Objective: To investigate the levels of NK cells and their relevant cytokines (IL-10, TGF-β and IFN-γ) in patients with primary immune thrombocytopenia (ITP) . Methods: All samples were obtained from 42 patients (22 newly diagnosed and 20 in remission) and 20 healthy volunteers. The levels of IL-10 and IFN-γ in blood serum were detected by enzyme-linked immunosorbent assay (ELISA) . The percentage of CD3(-) CD56(+) NK cell, CD3(-) CD56(bright) CD16(-) NK cell, CD3(-) CD56(dim) CD16(+) NK cell in peripheral blood lymphocyte were detected by flow cytometry. The NK cells were isolated by immunomagnetic microbeads. The mRNA expression levels of IL-10, TGF-β, and IFN-γ in NK cells were detected by real-time fluorescent quantitative PCR. Correlation between the above measured results was analyzed. Results: ① The blood serum level of IFN-γ in newly diagnosed ITP patients [ (653.0±221.6) ng/L] was higher than that in remission ITP patients [ (484.4±219.5) ng/L] and healthy control [ (390.9±253.5) ng/L] (P=0.022, P=0.001) . The blood serum level of IL-10 in newly diagnosed ITP patients was lower than that in healthy control [ (52.09±26.66) ng/L vs (79.44±38.43) ng/L, P=0.007]. ②The percentage of NK cell in newly diagnosed and remission ITP patients [ (9.53±3.93) %, (9.03±3.78) %] were significantly lower than that in healthy control [ (13.72±7.42) %] (P=0.013, P=0.007) . The ratio of CD3(-) CD56(bright) CD16(-) NK cell/total NK cells in newly diagnosed ITP patients was higher than that in healthy control [ (6.85±4.43) % vs (4.05±2.81) %, P=0.032]. The ratio of CD3(-)CD56(dim) CD16(-) NK cell/total NK cells in newly diagnosed ITP patients was lower than that in healthy control [ (93.14±4.43) % vs (95.94±2.81) %, P=0.032]. ③ There was no significant difference in the mRNA expression level of IFN-γ in NK cells of ITP patients and healthy control (all P>0.05) . The mRNA expression levels of IL-10 and TGF-β in NK cells in newly diagnosed ITP patients were significantly higher than that in healthy control (1.82±1.32 vs 1.02±1.03, P=0.023; 2.80±2.31 vs 1.46±1.37, P=0.028) . The ratio of CD3(-)CD56(bright) CD16(-) NK cell/total NK cells was positively correlated with the mRNA expression levels of IL-10, TGF-β in NK cells (r=0.424, P=0.001; r=0.432, P<0.001) . Conclusion: NK cells may compensate for the deficiency of the number by enhancing the secretion of negative regulation cytokines, acting as "protective" roles in the disease.
目的： 探讨原发免疫性血小板减少症（ITP）患者外周血中NK细胞及其相关细胞因子IFN-γ、IL-10、TGF-β的变化。 方法： 以22例初诊ITP患者（初诊组）、20例治疗后完全缓解ITP患者（完全缓解组）为研究对象，以20名健康志愿者为对照组。采用ELISA法检测三组受试者血清IFN-γ及IL-10水平；用流式细胞术检测NK细胞（CD3(-)CD56(+)）及其Bright亚群（CD3(-)CD56(bright) CD16(-)）、Dim亚群（CD3(-)CD56(dim) CD16(+)）水平；采用免疫磁珠法分离NK细胞，实时荧光定量PCR检测IFN-γ、IL-10、TGF-β基因mRNA的表达，并将以上测得结果做相关性分析。 结果： ①初诊组ITP患者血清IFN-γ浓度[（653.0±221.6）ng/L]高于完全缓解组[（484.4±219.5）ng/L]和对照组[（390.9±253.5）ng/L]（P值分别为0.022、0.001），血清IL-10浓度低于对照组[（52.09±26.66）ng/L对（79.44±38.43）ng/L，P=0.007]。②初诊组、完全缓解组患者外周血NK细胞比例[（9.53±3.93）%、（9.03±3.78）%]均低于对照组[（13.72±7.42）%]（P=0.013，P=0.007）；初诊患者外周血Bright亚群占NK细胞的比例高于对照组[（6.85±4.43）%对（4.05±2.81）%，P=0.032]；初诊组外周血Dim亚群占NK细胞的比例低于对照组[（93.14±4.43）%对（95.94±2.81）%，P=0.032]。③初诊组、完全缓解组及对照组NK细胞IFN-γ基因mRNA表达差异无统计学意义（P>0.05），初诊组NK细胞IL-10、TGF-β基因mRNA表达高于对照组（1.82±1.32对1.02±1.03，P=0.023；2.80±2.31对1.46±1.37，P=0.028）。外周血Bright细胞占NK细胞的比例与NK细胞IL-10及TGF-β基因mRNA表达呈正相关（r=0.424，P=0.001；r=0.432，P<0.001）。 结论： NK细胞可能通过加强分泌免疫负调控因子来代偿其数量的不足，在疾病中起保护作用。.
Keywords: Interferon-gamma; Interleukin-10; Killer Cell, natural; Thrombocytopenia; Transforming growth factor beta.