摘要
目的:探讨环状RNA BIRC6(CircBIRC6)对非小细胞肺癌(NSCLC)细胞(A549)顺铂(DDP)耐药性的影响。方法:RT-qPCR法检测DDP耐药及敏感肺癌组织与细胞中CircBIRC6表达,体外培养人NSCLC细胞株A549及DDP耐药细胞A549/DDP,将A549/DDP细胞分为Control组、pcDNA组、pcBIRC6组、si-CircBIRC6组、si-NC组、si-BIRC6+inhibitor NC组,si-BIRC6+miR-126-5p inhibitor组。RT-qPCR检测CircBIRC6、miR-126-5p表达;CCK-8法检测细胞增殖及对DDP的IC_(50)值;流式细胞术检测细胞凋亡;Western blot检测JARID2、Cyclin D1、P21、P-gp蛋白表达;双荧光素酶报告基因实验验证CircBIRC6、JARID2与miR-126-5p的靶向关系。结果:与DDP敏感肺癌组织相比,CircBIRC6在DDP耐药肺癌组织中表达水平显著升高,与A549细胞相比,A549/DDP细胞中CircBIRC6表达水平及DDP对A549/DDP细胞的IC_(50)值显著升高(P<0.05);双荧光素酶报告基因实验证实CircBIRC6、JAR-ID2与miR-126-5p存在靶向关系。与pcDNA组相比,pcBIRC6组A549/DDP细胞增殖、CircBIRC6、JARID2、Cyclin D1与P-gp蛋白表达及IC_(50)值显著升高,细胞凋亡率及miR-126-5p、P21蛋白表达水平显著降低(P<0.05);与si-NC组相比,si-BIRC6组细胞增殖、CircBIRC6、JARID2、Cyclin D1与P-gp蛋白表达及IC_(50)值显著降低,细胞凋亡率及miR-126-5p、P21蛋白表达水平显著升高(P<0.05);下调miR-126-5p表达可逆转抑制CircBIRC6表达对A549/DDP细胞的增殖抑制作用和凋亡促进作用。结论:CircBIRC6在A549/DDP细胞中高表达,可通过靶向抑制miR-126-5p表达,上调JARID2表达,促进A549/DDP细胞对DDP的耐药性。
Objective:To investigate the influence of circular RNA BIRC6(CircBIRC6)on cisplatin(DDP)resistance of non-small cell lung cancer(NSCLC)cells(A549).Methods:RT-qPCR was used to detect the expression of CircBIRC6 in DDP-resistant and sensitive lung cancer tissues and cells.Human NSCLC cell line A549 and DDP-resistant cells A549/DDP were cultured in vitro.A549/DDP cells were grouped into Control group,pcDNA group,pcBIRC6 group,si-CircBIRC6 group,si-NC group,si-BIRC6+inhibitor NC group,and si-BIRC6+miR-126-5p inhibitor group.RT-qPCR was applied to detect the expression of CircBIRC6 and miR-126-5p;CCK-8 assay was applied to detect cell proliferation and IC_(50)value for DDP;flow cytometry was applied to detect apoptosis;Western blot was applied to detect the protein expressions of JARID2,Cyclin D1,P21,and P-gp;dual-luciferase reporter gene assay was applied to verify the targeting relationship between CircBIRC6,JARID2 and miR-126-5p.Results:Compared with DDP-sensitive lung cancer tissues,the expression level of CircBIRC6 in DDP-resistant lung cancer tissues was greatly higher;compared with A549 cells,the expression level of CircBIRC6 in A549/DDP cells and the IC_(50) value of DDP on A549/DDP cells were greatly higher(P<0.05);the dual-luciferase reporter gene assay confirmed that CircBIRC6 and JARID2 had a targeting relationship with miR-126-5p.Compared with the pcDNA group,the A549/DDP cell proliferation,the protein expressions of CircBIRC6,JARID2,Cyclin D1 and P-gp,and IC_(50) value in the pcBIRC6 group were greatly higher,the apoptosis rate and the expression levels of miR-126-5p and P21 proteins were greatly lower(P<0.05);compared with the si-NC group,the A549/DDP cell proliferation,the protein expressions of CircBIRC6,JARID2,Cyclin D1 and P-gp,and IC_(50) value in the si-BIRC6 group were greatly lower,the apoptosis rate and the expression levels of miR-126-5p and P21 proteins were greatly higher(P<0.05);down-regulation of miR-126-5p expression reversed the proliferation-inhibiting and apoptosis-promoting effects of inhibiting CircBIRC6 expression on A549/DDP cells.Conclusion:CircBIRC6 is highly expressed in A549/DDP cells,which can inhibit the expression of miR-126-5p by targeting,up-regulate the expression of JARID2,and promote the resistance of A549/DDP cells to DDP.
作者
党乙
张梁
翟恒钰
李文海
DANG Yi;ZHANG Liang;ZHAI Hengyu(Xi'an International Medical Center Hospital,Shaanxi Xi'an 710100,China)
出处
《河北医学》
CAS
2022年第11期1768-1774,共7页
Hebei Medicine
基金
陕西省重点研发计划项目,(编号:S2022-YF-YBSF-0779)。
