CAJ | 학술논문

目的探讨重组人高迁移率族蛋白1(recombinant human HMGB1,rhHMGB1)对人T淋巴细胞白血病病毒1型(HTLV-1)感染的T细胞中与HMGB1相关膜受体表达的影响.方法病毒阳性细胞MT2中加入终浓度为1 μg/ml的人源化重组人rhHMGB1蛋白或等体积的PBS,培养24h后,以流式细胞术、免疫印迹、免疫荧光检测膜受体晚期糖基化终产物受体(receptor for advanced glycosylation endproducts,RAGE)、T细胞相关免疫球蛋白黏蛋白3(T cell immunoglobulin mucin 3,TIM-3)、Toll样受体4(Toll like receptor 4,TLR4)的表达及分布.结果流式细胞术检测结果显示1μg/ml rhHMGB1刺激MT2细胞后,与对照相比RAGE表达下降,TLR4的表达明显升高(P＜0.05);免疫印迹检测结果显示1 μg/ml rhHMGB1刺激MT2细胞后,细胞总蛋白和胞质中,膜受体RAGE和TLR4表达均明显升高(P＜0.05),而TIM-3的表达没有明显变化;免疫荧光检测结果显示rhHMGB1刺激MT2细胞后TLR4和RAGE的荧光强度均有增加,而TIM-3的荧光强度没有明显变化,且TLR4、TIM-3集中于细胞膜,而RAGE则集中于细胞质,细胞膜表达较少.结论胞外的HMGB1可促进HTLV-1感染T细胞RAGE和TLR4的表达.
목적 탐토중조인고천이솔족단백1(recombinant human HMGB1,rhHMGB1)대인T림파세포백혈병병독1형(HTLV-1)감염적T세포중여HMGB1상관막수체표체적영향.방법 병독양성세포MT2중가입종농도위1 μg/ml적인원화중조인rhHMGB1단백혹등체적적PBS,배양24h후,이류식세포술、면역인적、면역형광검측막수체만기당기화종산물수체(receptor for advanced glycosylation endproducts,RAGE)、T세포상관면역구단백점단백3(T cell immunoglobulin mucin 3,TIM-3)、Toll양수체4(Toll like receptor 4,TLR4)적표체급분포.결과 류식세포술검측결과현시1μg/ml rhHMGB1자격MT2세포후,여대조상비RAGE표체하강,TLR4적표체명현승고(P＜0.05);면역인적검측결과현시1 μg/ml rhHMGB1자격MT2세포후,세포총단백화포질중,막수체RAGE화TLR4표체균명현승고(P＜0.05),이TIM-3적표체몰유명현변화;면역형광검측결과현시rhHMGB1자격MT2세포후TLR4화RAGE적형광강도균유증가,이TIM-3적형광강도몰유명현변화,차TLR4、TIM-3집중우세포막,이RAGE칙집중우세포질,세포막표체교소.결론 포외적HMGB1가촉진HTLV-1감염T세포RAGE화TLR4적표체.
Objective To investigate the influences of recombinant human high mobility group protein 1 (rhHMGB1) on the expression of HMGB1-related receptors in Human T-cell leukemia virus type 1 (HTLV-1) infected MT2 T cells.Methods rhHMGB1 was added to HTLV-l-positive MT2 cells at a final concentration of 1 μg/ml.Equal volume of PBS was used to set up the control.After 24 h of culture,the expression and distribution of surface molecules including advanced glycosylation end products (RAGE),T cell immunoglobulin mucin 3 (TIM-3) and toll like receptor 4 (TLR4) were detected by flow cytometry,Western blot and immunofluorescence.Resulls The results of flow cytometry analysis showed a decreased expression of RAGE,but a significantly increased expression of TLR4 (P＜0.05) on MT2 cells stimtlated by 1μg/ml rhHMGB1,as compared with control group.The expression of RAGE and TLR4 among total cellular proteins and cytoplasmic proteins were significantly increased (P＜0.05),while the expression of TIM-3 showed no significant change as indicated by Western Blot.The immunofluorescence staining demonstrated that the fluorescence intensity of TLR4 and RAGE were increased,while the fluorescence intensity of TIM-3 showed no change upon rhHMGB1 treatment.Furthermore,TLR4 and TIM-3 were mainly distributed on the cell membrane,while RAGE was found in the cytoplasm.Conclusion Recombinant human HMGB1 can enhance the expression of RAGE and TLR4 in HTLV-1-infected T cell line MT2.