[摘 要] 目的：观察白细胞介素-37（IL-37）在乳腺癌患者的表达变化对CD8+ T细胞活性的影响。方法：纳入2020年7月至 2022年9月在新乡医学院第一附属医院就诊的46例乳腺癌患者、24例乳腺良性肿瘤患者、20例对照者。采集外周血，分离血浆 和外周血单个核细胞（PBMC），收集接受手术治疗的乳腺癌患者肿瘤组织和癌旁组织，分离组织中肿瘤浸润淋巴细胞 （TIL），纯化CD8+ T细胞。ELISA法检测IL-37、可溶型单免疫球蛋白IL-1受体相关蛋白（SIGIRR）表达，实时定量PCR法检测组 织中IL-37 mRNA，流式细胞术检测CD8+ T细胞中IL-18受体α链（IL-18Rα）和SIGIRR表达。外源性IL-37刺激纯化的CD8+ T细 胞，与乳腺癌细胞系MCF-7共培养，通过测定乳酸脱氢酶水平计算靶细胞死亡比例，ELISA法检测上清中穿孔素、颗粒酶B、 干扰素-γ（IFN-γ）、肿瘤坏死因子-α（TNF-α）水平。结果：乳腺癌患者血浆IL-37水平高于乳腺良性肿瘤患者[（554.17 ± 96.63）pg/ mL vs （499.52 ± 78.66）pg/mL，P = 0.020]和对照者[（483.97 ± 47.23）pg/mL，P = 0.003]。乳腺癌患者肿瘤组织中IL-37 mRNA相对 表达量高于癌旁组织[（1.88 ± 0.21） vs （1.00 ± 0.53）pg/mL，P < 0.001]。外周血IL-18Rα+ CD8+细胞比例、SIGIRR+ CD8+细胞比例、 血浆可溶型SIGIRR水平在乳腺癌患者、乳腺良性肿瘤患者、对照者之间的差异无统计学意义（均P > 0.05）。CD8+ TIL表达IL 18Rα和SIGIRR的比例在肿瘤组织和癌旁组织之间的差异无统计学意义（P > 0.05）。重组人IL-37刺激后，CD8+ T细胞诱导靶细 胞死亡比例、上清中IFN-γ和TNF-α水平在直接接触和间接接触共培养系统中均低于无刺激（均P < 0.05）。在直接接触共 培养系统中，IL-37刺激后上清中穿孔素和颗粒酶B水平均低于无刺激（均P < 0.001），但在间接接触共培养系统中，上清中穿孔 素和颗粒酶B水平在无刺激和IL-37刺激之间的差异无统计学意义（均P > 0.05）。结论：乳腺癌患者中IL-37水平升高可能参与 诱导外周血和肿瘤微环境中CD8+ T细胞功能衰竭。

[Abstract] Objective: To investigate the effects of changes in interleukin-37 (IL-37) expression on CD8+ T cell vitality in patients with breast cancers. Methods: Forty-six patients with breast cancers, twenty-four patients with benign breast tumors, and twenty controls enrolled at the First Affiliated Hospital of Xinxiang Medical University between July 2020 and September 2022 were included in the study. Peripheral blood was collected. Plasma and peripheral blood mononuclear cells (PBMC) were isolated. Tumor tissues and para-cancerous tissues were collected from breast cancer patients who received surgical treatment. Tumor infiltrating lymphocytes (TILs) were isolated and CD8+ T cells purified. IL-37 and the expressions of soluble single immunoglobin IL-1 receptor-related (SIGIRR) proteins were detected by enzyme-linked immunosorbent assay (ELISA). IL-37 mRNA in the tissue was measured by real time PCR. IL-18 receptor α chain (IL-18Rα) and SIGIRR expression in CD8+ T cells were measured by flow cytometry. Purified CD8+ T cells were stimulated with exogenous IL-37, and were co-cultured with breast cancer cell line MCF-7. The percentage of target cell death was calculated by measuring the level of lactate dehydrogenase. Perforin, granzyme B, interferon-γ（IFN-γ）, and the level of tumor necrosis factor-α (TNF-α) in the supernatants was detected by ELISA. Results: Plasma IL-37 level was significantly elevated in patients with breast cancer compared with that in patients with benign breast tumors ([554.17 ± 96.63] vs [499.52 ± 78.66] pg/mL, P = 0.020) and that in the controls ([483.97 ± 47.23] pg/mL, P = 0.003). The expression of IL-37 mRNA in the tumor tissues was higher than that in the para-cancerous tissues in patients with breast cancer ([1.88 ± 0.21] vs [1.00 ± 0.53] pg/mL, P < 0.001). There were no statistically significant differences of peripheral IL-18Rα+CD8+ cell percentage, SIGIRR+ CD8+ cell percentage, or the level of soluble SIGIRR among patients with breast cancer, patients with benign breast tumors, and the controls (P > 0.05). There were also no statistically significant differences of the percentages of IL-18Rα+ or SIGIRR+ expression in CD8+ TIL cells between the tumor tissues and the para-cancerous tissues (P > 0.05). The percentage of CD8+ T cell-induced target cell death, and the levels of IFN-γ and TNF-α in the supernatants in both direct contact and indirect contact co-culture systems were reduced with IL-37 stimulation compared with those without IL-37 stimulation(P < 0.05). In direct contact co-culture system, the levels of perforin and granzyme B in the supernatants were down-regulated in response to IL-37 stimulation compared with those without stimulation (P < 0.001). However, in indirect contact co-culture system there were no significant differences in the levels of perforin or granzyme B in the supernatants with or without IL-37 stimulation (P > 0.05). Conclusion: Elevated IL-37 level might take part in the induction of CD8+ T cell exhaustion in peripheral blood and tumor microenvironments in breast cancer patients.

新乡市科技攻关项目（No. GG2020031）