调节性T细胞在阵发性睡眠性血红蛋白尿症异常克隆中的免疫机制研究

doi:10.3760/cma.j.issn.1007-1245.2022.22.006

国际医药卫生导报 ›› 2022, Vol. 28 ›› Issue (22): 3135-3139.DOI: 10.3760/cma.j.issn.1007-1245.2022.22.006

调节性T细胞在阵发性睡眠性血红蛋白尿症异常克隆中的免疫机制研究

林莺张荣东陈琦陈仁利

宁德师范学院附属宁德市医院血液科，宁德　352100

收稿日期:2022-08-30 出版日期:2022-11-15 发布日期:2022-11-18
通讯作者: 林莺，Email：linying806@126.com
基金资助:
福建省自然科学基金面上项目（2020J011346）；2019年宁德市指导性科技计划项目（20190051）

Study of immune mechanism of regulatory T cells in abnormal clones of paroxysmal nocturnal hemoglobinuria

Lin Ying, Zhang Rongdong, Chen Qi, Chen Renli

Department of Hematology, Ningde Municipal Hospital of Ningde Normal University, Ningde 352100, China

Received:2022-08-30 Online:2022-11-15 Published:2022-11-18
Contact: Lin Ying, Email: linying806@126.com
Supported by:
General Project of Fujian Natural Science Foundation (2020J011346); Ningde City Guiding Science and Technology Plan Project in 2019 (20190051)

摘要/Abstract

摘要： 目的　测定阵发性睡眠性血红蛋白尿症（PNH）患者调节性T细胞的变化，探讨其在PNH异常克隆中可能的免疫机制。方法　2018年1月至2022年1月在宁德师范学院附属宁德市医院血液科就诊的31例PNH患者为病例组，其中发作性PNH 15例（男10例、女5例，年龄23～69岁）、再生障碍性贫血-阵发性睡眠性血红蛋白尿（AA-PNH）16例（男11例、女5例，年龄20～68岁）；25例健康体检者为对照组（男14例、女11例，年龄19～56岁）。用流式细胞仪检测各组外周血T细胞和Th细胞亚群变化以及CD4⁺CD25⁺T细胞和Th3细胞数量，用实时荧光定量多聚核苷酸链式反应（RT-PCR）测定转化生长因子-β（TGF-β）和叉头框蛋白P3（FOXP3）的表达水平，用酶联免疫吸附试验（ELISA）检测TGF-β和白细胞介素-10（IL-10）浓度。采用F检验、Bonferroni方法、H检验、χ²检验。结果　AA-PNH组CD4⁺/CD3⁺细胞和CD4⁺/CD8⁺比例低于对照组［（44.55±11.91）%比（57.61±6.90）%、0.66（0.38，1.26）比1.32（0.86，1.64）］，CD8⁺/CD3⁺细胞高于对照组［（62.31±10.76）%比（45.51±5.54）%］，差异均有统计学意义（均P<0.05）；AA-PNH组Th1/CD3⁺CD4⁺细胞高于发作性PNH组及对照组［（24.27±4.30）%比（12.61±3.02）%、（4.07±1.93）%］，差异有统计学意义（P<0.05）。AA-PNH组及发作性PNH组CD4⁺CD25⁺T细胞和Th3细胞的数量高于对照组［（67.82±15.67）×10⁶/L、（97.11±17.10）×10⁶/L比（43.86±9.52）×10⁶/L，（57.15±10.58）×10⁶/L、（82.27±15.15）×10⁶/L比（42.35±9.33）×10⁶/L］，差异均有统计学意义（均P<0.05）。发作性PNH组、AA-PNH组中FOXP3及TGF-β的mRNA的阳性表达率和TGF-β及IL-10浓度均高于对照组［86.67%（13/15）、75.00%（12/16）比40.00%（10/25），73.33%（11/15）、62.50%（10/16）比36.00%（9/25），（49.76±2.03）μg/L、（37.29±1.78）μg/L比（17.01±3.27）μg/L，（7.86±1.17）pg/ml、（5.57±1.13）pg/ml比（1.33±0.49）pg/ml］，差异均有统计学意义（均P<0.05）。结论　PNH患者调节性T细胞数量增加，功能亢进，推测其免疫抑制作用可能使其在PNH异常克隆中逃避免疫监视。

关键词: 阵发性睡眠性血红蛋白尿症, 调节性T细胞, Th3细胞, 转化生长因子-β, 叉头框蛋白P3

Abstract: Objective　To determine the change of regulatory T cells in paroxysmal nocturnal hemoglobinuria (PNH) patients and analyze its probable immune mechanism in abnormal clones of PNH. Methods　Thirty-one PNH patients admitted to Department of Hematology, Ningde Municipal Hospital of Ningde Normal University from January 2018 to January 2022 were selected as a case group, including fifteen cases of episodic PNH (10 males and 5 females, aged 23-69 years) and sixteen cases of aplastic anemia-paroxysmal nocturnal hemoglobinuria (AA-PNH) (11 males and 5 females, aged 20-68 years). Twenty-five healthy subjects were selected as a control group (14 males and 11 females, aged 19-56 years). Whole blood samples in subgroups were assayed by flow cytometer to detect T cells, Th lymphocyte subsets, and the numbers of CD4⁺CD25⁺ regulatory T cells and Th3 cells. The expression levels of transforming growth factor-β (TGF-β) and forkhead box P3 (FOXP3) were analyzed by real-time quantitative polymerase chain reaction (RT-PCR). Enzyme-linked immunosorbent assay (ELISA) kits were used to detect the plasma concentrations of TGF-β and interleukin-10 (IL-10). F test, Bonferroni method, H test, and chi-square test were used. Results　The number of CD4⁺/CD3⁺ cells and the CD4⁺/CD8⁺ ratio in the AA-PNH group were lower than those in the control group [(44.55±11.91)% vs. (57.61±6.90)%, 0.66 (0.38, 1.26) vs. 1.32 (0.86, 1.64)], but the CD8⁺/CD3⁺ cells were higher than those in the control group [(62.31±10.76)% vs. (45.51±5.54)%], with statistically significant differences (all P<0.05). The number of Th1/CD3⁺CD4⁺ cells in the AA-PNH group was significantly higher than those in the episodic PNH group and the control group [(24.27±4.30)% vs. (12.61±3.02)%, (4.07±1.93)%], with a statistically significant difference (P<0.05). The numbers of CD4⁺CD25⁺ T cells and Th3 cells in the AA-PNH group and the episodic PNH group were higher than those in the control group [(67.82±15.67) ×10⁶/L, (97.11±17.10) ×10⁶/L vs. (43.86±9.52) ×10⁶/L; (57.15±10.58) ×10⁶/L, (82.27±15.15) ×10⁶/L vs. (42.35±9.33) ×10⁶/L], with statistically significant differences (all P<0.05). The positive expression rates of FOXP3 and TGF-β mRNA and the concentrations of TGF-β and IL-10 in the episodic PNH group and the AA-PNH group were higher than those in the control group [86.67% (13/15), 75.00% (12/16) vs. 40.00% (10/25); 73.33% (11/15), 62.50% (10/16) vs. 36.00% (9/25); (49.76±2.03) μg/L, (37.29±1.78) μg/L vs. (17.01±3.27) μg/L; (7.86±1.17) pg/ml, (5.57±1.13) pg/ml vs. (1.33±0.49) pg/ml], with statistically significant differences (all P<0.05). Conclusion　The number and function of regulatory T cells in PNH patients are increased and its inhibitive function might contribute to abnormal clones of PNH to evade the immune surveillance.

Key words: Paroxysmal nocturnal hemoglobinuria, Regulatory T lymphocytes, Th3 cells, Transforming growth factor-β, Forkhead box P3

林莺张荣东陈琦陈仁利. 调节性T细胞在阵发性睡眠性血红蛋白尿症异常克隆中的免疫机制研究[J]. 国际医药卫生导报, 2022, 28(22): 3135-3139.

Lin Ying, Zhang Rongdong, Chen Qi, Chen Renli. Study of immune mechanism of regulatory T cells in abnormal clones of paroxysmal nocturnal hemoglobinuria[J]. International Medicine and Health Guidance News, 2022, 28(22): 3135-3139.

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调节性T细胞在阵发性睡眠性血红蛋白尿症异常克隆中的免疫机制研究

Study of immune mechanism of regulatory T cells in abnormal clones of paroxysmal nocturnal hemoglobinuria

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