目的: 探讨IL-12 逆转化疗药物对NK细胞免疫功能的抑制及其机制。方法: 纯化的NK细胞在PMA和Ionomycin刺激条件下、加或不加化疗药物顺铂（cisplatin，DDP）和IL-12，用ELISA法检测培养上清中IFN-γ 和TNF-α 的分泌水平，流式细胞术检测IFN-γ 和TNF-α、肿瘤坏死因子相关的凋亡诱导配体（TNF-related apoptosis inducing ligand, TRAIL）和转录因子（T-bet 和p-STAT-4）的表达百分率；纯化NK细胞加或不加化疗药物和IL-12 预处理48 h，流式细胞术检测其对白血病Jurkat 细胞株的杀伤效应。结果: 化疗药物明显抑制NK细胞分泌IFN-γ、TNF-α 以及TRAIL 的表达，IL-12 可以明显逆转DDP对NK细胞分泌IFN-γ、TNF-α 和TRAIL 的抑制（P<0.05 或P<0.01）。DDP抑制IFN-γ 和TNF-α 的转录因子p-STAT-4 的表达，而IL-12 通过上调磷酸化STAT-4 恢复了NK细胞的IFN-γ 和TNF-α 的分泌功能（P<0.01）。杀伤实验显示，DDP抑制NK细胞对白血病细胞Jurkat 的杀伤，而IL-12 通过上调TRAIL恢复NK细胞的杀伤功能（P<0.05 或P<0.01）。结论: 化疗药物DDP抑制NK细胞的杀伤功能以及细胞因子（IFN-γ 和TNF-α）的分泌，IL-12 通过上调TRAIL和转录因子STAT-4 的磷酸化恢复NK细胞的免疫功能，为临床应用IL-12 重建肿瘤化疗患者的免疫功能提供实验依据。

Objective: To investigate the reverse effect and mechanism of IL-12 on chemotherapeutic medicine suppressing the immune function of NK cells. Methods: Purified NK cells were stimulated with PMA plus Ionomycin in the presence or absent of Cisplatin (DDP) and IL-12. The levels of IFN- γ and TNF- α in culture supernatants were detected by enzyme-linked immunosorbent assay (ELISA); The content of IFN-γ and TNF-α, TRAIL (TNF-related apoptosis inducing ligand) and transcription factors including T-bet and p-STAT-4 in NK cells were analyzed by Flow cytometry. The cytotoxicity of purified NK cells (pretreated with/without chemotherapeutics and IL-12 for 48 h) to Jurkat cells was measured by Flow cytometry. Results: Chemotherapeutics significantly inhibited the production of IFN-γ, TNF-α and the expression of TRAIL in NK cells, which were significantly reversed by IL-12 (P<0.05 or P<0.01). Further study revealed that chemotherapeutics down-regulated while IL-12 reversed the expression of p-STAT4 to restore cytokine production.In addition, DDP also inhibited but IL-12 recovered the cytotoxicity of NK cells against tumor cells by inducing the expression of TRAIL (P<0.05 or P<0.01). Conclusion: Chemotherapeutics inhibited the cytotoxicity of NK cells and its secretion of cytokines (IFN-γ and TNF-α), which were reversed by IL-12 via up-regulating TRAIL and p-STAT-4; this might provide experimental evidence for the clinical application of IL-12 for rebuild the immune function of tumor patients receiving chemotherapy.

国家自然科学基金资助项目（No.31470888)