miR-548t-3p靶向CITED2调控甲状腺癌细胞的凋亡研究

Study on miR-548t-3p Targeting CITED2 to Regulate Apoptosis of Thyroid Cancer Cells

目的探讨miR-548t-3p调控甲状腺癌细胞凋亡的潜在机制。方法选取甲状腺癌患者62例,检测其癌组织和癌旁组织中miR-548t-3p的表达水平。过表达或敲低miR-548t-3p后,检测甲状腺癌细胞SW579、B-CPAP、8305C、BHT101、CAL-62、KHM-5M、TPC-1、FTC-133、HTh-7的凋亡水平。通过miRDB在线分析和荧光素酶报告实验鉴定miR-548t-3p的底物。结果miR-548t-3p在甲状腺癌组织的表达水平显著高于甲状腺癌旁组织(P<0.05)。过表达miR-548t-3p后,甲状腺癌细胞SW579、B-CPAP、8305C、BHT101、CAL-62、KHM-5M、TPC-1、FTC-133、HTh-7的凋亡水平均下降(P<0.05);而敲低miR-548t-3p后,甲状腺癌细胞的凋亡水平均上升(P<0.05)。过表达miR-548t-3p后,miR-548t-3p能够显著降低SNRK、VASP、CITED2的mRNA水平(P<0.05)。敲低SNRK、VASP、CITED2后,仅当敲低CITED2时,甲状腺癌细胞SW579的凋亡水平显著下降(P<0.05)。过表达CITED2后,甲状腺癌细胞SW579的凋亡水平显著上升(P<0.05)。敲低miR-548t-3p后,CITED2的蛋白表达水平显著上升。过表达miR-548t-3p后,CITED2的蛋白表达水平显著下降。此外,miR-548t-3p靶向CITED2的3端非编码区(P<0.05)。同时敲低miR-548t-3p和CITED2,SW579细胞的凋亡水平无显著改变(P>0.05);同时过表达miR-548t-3p和CITED2,SW579细胞的凋亡水平无显著改变(P>0.05)。结论miR-548t-3p通过靶向CITED2 mRNA的3端非编码区,降解了CITED2 mRNA并减少了CITED2的蛋白表达,最终抑制了甲状腺癌细胞的凋亡。

Objective To explore the potential mechanism of miR-548t-3p regulating the apoptosis of thyroid cancer cells.Methods A total of 62 patients with thyroid cancer were enrolled,and the expression levels of miR-548t-3p in cancer tissues and para-cancerous tissues were detected.After overexpression or knockdown of miR-548t-3p,the apoptosis levels of thyroid cancer cells SW579,B-CPAP,8305C,BHT101,CAL-62,KHM-5M,TPC-1,FTC-133,and HTh-7 were detected.The miR-548t-3p substrate was identified by miRDB online analysis and luciferase report experiment.Results The expression level of miR-548t-3p in thyroid cancer tissues was significantly higher than that in adjacent thyroid cancer tissues(P<0.05).After overexpression of miR-548t-3p,the apoptosis level of thyroid cancer cells SW579,B-CPAP,8305C,BHT101,CAL-62,KHM-5M,TPC-1,FTC-133,HTh-7 decreased(P<0.05);after knocking down miR-548t-3p,the apoptosis level of thyroid cancer cells increased(P<0.05).After overexpression of miR-548t-3p,miR-548t-3p can significantly reduce the mRNA levels of SNRK,VASP,and CITED2(P<0.05).After knocking down SNRK,VASP,CITED2,only when knocking down CITED2,the apoptosis level of thyroid cancer cell SW579 decreased significantly(P<0.05).After overexpression of CITED2,the apoptotic level of thyroid cancer cell SW579 increased significantly(P<0.05).After knocking down miR-548t-3p,the protein expression level of CITED2 increased significantly.After overexpression of miR-548t-3p,the protein expression level of CITED2 decreased significantly.In addition,miR-548t-3p targets the 3rd non-coding region of CITED2(P<0.05).Knock down miR-548t-3p and CITED2 at the same time,the apoptosis level of SW579 cells did not change significantly(P>0.05);while overexpressing miR-548t-3p and CITED2,the apoptosis level of SW579 cells did not change significantly(P>0.05).Conclusion miR-548t-3p degrades CITED2 mRNA and reduces the protein expression of CITED2 by targeting the 3 non-coding region of CITED2 mRNA,and finally inhibits the apoptosis of thyroid cancer cells.