Objective: To investigate natural killer (NK) cell quantities and function in patients with immune thrombocytopenia (ITP) . Methods: A total of 66 ITP patients (34 newly diagnosed and 32 in complete remission) were collected from September 2015 to May 2016 in Tianjin Medical University General Hospital, and 30 healthy volunteers were recruited as controls. The percentages of NK cells and their subsets in peripheral blood, the expression of activating receptor (NKp44), inhibitory receptor (NKG2A) and CD16, perforin and granzyme β were detected by flow cytometry. The correlation between the above parameters and patients' immune status and platelet level were evaluated. Results: (1)The percentage of CD3(-)CD56(+) NK cells in newly diagnosed patients (10.99%±4.89%)and patients in complete remission (9.73%±6.75%) were significantly lower than that in healthy controls (14.67%±7.24%)(P=0.023, 0.003). The percentage of NK cells Bright subset was significantly lower in the newly diagnosed patients(0.48%±0.23%)and those in complete remission (0.41%±0.33%) than in healthy controls(0.64%±0.32%)(P=0.037, 0.002); the percentage of Dim subset was also significantly lower in the newly diagnosed (10.16%±5.02%) and patients in complete remission (8.07%±5.74%) than in healthy controls(14.16%±7.19%) (P=0.009, 0.007). (2)The proportion of Bright subset in total NK cells in new diagnosed ITP patients (6.48%±4.33%) was significantly higher than that in healthy controls (4.21%±2.70%)(P=0.020); the proportion of Dim NK cells subset in new diagnosed ITP patients (93.51%±4.33%) was significantly lower than that in healthy controls(95.79%±2.70%) (P=0.020). (3)The expression of activating receptor NKp44 in new diagnosed ITP patients was significantly lower than that in complete remission group and healthy controls[0.28%(0.95%)vs 0.61%(2.05%), 0.92%(0.90%); P=0.047, 0.048]; the expression of inhibitory receptor NKG2A in new diagnosed ITP patients was significantly higher than that in healthy controls(42.34%±23.86% vs 29.25%±12.83%, P=0.009). The proportion of CD16 was significantly lower in the newly diagnosed patients than in healthy controls(93.51%±4.33%95.79%±2.70%, P=0.020). (4)The expression of perforin in the newly diagnosed ITP patients was significantly lower than that in healthy controls [87.52%(25.29%)vs 91.55%(8.29%), P=0.025]; the expression of granzyme β in ITP patients and controls showed no statistically significant difference. (5)The level of NK cells in ITP patients was negatively correlated with CD3(+) CD8(+) T cells (r=-0.387, P=0.012) and CD5(+) CD19(+) B cells in peripheral blood (r=-0.273, P=0.028), positively correlated with the ratio of CD3(+) CD4(+) /CD3(+) CD8(+) (r=0.358, P=0.028) and peripheral platelet count (r=0.314, P=0.011). Conclusion: Deceased quantities and impaired total NK function, insufficient suppression of autoreactive T and B cells might play a role in the pathogenesis of ITP.
目的: 探讨免疫性血小板减少症(ITP)患者外周血中自然杀伤(NK)细胞数量及功能的变化。 方法: 选取2015年9月至2016年5月天津医科大学总医院血液科住院的初诊ITP患者34例,完全缓解ITP患者32例及健康志愿者30名。采用流式细胞术检测外周血NK细胞及其亚群比例,NK细胞表面分子激活性受体NKp44、抑制性受体NKG2A及CD16,胞质蛋白穿孔素及颗粒酶β的表达水平,并将其与ITP患者的免疫状态及血小板水平做相关性分析。 结果: (1)NK细胞及其亚群占外周血淋巴细胞比例:NK细胞:初诊组(10.99%±4.89%)及完全缓解组(9.73%±6.75%)低于对照组(14.67%±7.24%)(P=0.023、0.003);Bright亚群:初诊组(0.48%±0.23%)及完全缓解组(0.41%±0.33%)低于对照组(0.64%±0.32%)(P=0.037、0.002);Dim亚群:初诊组(10.16%±5.02%)及完全缓解组(8.07%±5.74%)低于对照组(14.16%±7.19%)(P=0.009、0.007)。(2)各亚群占NK细胞的比例:Bright亚群:初诊组(6.48%±4.33%)高于对照组(4.21%±2.70%)(P=0.020);Dim亚群:初诊组(93.51%±4.33%)低于对照组(95.79%±2.70%)(P=0.020)。(3)激活性受体NKp44:初诊组低于完全缓解组及对照组[0.28%(0.95%)比0.61%(2.05%)、0.92%(0.90%),P=0.047、0.048];抑制性受体NKG2A:初诊组高于对照组(42.34%±23.86%比29.25%±12.83%,P=0.009);CD16:初诊组低于对照组(93.51%±4.33%比95.79%±2.70%,P=0.020)。(4)穿孔素:初诊组低于对照组[87.52%(25.29%)比91.55%(8.29%),P=0.025];颗粒酶β 3组之间差异无统计学意义。(5)NK细胞与外周血杀伤T细胞(CD3(+)CD8(+))、B1细胞(CD5(+)CD19(+))呈负相关(r=-0.387、-0.273,P=0.012、0.028),与外周血辅助/杀伤T细胞比值(CD3(+)CD4(+)/CD3(+)CD8(+))、血小板计数呈正相关(r=0.358、0.314,P=0.028、0.011)。 结论: ITP患者外周血NK细胞数量下降,发育障碍、功能降低,对自身反应性T、B细胞的抑制不足导致疾病的发生。.
Keywords: Granzymes; Killer cells, natural; Perforin; Receptors, natural killer cell; Thrombocytopenia.