外泌体APE1对非小细胞肺癌A549细胞顺铂敏感度的影响

戴楠; 赵晓龙; 代晓燕; 李梦侠

doi:10.3971/j.issn.1000-8578.2020.19.1609

外泌体APE1对非小细胞肺癌A549细胞顺铂敏感度的影响

Effect of Exosomal APE1 on Sensitivity of NSCLC A549 Cells to Cisplatin

摘要

摘要:
目的探讨外泌体脱嘌呤脱嘧啶核酸内切酶（APE1）对非小细胞肺癌（NSCLC）A549细胞顺铂（CDDP）敏感度的影响。

方法 Western blot检测顺铂处理和APE1基因重组慢病毒载体转染A549细胞后，细胞及其分泌的外泌体中APE1的变化情况。PKH26对外泌体染色后，激光共聚焦观察其进入细胞内的情况。将外泌体和带GST标签的APE1纯化蛋白（GST-APE1）与A549细胞共培养，观察细胞内GST-APE1的表达情况。分别将顺铂处理后的细胞外泌体（EXO^CDDP）和慢病毒转染细胞产生的外泌体（EXO^APE1）与A549细胞共培养48 h，观察外泌体对A549细胞顺铂敏感度的影响。敲低A549细胞内APE1后，采用APE1的功能抑制剂处理与EXO^APE1共培养的A549细胞，观察处理后A549细胞对CDDP的敏感度以及γ-H2AX蛋白变化情况。

结果高表达APE1的A549细胞产生高表达APE1的外泌体。APE1能够以外泌体包裹的形式被受体细胞所吸收。将EXO^CDDP和EXO^APE1分别与A549细胞共培养后，A549细胞对顺铂的敏感度明显低于对照组。采用APE1碱基切除修复功能抑制剂，能够逆转EXO^APE1所致的细胞耐药性。

结论 A549细胞可通过外泌体传递APE1降低受体细胞对顺铂的敏感度。

Abstract:
Objective To investigate the effect of exosomal Apurinic aprimidinic endonuclease 1 (APE1) on the sensitivity of non-small cell lung cancer (NSCLC) cells to cisplatin(CDDP).
Methods The expression of APE1 in A549 cells and its exosomes were detected by Western blot after cisplatin treatment or APE1 transfection. After PKH26 exosomal staining, laser confocal observation was carried out to observe its location in cells. Exosomes and GST-APE1 were co-cultured with A549 cells to observe the expression of GST-APE1. EXO^CDDP and EXO^APE1 were cultured with A549 cells for 48h to observe their effect on sensitivity of A549 cells to cisplatin. After being co-cultured with EXO^APE1, A549 cells with APE1 knockdown were treated with APE1 functional inhibitors. The sensitivity of A549 cells to CDDP and the change of γ- H2AX protein were observed.
Results A549 cells with high expression of APE1 produced the exosomes with high expression of APE1. APE1 could be absorbed by receptor cells in the form of exosomes. When EXO^CDDP and EXO^APE1 were co-cultured with A549 cells, the sensitivity of A549 cells to cisplatin was significantly lower than that of the control group. The drug resistance of EXO^APE1 co-cultured cells could be reversed by APE1 base excision and repair function inhibitors.
Conclusion A549 cells could transmit APE1 through exosomes to reduce the sensitivity of receptor cells to cisplatin.

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