目的·研究舌下含服免疫治疗(sublingual immunotherapy,SLIT)对变应性鼻炎(allergic rhinitis,AR)炎症反应中效应性T细胞(effector T cell,Teff)和调节性T细胞(regulatory T cell,Treg)功能的影响。方法·选择复旦大学附属华山医院北院耳鼻咽喉头颈外科招募的20例AR患者,分别在治疗前及治疗24个月和30个月时获取其外周血单个核细胞(peripheral blood mononuclear cell,PBMC),检测PBMC中Teff和Treg所占CD4+ T细胞百分比,并检测PBMC中2种细胞钙释放激活钙通道调节分子1(calcium release-activated calcium channel modulator 1,ORAI1)蛋白的浓度和Ca2+平均荧光强度(mean fluorescence intensity,MFI)。然后将治疗30个月时的2种细胞进行体外培养,检测2种细胞中ORAI1蛋白的表达以及培养基中白细胞介素-4(interleukin-4,IL-4)和IL-10浓度。制备含人ORAI1短发夹RNA的慢病毒并将其转染至2种细胞,再次检测2种细胞中Ca2+ MFI和培养基中IL-4和IL-10的浓度。结果·在治疗24个月和30个月时PBMC中Teff所占CD4+ T细胞百分比逐渐减少(P=0.000,P=0.027),而Treg所占CD4+ T细胞百分比逐渐增多(P=0.000,P=0.008);Teff中ORAI1蛋白表达量(P=0.000,P=0.003)和Ca2+ MFI(P=0.000,P=0.000)也逐渐减少,而Treg中ORAI1蛋白表达量(P=0.000,P=0.007)和Ca2+ MFI(P=0.000,P=0.000)随着治疗时间延长逐渐增多。体外培养的2种细胞均表达ORAI1蛋白,而且在慢病毒转染后,细胞中Ca2+ MFI降低(P=0.004,P=0.000),培养基中IL-4(P=0.009)和IL-10(P=0.000)的浓度均显著降低。结论·SLIT可以通过调节ORAI1蛋白的表达控制Teff和Treg细胞的功能。
Objective · To investigate the effect of sublingual immunotherapy (SLIT) on the functions of effector T cells (Teff) and regulatory T cells (Treg) under allergic rhinitis (AR) condition. Methods · Peripheral blood mononuclear cells (PBMC) were obtained from 20 AR patients who were recruited from Department of Otorhinolaryngology-Head and Neck Surgery, Huashan Hospital North of Fudan University before and after the 24- and 30-month treatments of SLIT. The percentages of Teff and Treg in CD4+ T cells in PBMC were examined and the contents of calcium release-activated calcium channel modulator 1 (ORAI1) protein and Ca2+ mean fluorescence intensity (MFI) in PBMC were measured. Then, Teff and Treg after the 30-month treatment were cultured in vitro, and were evaluated for the expression of ORAI1 protein and concentrations of interleukin-4 (IL-4) and IL-10 in the cultures. Lentivirus vector (lenti) that encoded short hairpin RNA against ORAI1 (lenti-ORAI1) was prepared and transfected into Teff and Treg, and then Ca2+ MFI in Teff and Treg and concentrations of IL-4 and IL-10 in the cultures were assessed again. Results · The percentage of Teff in CD4+ T cells in PBMC reduced gradually after the 24- and 30-month treatments (P=0.000, P=0.027); however, the percentage of Treg was increased (P=0.000, P=0.008). The expression of ORAI1 protein (P=0.000, P=0.003) and Ca2+ MFI (P=0.000, P=0.000) in Teff were also decreased gradually after the 24- and 30-month treatments; however, the expression of ORAI1 protein (P=0.000, P=0.007) and Ca2+ MFI in Treg were enhanced (P=0.000, P=0.000). ORAI1 protein was expressed in these two types of cells cultured in vitro. Furthermore, Ca2+ MFI in them (P=0.004, P=0.000) and IL-4 (P=0.009) and IL-10 (P=0.000) in the cultures were all decreased after the transfection of lenti-ORAI1. Conclusion · SLIT can control functions of Teff and Treg through regulating the expression of ORAI1 protein.