CAJ | 학술논문

目的探讨谷氨酸信号通路在人外周血淋巴细胞活化及白癜风免疫发病机制中的作用.方法流式细胞仪测定离子型谷氨酸受体NMDAR的非竞争性拮抗剂MK801对健康对照外周血淋巴细胞CD25表达及IFN-γ水平的影响；NMDAR的激动剂NMDA和拮抗剂MK801对淋巴细胞内活性氧簇水平的影响.实时定量PCR和流式细胞仪测定白癜风患者和健康对照外周血淋巴细胞中NMDAR亚型NMDAR1和NMDAR2A表达差异.免疫组化方法观察白癜风患者皮损和健康对照皮肤组织中NMDAR1和NMDAR2A的表达.结果 MK801组外周血淋巴细胞CD25表达(7.28±0.18)％较阴性对照组(16.02±0.42)％明显下降(P＜ 0.01)；MK801组淋巴细胞中CD25+ IFN-γ+比例(1.79±0.09)％较阴性对照组(0.78±0.06)％明显增高(P＜0.01).NMDA组淋巴细胞内ROS水平(101.1±3.50)明显高于阴性对照组(69.80±2.08)(P＜0.01).白癜风患者外周血淋巴细胞NMDAR1的表达(3.85±2.17)较健康组(0.97±0.55)明显增高(P＜0.05).结论谷氨酸信号通路对活化淋巴细胞的IFN-γ分泌及ROS水平的影响有可能是白癜风的免疫发病机制之一.
목적 탐토곡안산신호통로재인외주혈림파세포활화급백전풍면역발병궤제중적작용.방법 류식세포의측정리자형곡안산수체NMDAR적비경쟁성길항제MK801대건강대조외주혈림파세포CD25표체급IFN-γ수평적영향；NMDAR적격동제NMDA화길항제MK801대림파세포내활성양족수평적영향.실시정량PCR화류식세포의측정백전풍환자화건강대조외주혈림파세포중NMDAR아형NMDAR1화NMDAR2A표체차이.면역조화방법관찰백전풍환자피손화건강대조피부조직중NMDAR1화NMDAR2A적표체.결과 MK801조외주혈림파세포CD25표체(7.28±0.18)％교음성대조조(16.02±0.42)％명현하강(P＜ 0.01)；MK801조림파세포중CD25+ IFN-γ+비례(1.79±0.09)％교음성대조조(0.78±0.06)％명현증고(P＜0.01).NMDA조림파세포내ROS수평(101.1±3.50)명현고우음성대조조(69.80±2.08)(P＜0.01).백전풍환자외주혈림파세포NMDAR1적표체(3.85±2.17)교건강조(0.97±0.55)명현증고(P＜0.05).결론 곡안산신호통로대활화림파세포적IFN-γ분비급ROS수평적영향유가능시백전풍적면역발병궤제지일.
Objective To investigate the roles of glutamate signaling pathway in the activation of human peripheral blood lymphocytes(PBLs) and pathogenesis of vitiligo.Methods Peripheral blood lymphocytes (PBLs) isolated from 5 patients with generalized vitiligo and 5 healthy controls were cultured in vitro.Flow cytometry was performed to quantify the expression of CD25 and interferon-γ on PBLs derived from healthy controls and treated with MK801 (a non-competitive antagonist of N-methyl-D-aspartic acid receptor,NMDAR) at 100 μmol/L or phosphate buffered saline (PBS) for 48 hours,as well as the level of reactive oxygen species (ROS) in the controlderived PBLs treated with MK801 at 100 μmol/L,NMDA (an agonist of N-methyl-D-aspartic acid receptor) at 0.5 mmol/L or PBS for 48 hours.The protein and mRNA expressions of NMDAR1 and NMDAR2A were measured by flow cytometry and real-time PCR,respectively,in PBLs from the healthy controls and vitiligo patients.Immunohistochemistry was used to observe the expressions of NMDAR1 and NMDAR2A in tissue specimens from depigmented and postinflammatory hyperpigmentation lesions of the patients with vitiligo and from normal skin of the healthy controls.Results Compared with the PBS-treated PBLs from the healthy controls,the MK801-treated PBLs showed a downregulated expression of CD25 (7.28％ ± 0.18％ vs.16.02％ ± 0.42％,P ＜ 0.01),but an upregulated proportion of CD25+IFN-γ+ lymphocytes (1.79％ ± 0.09％ vs.0.78％ ± 0.06％,P ＜ 0.01),and the NMDA-treated PBLs displayed a higher ROS level (101.1 ± 3.50 vs.69.80 ± 2.08,P＜ 0.01 ).The protein expression of NMDAR1 in PBLs was significantly higher in vitiligo patients than in the healthy controls (3.85 ± 2.17 vs.0.97 ±0.55,P ＜ 0.05).Conclusion Glutamate signaling pathway may be involved in the immunopathogenesis of vitiligo via affecting the secretion of interferon-γ by,and ROS level in,activated lymphocytes.