摘要
目的 探讨CircCPA4调节miR-377-3p/GGA3轴对肺癌细胞增殖、凋亡和上皮间质转化(EMT)的影响。方法 qRT-PCR、Western blot分别检测肺癌组织、相邻正常组织、人肺癌A549细胞、人正常肺上皮细胞BEAS-2B中CircCPA4、miR-377-3p、GGA3表达;将A549细胞分为NC组(未做任何处理)、si-NC组(转染si-NC)、si-CircCPA4组(转染si-CircCPA4)、si-CircCPA4+inhibitor NC组(转染si-CircCPA4+inhibitor NC)、si-CircCPA4+miR-377-3p inhibitor组(转染si-CircCPA4+miR-377-3p inhibitor)。双荧光素酶报告基因实验检测CircCPA4、miR-377-3p、GGA3的关系;qRT-PCR检测A549细胞中CircCPA4、miR-377-3p表达;CCK-8法检测A549细胞增殖情况;流式细胞术检测A549细胞凋亡;Transwell检测A549细胞侵袭、迁移数量;Western blot检测A549细胞GGA3、E-cadherin、Vimentin、N-cadherin的蛋白水平。结果 在肺癌组织和肺癌A549细胞中CircCPA4、GGA3水平显著上调(P<0.05),miR-377-3p表达量显著下调(P<0.05)。与NC组、si-NC组比较,si-CircCPA4组A549细胞OD_(450)值、迁移、侵袭细胞数量、CircCPA4表达量、GGA3、N-cadherin、Vimentin蛋白水平显著下降(P<0.05),A549细胞凋亡率、miR-377-3p表达量、E-cadherin蛋白水平显著升高(P<0.05);而miR-377-3p表达下调消除了沉默CircCPA4对A549细胞恶性生物学行为的抑制作用;CircCPA4靶向调节miR-377-3p/GGA3轴。结论 沉默CircCPA4可能通过上调miR-377-3p来抑制GGA3表达,抑制A549细胞增殖、EMT,促进细胞凋亡。
Objective To investigate the regulatory role of Circular RNA carboxypeptidase A4(circCPA4)in the proliferation,apoptosis and epithelial-mesenchymal transformation(EMT)of lung cancer cells by regulating the microRNA-377-3p(miR-377-3p)/Golgi-localized gamma-ear-containing ARF-binding protein 3(GGA3)axis.Methods The mRNA and protein expressions of circCPA4,miR-377-3p and GGA3 in lung cancer tissues,adjacent normal tissues,the human lung cancer cell line A549 and the human bronchial epithelial cells(BEAS-2B)were detected by Quantitative real-time polymerase chain reaction(QRT-PCR)and Western blot respectively.A549 cells were treated with blank control,or transfected with si-NC,si-circCPA,si-circCPA4+inhibitor NC and si-circCPA4+miR-377-3p inhibitor.The relationship between circCPA4,miR-377-3p and that between miR-377-3p and GGA3 was detected by dual-luciferase reporter gene assay.The expressions of circCPA4 and miR-377-3p in A549 cells was detected by qRT-PCR.The proliferation and apoptosis in A549 cells were detected by Cell Counting Kit-8(CCK-8)assay and flow cytometry,respectively.The number of invasive and migratory cells was detected by Transwell assay.Western blot was applied to detect the protein levels of GGA3,E-cadherin,Vimentin and N-cadherin in A549 cells.Results CircCPA4 and GGA3 were significantly upregulated in lung cancer tissues and the lung cancer cell line A549(P<0.05),while miR-377-3p was significantly downregulated(P<0.05).Compared with those treated with blank control or transfected with si-NC,A549 cells transfeted with si-circCPA4 presented significantly lower OD450 value,the number of invasive and migratory cells,relative level of circCPA4,GGA3,and protein expressions of N-cadherin and Vimentin(P<0.05),and significantly higher apoptotic rate,relative level of miR-377-3p and protein expression of E-cadherin(P<0.05).Knockdown of miR-377-3p reversed the inhibitory effects of silenced circCPA4 on malignant behaviors of A549 cells.CircCPA4 targeted the miR-377-3p/GGA3 axis.Conclusion Silencing circC
作者
金曼
王彤辉
任小飞
李淼
JIN Man;WANG Tonghui;REN Xiaofei(Department of Oncology,the Fifth Peopl’s Hospital of Qinghai Province,Qinghai,Xining 810000,China)
出处
《河北医药》
CAS
2023年第18期2731-2735,2740,共6页
Hebei Medical Journal
基金
青海省科技计划项目(编号:2017-ZJ-707)。
关键词
CircCPA4
miR-377-3p
肺癌
增殖
凋亡
上皮间质转化
GGA3
circular RNA carboxypeptidase A4(circCPA4)
microRNA-377-3p(miR-377-3p)
lung cancer
proliferation
apoptosis
epithelial-mesenchymal transformation
Golgi-localized gamma-ear-containing ARF-binding protein 3(GGA3)
