低剂量辐射对恶性肿瘤患者自体CIK细胞免疫功能影响的实验研究

目的 观察低剂量辐射对恶性肿瘤患者自体CIK细胞增殖、表型和杀伤活性的影响,为临床应用CIK细胞过继免疫治疗提供依据。方法 取10例恶性肿瘤患者外周血,常规分离出单个核细胞,用不同细胞因子培养诱导CIK细胞。培养10 d后,以30、50、80、100和200 mGy X射线照射,24 h后采用³H-TdR掺入法检测对CIK细胞增殖的影响;流式细胞术检测对CD3+CD56+表型CIK细胞百分率的影响;³H-TdR释放法检测对其杀伤活性的影响。采用³H-TdR释放法检测80 mGy X射线照射对自体CIK细胞在12、24、48和72 h杀伤活性的影响,及80 mGy连续照射3 d对CIK细胞杀伤活性的影响。结果 CIK细胞增殖活性与对照组(0 mGy)相比均有明显增高(t =2.2、3.5、3.3和2.2, P ＜0.05)。50、80和100 mGy组的CD3+CD56+细胞的百分率均有显著性增高(t =2.3、4.2和2.4, P ＜0.05)。CIK细胞接受LDI,80 mGy组和100 mGy组与对照组(0 mGy)相比,CIK细胞杀伤活性均有显著性增高(t =3.3和2.3, P ＜0.05)。80 mGy照射后24 h,CIK细胞的杀伤活性出现高峰,为(54.2±5.0)%(t =3.2, P ＜0.01),48和72 h下降到正常水平。80 mGy连续照射3 d,24和48 h CIK细胞杀伤活性分别为(55.2±5.3)%和(61.9±4.4)%,72 h达到最高水平为(67.2±5.7)%(t =2.6、4.7和5.7, P ＜0.05)。结论 低剂量辐射对肿瘤患者CIK细胞显示兴奋效应,具有临床应用前景。

Experimental study on effect of low dose radiation on autologous CIK cell immunity functions of tumor patients

Objective To observe the effects of low dose irradiation (LDI) on autoiogous CIK cell proliferation, phenotype and killing activity in tumor patients, and to provide the evidence for clinical application of adoptive immunotherapy with CIK cells. Methods Peripheral blood mononuclear cells (PBMC) were separated from 10 patients with malignant tumor, and CIK cells were cultured with different cytokines. (1) After 10 d culture, C1K cells were irradiated with different doses as 30, 50, 80, 100 and 200 Gy of X-rays was also detected. The CIK cell proliferation and killing activity were measured with 3H-TdR incorporation assay and 3H-TdR release assay, respectively and the percentage variation of CD3 +CD56 + were measured with flow cytometry after 24 h. ( 1 ) Autologous CIK cells were irradiated with 80 mGy X-rays. At different culture time ( 12, 24, 48, 72 h) after irradiation, the killing activity was measured with 3H-TdR release assay. (3) The effect of 3d low dose irradiation of 80 mGy X-rays on thekilling activity of CIK cells was also detected. Results After the CIK cells were irradiated with different doses as 50, 80, 100, 200 mGy of X-rays, the CPM values were 20 048.6 ± 2332. 2 ( t = 2.2, P ＜0.05), 21 832.2 ±2975.9 (t=3.5, P＜0.01), 21 000.3 ±2451.1 (t=3.3, P＜0.01), 19908.1 ±2051.0 ( t = 2.2, P ＜ 0.05 ), respectively and the proliferation of CIK cells were significantly higher than that of control group. The CD3 + CD56 + cell percentage of 50, 80, 100 mGy groups were ( 30.3 ±3.8)% (t=2.3, P＜0.05), (32.3±3.4)% (t=4.2, P＜0.01), (29.742.9)% (t = 2.4, P＜0.05 ), respectively. The killing activity of CIK cells of 80, 100 mGy groups were 55.2 ± 5.0 ( t = 3.3, P ＜ 0.01 ), 52.8 ± 4.1 ( t = 2.3, P ＜ 0.05 ), respectively. The killing activity of CIK cells up-regulated significantly at 24 h, dropped to normal levels at 48 h and 72 h. After 80 mGy X-ray irradiation for 3 consecutive days, the killing activity of CIK cells at different time points were 55.2 ± 5.3 (t = 2.6, P ＜0.05),61.9 ± 4.4 (t = 4.7, P ＜0.01), 67.2 ±5.7 (t = 5.7, P ＜0.01) for 24, 48, 72 h,respectively. Conclusions LDI might have the hormesis effect on CIK cells.