自然杀伤细胞联合他莫昔芬对乳腺癌细胞的杀伤作用及其机制

发布时间：2018-04-23 09:28

本文选题：乳腺肿瘤 + 他莫昔芬　；参考：《吉林大学学报(医学版)》2017年02期

【摘要】：目的:通过体外实验观察自然杀伤细胞(NK细胞)联合他莫昔芬(TAM)对乳腺癌细胞(BCC)的协同杀伤作用,初步探讨其作用机制。方法:选用3种受体表达程度不同的BCC,设置空白对照组及TAM各浓度/时间梯度实验组,MTT法检测各组细胞增殖抑制率,并确定最终实验浓度为5μmol·L-1。设自然释放组、最大释放组、TAM组、NK细胞组和联合实验组(BCC+NK细胞+TAM),钙黄绿素-AM释放法观察不同效靶比下两者的联合杀伤作用。设空白对照组(NK细胞)、NK细胞+TAM组、NK细胞+BCC组和联合实验组,ELISA法检测各组NK细胞中TNF-α和IFN-γ水平。设空白对照组(NK细胞)、NK细胞+TAM组、NK细胞+BCC组和联合实验组,流式分析法分别检测NK活化性受体(NKp46)、NK抑制性受体(CD158a、CD158b、CD158b2和CD158e)表达水平。设空白对照组(BCC)、BCC+TAM组、BCC+NK细胞组和联合实验组,流式细胞术检测各组细胞中活化性配体(MICA、ULBP1和ULBP2)的表达水平。结果:MTT法检测,TAM对3种BCC的增殖抑制率具有明显的时间和浓度依赖性(P0.05)。钙黄绿素-AM释放法检测,随着效靶比的增加,与NK细胞组比较TAM组BCC杀伤率明显升高(P0.01),且联合组杀伤率明显高于NK细胞组和TAM组(P0.05)。ELISA法检测,与空白对照组比较,各实验组无论有无BCC存在,NK细胞中分泌TFN-α和IFN-γ水平均升高(P0.05或P0.01),且与TAM联合后TFN-α和IFN-γ水平明显升高(P0.05)。流式细胞术检测,与空白对照组比较,各实验组NK细胞中NKp46表达水平均升高(P0.05);各实验组NK细胞中CD158a、CD158b、CD158b2和CD158e表达水平均明显下降(P0.05);各实验组BCC中MICA、ULBP1和ULBP2表达水平均明显升高(P0.05)。结论:NK细胞与TAM联合体外杀伤BCC具有协同作用,其协同作用机制可能为TAM能够通过促进NK细胞分泌TNF-α和IFN-γ而增强其杀伤能力;通过增加NK细胞活化性受体和活化性配体表达水平、降低NK细胞抑制性受体表达水平,进而增加NK细胞的杀伤能力。
[Abstract]:Aim: to observe the synergistic killing effect of natural killer cell (NK) combined with tamoxifen (TAM) on breast cancer cell line BCCs in vitro and to explore its mechanism. Methods: three BCCs with different levels of receptor expression were used to determine the inhibition rate of cell proliferation in each group by setting up blank control group and TAM concentration / time gradient test group. The final experimental concentration was 5 渭 mol L-1. The natural release group, the maximum release group, the TAM group and the combined experimental group were used to observe the combined effect of TAMN and calcitonin -AM release method. The levels of TNF- 伪 and IFN- 纬 in NK cells were detected by Elisa in TAM group and combined experimental group. The NK cell BCC group and the combined experimental group were used to detect the expression of NK cell BCC and CD158b2, CD158b2 and CD158e in NK activated receptor (NKp46) and its inhibitory receptor (CD158b2 and CD158e), respectively. The expression levels of activated ligands MICAULBP1 and ULBP2were detected by flow cytometry. Results the proliferation inhibition rate of Tam on three kinds of BCC was significantly time-and concentration-dependent (P 0.05). The cytotoxicity of BCC in TAM group was significantly higher than that in NK cell group, and the killing rate of BCC in combination group was significantly higher than that in NK cell group and TAM group. The levels of TFN- 伪 and IFN- 纬 in NK cells with or without BCC increased significantly (P0.05 or P0.01), and the levels of TFN- 伪 and IFN- 纬 increased significantly after combined with TAM. Compared with the blank control group, the expression of NKp46 in NK cells in each experimental group was significantly higher than that in the control group by flow cytometry, while the expression levels of CD158b ~ + CD158b ~ (2) and CD158e in NK cells of each experimental group were significantly lower than those of control group (P _ (0.05)), and the expression levels of BCC and ULBP2 in each experimental group were significantly increased (P _ (0.05)). Conclusion there is a synergistic effect between TAM and the fraction of NK cells in killing BCC. The synergistic mechanism may be that TAM can enhance the cytotoxicity of NK cells by promoting the secretion of TNF- 伪 and IFN- 纬 by NK cells. By increasing the expression level of activated receptor and activated ligand of NK cell, the expression level of inhibitory receptor of NK cell was decreased, and the killing ability of NK cell was increased.
【作者单位】：吉林大学第一医院肿瘤中心;吉林大学第一医院生殖中心;
【基金】：吉林省科技厅自然科学基金资助课题(20160101028JC)
【分类号】：R737.9

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