舌下含服免疫治疗对效应性T细胞和调节性T细胞功能的影响

Effect of sublingual immunotherapy on functions of effector T cells and regulatory T cells

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.