花粉症患者发作期外周血Ⅱ型固有淋巴样细胞(ILC2)亚群的水平变化及其临床意义

Dynamic changes in peripheral typeⅡinnate lymphoid cell(ILC2)subpopulation and its clinical significance in children with hay fever during the pollen season

目的初步探讨花粉症患者在花粉季节发作期外周血Ⅱ型固有淋巴样细胞(ILC2)亚群表达水平、功能变化及其胞内STAT6信号通路水平变化。方法纳入10例花粉症患者、10例尘螨致敏哮喘患者以及12例健康儿童,分别在花粉季节发作期和非发作期采用流式细胞术监测其外周血ILC2细胞及其胞内Th2型因子水平变化。分选各组外周血Lin-细胞亚群,体外给予前炎症因子(IL-25和IL-33)刺激培养7 d,酶联免疫吸附法检测细胞培养上清中IL-5和IL-13的水平,Western blot法检测各组细胞总蛋白质中信号传导及转录激活因子6(STAT6)的磷酸化水平。结果在花粉季节发作期内,花粉症患者外周血ILC2细胞表达水平[(23.09±7.86)%]显著高于尘螨过敏哮喘组[(6.84±3.85)%]以及健康对照组[(1.69±0.87)%],差异均有统计学意义(P<0.05)。在非发作期,花粉症患者外周血ILC2细胞水平呈现下降趋势[(11.30±2.45)%],但依旧高于同期的尘螨致敏哮喘组[(3.76±1.96)%]以及健康对照组[(1.32±0.91)%],差异有统计学意义(P<0.05)。此外,在花粉季节发作期内,花粉症患者外周血IL-13+ILC2[(6.94±3.16)%vs(4.17±1.98)%,P<0.05]、尘螨致敏哮喘患者外周血IL-13+ILC2[(1.89±0.70)%vs(1.44±0.55)%,P<0.05]均显著高于非发作期。体外给予IL-25或IL-33刺激,均可以上调花粉症组和尘螨致敏哮喘组培养上清中IL-5和IL-13的水平,且当联合应用IL-25和IL-33可以协同放大此种刺激效应。Western blot证实应用IL-25和IL-33协同刺激可以显著上调花粉症患者组Lin-细胞胞内STAT6磷酸化水平。结论花粉季节发作期内,花粉症患者体内ILC2细胞数目及功能异常可能是导致花粉症患者在短时间内临床症状出现或急性加重的另一诱因。

Objective To analyze the dynamic changes in the expression and function of peripheral typeⅡinnate lymphoid cell(ILC2)subpopulation and the activity of signal transducers and activators of transcription(STAT6)in children with hay fever during pollen season.Methods A total of 10 patients with hay fever,10 patients with house dust mite(HDM)-sensitized asthma and 12 healthy controls(HC)were enrolled in this study.Changes in peripheral ILC2 and the intracellular expression of Th2-related cytokines were detected by flow cytometry during and outside the pollen season.Peripheral Lin-cell population was isolated from each group and cultured with the presence of IL-25 or IL-33 for 7 d.The concentrations of IL-5 and IL-13 in culture supernatants were measured by ELISA.Expression of phospho-STAT6 at protein level was quantified by Western blot.Results Within the pollen season,the percentage of peripheral ILC2 cells was significantly higher in children with hay fever[(23.09±7.86)%]than in children with HDM-sensitized asthma[(6.84±3.85)%,P<0.05]and healthy children[(1.69±0.87)%,P<0.05].In the non-pollen season,the peripheral ILC2 cells in children with hay fever presented a decreasing trend[(11.30±2.45)%],but was still higher than that in HDM-sensitized asthmatics[(3.76±1.96)%,P<0.05]and HC[(1.32±0.91)%,P<0.05]at the same time point.Moreover,peripheral IL-13+ILC2 cells in children with hay fever[(6.94±3.16)%vs(4.17±1.98)%,P<0.05]and in HDM-sensitized asthmatics[(1.89±0.70)%vs(1.44±0.55)%,P<0.05]during the pollen season were significantly higher than those in the non-pollen season.After the in vitro stimulation with IL25 or IL-33,the levels of IL-5 and IL-13 in culture supernatants were both increased in children with hay fever and HDM-sensitized asthmatics,and a synergistic action was observed when IL25 and IL-33 were used in combination.Meanwhile,the protein level of phospho-STAT4 in Lin-cells was significantly up-regulated in the hay fever group after stimulation with IL25 and IL-33.Conclusions During the pollen season,the abnormal number and function of ILC2 subpopulation in children with hay fever might be another cause of the occurrence of clinical symptoms in a short period of time or acute exacerbation.