expression is significantly upregulated in thyroid cancer tissue and cell lines.Low-expression PSMB8-AS1 can inhibit the proliferation and migration of FTC-133 cells targeting and regulating miR-144-5p.