摘要
目的探讨树突状细胞(DC)与杀伤细胞(CIK)共培养后对宫颈癌细胞(Hela)凋亡的影响。方法采集宫颈癌患者外周血,分离外周血单个核细胞(PBMC),分别诱导DC和CIK细胞,并扩增培养,显微镜下观察细胞形态,流式细胞仪检测DC细胞表面分子CD8、CD40及CIK细胞CD3、CD8、CD56。采用CCK-8检测DC-CIK细胞对Hela细胞存活率的影响,Annexin V/PI双染检测Hela细胞的凋亡,逆转录聚合酶链反应检测Hela细胞Bax/Bcl-2 m RNA比值和c-myc m RNA水平。结果 DC细胞培养第7天时CD8的阳性表达率为21.62%,CD40的阳性表达率为76.67%。CIK细胞培养第14天时CD3、CD8及CD56的阳性表达率分别为86.85%、82.69%和47.65%。DC-CIK细胞与Hela细胞共培养后,Hela细胞的存活率下降58.40%。流式细胞仪检测Hela+DC-CIK细胞凋亡率增加4.12倍。Hela+DC-CIK共培养的Hela组Bax/Bcl-2 m RNA比值为Hela组的(3.49±0.08)倍(P<0.05),c-myc m RNA水平提高60.72%(P<0.05)。结论该实验成功构建DC-CIK共培养体系,初步验证DC-CIK可能通过调控Bax/Bcl-2及c-myc基因的表达,诱导Hela细胞凋亡。
Objective To explore the influence of co-culture of dendritic cells(DCs)and cytokine induced killer cells(CIKs)on apoptosis of cervical cancer Hela cells.Methods The peripheral blood of cervical cancer patients was collected,and the peripheral blood mononuclear cells(PBMCs)were separated,then DCs and CIKs were induced and cultured.The cell morphology was observed under microscope.Flow cytometry was used to investigate the surface markers CD8 and CD40 of the DCs,and the surface marker CD3,CD8 and CD56 of the CIKs.CCK-8 was used to detect the Hela cell survival rate after being treated with DCs-CIKs.Annexin V/PI double-staining was used to investigate the apoptosis of Hela cells after being treated with DCs-CIKs.The Bax/Bcl-2 mRNA ratio and the level of c-myc mRNA in Hela cells were detected with reverse transcription-polymerase chain reaction(RT-PCR)after treatment with DCs-CIKs.Results The positive expression rates of CD8 and CD40 were 21.62%and 76.67%respectively after DCs were cultured for 7 days.The positive expression rates of CD3,CD8 and CD56 were 86.85%,82.69%and 47.65%respectively after CIKs were cultured for 14 days.After co-cultured with DCs and CIKs,the survival rate of Hela cells decreased by 58.40%(P<0.05),and the apoptosis rate of Hela cells increased 4.12 times.The Bax/Bcl mRNA ratio in the DCs-CIKs and Hela co-culture group was(3.49±0.08)times that in the simple Hela group(P<0.05),and the level of c-myc mRNA increased by 60.72%(P<0.05).Conclusions The co-culture system of DCs-CIKs was successfully constructed.It was basically validated that DCs-CIKs could induce the apoptosis of Hela cells by regulating the expression of Bax/Bcl-2 and c-myc genes.
作者
韦玮
陈心秋
Wei Wei;Xin-qiu Chen(Department of Gynecology and Obstetrics,Liuzhou Worker's Hospital,Liuzhou,Guangxi 545005,China;Department of Gynecologic Oncology,the Affiliated Cancer Hospital of Guangxi Medical University,Nanning,Guangxi 530021,China)
出处
《中国现代医学杂志》
CAS
2018年第20期34-40,共7页
China Journal of Modern Medicine
