miR-324-5p通过靶向调控DAPK1降低胰腺癌细胞的放射敏感性

miR-324-5p decreases the radiosensitivity of pancreatic cancer cells by targeting DAPK1

目的:探讨死亡相关蛋白激酶1(death-associated protein kinase 1,DAPK1)在胰腺癌(pancreatic cancer,PaC)细胞放射敏感性中的作用,验证miR-324-5p通过靶向调控DAPK1影响胰腺癌细胞放射敏感性的机制。方法:通过生物信息学预测靶向DAPK1的miRNAs,并利用双荧光素酶报告基因检测miR-324-5p对DAPK1的调控作用。在PANC-1和MIA PaCa-2细胞中过表达miR-324-5p和DAPK1或抑制miR-324-5p后,对各细胞株进行放射诱导,检测细胞增殖和凋亡情况,以及凋亡相关分子的表达情况。结果:GEO数据集结果显示,胰腺癌组织中miR-324-5p的表达水平高于正常组织。与正常胰腺导管上皮细胞系(HPDE6-C7)相比,胰腺癌细胞系(Capan-1、Bxpc-3、PANC-1和MIA PaCa-2)中miR-324-5p表达水平更高(P均<0.001)。双荧光素报告基因检测结果表明,miR-324-5p靶向DAPK1的3'UTR,并且可下调DAPK1的表达。细胞实验结果证实,过表达miR-324-5p通过靶向调控DAPK1降低放射诱导的细胞凋亡和DNA的损伤,进而降低了胰腺癌细胞的放射敏感性。结论:miR-324-5p通过负调控DAPK1降低胰腺癌细胞对放射的敏感性,从而影响DNA修复和细胞凋亡。miR-324-5p/DAPK1途径可能为胰腺癌的靶向治疗提供了潜在的治疗靶点。

Objective:To explore the role of death-associated protein kinase 1(DAPK1)in the radiosensitivity of pancreatic cancer(PaC)cells,and to demonstrate the mechanism by which microRNA(miR-324-5p)affects the radiosensitivity of PaC cells by targeting DAPK1.Methods:Bioinformatics was used to predict the microRNAs targeting DAPK1,and dual-luciferase reporter gene assay was used to detect the regulatory effect of miR-324-5p on DAPK1.After cells were overexpressed miR-324-5p and DAPK1 or inhibited miR-324-5p in PANC-1 and MIA PaCa-2 cell lines,and irradiated,cell proliferation,apoptosis,and the expression of apoptosis-related molecules were detected.Results:The results of GEO data set showed that the level of miR-324-5p in pancreatic cancer tissues was higher than that in normal tissues.Compared with normal pancreatic ductal epithelial cell lines(HPDE6-C7),pancreatic cancer cell lines(CAPAN-1,BXPC-3,PANC-1,and MIA PACA-2)showed higher expression of miR-324-5p(all P<0.001).Dual luciferin reporter gene assay results showed that miR-324-5p targeted the 3'UTR of DAPK1 and down-regulated the expression of DAPK1.Cell experiments showed that overexpression of miR-324-5p reduced radiation-induced apoptosis and DNA damage by targeting DAPK1,thereby reducing the radiosensitivity of PaC cells.Conclusion:miR-324-5p reduces the sensitivity of pancreatic cancer cells to radiation through negatively regulating DAPK1,which provides novel insights for the regulation of miR-324-5p and DAPK1 levels.The miR-324-5p/DAPK1 pathway may provide a potential therapeutic target for targeted therapy of pancreatic cancer.