| 熊利婷,李春晓,王俊杰.高剂量分割照射对NK细胞效应功能的影响及应用[J].中华放射医学与防护杂志,2025,45(7):623-628.Xiong Liting,Li Chunxiao,Wang Junjie.The impact and applications of high-dose fractionated irradiation on the effector function of NK cells[J].Chin J Radiol Med Prot,2025,45(7):623-628 |
| 高剂量分割照射对NK细胞效应功能的影响及应用 |
| The impact and applications of high-dose fractionated irradiation on the effector function of NK cells |
| 投稿时间：2025-03-18 |
| DOI：10.3760/cma.j.cn112271-20250318-00093 |
| 中文关键词: 高剂量分割照射 自然杀伤细胞 T细胞免疫球蛋白和ITIM结构域蛋白(TIGIT) |
| 英文关键词:High-dose fractionated irradiation Natural killer cells T-cell immunoreceptor with Ig and ITIM domains (TIGIT) |
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| 中文摘要: |
| 目的 探讨高剂量分割照射对自然杀伤(NK)细胞功能及免疫检查点表达的影响。方法 取10只6~8周龄C57BL/6N雌性小鼠,右侧肩背部皮下注射Lewis肺癌细胞,建立皮下荷瘤肺癌模型。采用随机数表法分为对照组和照射(RT)组,每组5只。RT组接受分割照射8 Gy×3次的照射,对照组不照射。照后3 d采用流式细胞术,比较两组间脾脏、引流淋巴结、肿瘤组织内NK细胞的数量、NK细胞免疫检查点程序性死亡受体1(PD-1)、具有Ig和ITIM结构域的T细胞免疫受体(TIGIT)以及NK细胞表面抗体溶酶体相关膜蛋白1(CD107a)的表达变化。用NK细胞表面抗体CD107a的表达评估其活化状态和细胞毒性。取20只荷瘤小鼠采用随机数表法分为对照组、单纯照射(RT)组、单纯抗TIGIT治疗(αTIGIT)组和照射联合抗TIGIT治疗组(RT+αTIGIT)4组,每组5只,RT组和RT+αTIGIT组接受高剂量分割照射,αTIGIT组和RT+αTIGIT组每3天给予αTIGIT抗体0.5 mg/kg,对照组用同型IgG处理,监测肿瘤大小。照后20 d,分别采用Ki67免疫组织化学染色和TUNEL免疫荧光染色检测肿瘤细胞增殖水平和凋亡事件。结果 与对照组相比,高剂量分割照射导致RT组的脾脏、肿瘤引流淋巴结及肿瘤微环境中的NK细胞数量减少(t=6.79~7.54,P<0.05),PD-1和TIGIT (t=4.16~5.65,P<0.05;t= 6.12~21.13,P<0.05)的平均荧光强度显著上调。CD107a+表达下降(t= 5.64~8.99,P<0.05)。与RT组相比,RT+αTIGIT组肿瘤质量和体积降低(t=2.50、9.12,P<0.05),且Ki67表达下降(t=12.10,P<0.05),TUNEL表达升高(t=5.42,P<0.05)。结论 高剂量分割照射会削弱NK细胞功能,上调免疫检查点表达,从而限制免疫应答,照射联合抗TIGIT抗体治疗能够在一定程度上逆转这一趋势,改善肿瘤治疗效果。 |
| 英文摘要: |
| Objective To investigate the impact of high-dose fractionated irradiation on NK cell function and immune checkpoint expression. Methods A total of 10 female C57BL/6N mice aged 6-8 weeks were subcutaneously injected with Lewis lung carcinoma (LLC) cells on the right shoulder to establish a subcutaneous tumor model. Mice were randomly divided into two groups: the control (Ctrl) group and the radiation (RT) group, with five mice in each group. The RT group received fractionated irradiation of 8 Gy×3 fractions, while the Ctrl group was not irradiated. Three days later, flow cytometry was performed to compare the numbers of NK cells and the expressions of immune checkpoints, including programmed death-1 (PD-1) and T-cell immunoreceptor with Ig and ITIM domains (TIGIT), in the spleen, draining lymph nodes, and tumor tissue between the two groups. The activation state and cytotoxicity of NK cells were assessed by the expression of lysosomal-associated membrane protein 1 (CD107a) on the NK cell surface. Additionally, a second experiment was conducted with 20 tumor-bearing mice divided into four groups: Ctrl, RT, anti-TIGIT (αTIGIT) treatment, and RT combined with αTIGIT treatment (RT+αTIGIT), with five mice per group. The RT and RT+αTIGIT groups received fractionated RT, while the αTIGIT and RT+αTIGIT groups were treated with blocking antibody αTIGIT at 0.5 mg/kg every 3 days and the Ctrl group was treated with isotype control IgG. Tumor size was monitored during the experiment. On day 20, mice were euthanized, Ki67 immunohistochemistry and TUNEL immunofluorescence staining were used to evaluate tumor cell proliferation and apoptosis. Results Compared with the Ctrl group, fractionated high-dose RT resulted in a decrease in NK cell numbers in the spleen, tumor-draining lymph nodes, and tumor microenvironment in the RT group (t=6.79-7.54, P<0.05), together with a significant upregulation of the mean fluorescence intensity of PD-1 and TIGIT (t=4.16-5.65, P<0.05; t=6.12-21.13, P<0.05), and a decrease in CD107a+ expression (t=5.64-8.99, P<0.05). Compared with the RT group, the RT+αTIGIT group showed reduced tumor weight (t=2.50, 9.12, P<0.05), decreased Ki67 expression (t=12.10, P<0.05), and increased TUNEL expression (t=5.42, P<0.05). Conclusions The fractioned high-dose RT can impair NK cell function and upregulate immune checkpoint expression, thereby limiting the immune response. The combination of irradiation with anti-TIGIT antibody treatment can partially reverse this trend and improve tumor treatment efficacy. |
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