脂多糖对结核病患者外周血单核细胞Toll样受体4表达及细胞内活性氧产生的影响

摘要

Objective:To explore the effects of lipopolysaccharide ( LPS ) on the expression of Toll-like receptor 4 ( TLR4 ) and production of intracellular reactive oxygen species ( ROS ) in peripheral blood mononuclear cells of tuberculosis ( TB ) patients. Methods:The peripheral blood samples from 13 TB patients and 13 health volunteers were collected,which was dyed with ROS probe DHR123 or anti-TLR4-PE. The expression of TLR4 and production of intracellular ROS in peripheral blood mononuclear cells of TB patients and health volunteers were detected using flow cytometry. The 100 μL anticoagulant blood were cultured in 96-well plates,and divided into group A,group B,group C(6 wells each group) and cultured for 24 h,and treated with 100 ng/mL LPS. The expression of TLR4 and production of intracellular ROS in peripheral blood mononuclear cells were detected by flow cytometry. Results:The positive expression rate of TLR4 in peripheral blood mononuclear cells in TB patients[(17. 6 ± 3. 6)%] was significantly lower than that in health volunteer[(30. 0 ± 2. 4)%](P<0. 01). The level of ROS in peripheral blood mononuclear cells in TB patients(74. 8 ± 11. 4) was significantly lower than that in health volunteer(109. 6 ± 13. 2)(P<0. 01). After LPS stimulating for 24 h,the difference of the level of intracellular ROS between group C(182. 1 ± 11. 3) and group B(127. 6 ± 14. 1) was statistically significant(P<0. 01),and the difference of the expression level of intrcellular TLR4 between group C[(62. 1 ± 3. 3)%] and group B[(47. 6 ± 4. 7)%] was statistically significant(P <0. 01). Conclusions:LPS can increase the TLR4 expression and intracellular ROS production of peripheral blood mononuclear cells in TB patients.%目的:探讨脂多糖(LPS)对结核病患者外周血单核细胞Toll样受体4(TLR4)表达及细胞内活性氧(ROS)产生的作用.方法:收集健康志愿者外周血标本和结核病患者外周血标本,各13例;取二者肝素抗凝血100μL,荧光探针DHR123或抗人anti-TLR4-PE荧光素标记抗体,全血染色后,流式细胞仪技术检测分析二者外周血单核细胞TLR4表达及细胞内ROS产生水平;再取二者抗凝血100μL于96孔培养板中,按如下分组进行处理:健康志愿者抗凝血为A组,结核病患者抗凝血为B组,结核病患者抗凝血+LPS 100 ng/mL为C组,每组6孔,培养24 h后,流式细胞仪技术检测单核细胞TLR4表达及细胞内ROS产生水平.结果:结核病患者外周血单核细胞TLR4阳性表达率为(17.6±3.6)%,较健康志愿者的(30.0±2.4)%显著降低(P<0.01);结核病患者外周血单核细胞细胞内ROS水平(74.8±11.4)较健康志愿者(109.6±13.2)显著降低(P<0.01).LPS刺激24 h后,C组单核细胞胞内ROS水平(182.1±11.3)与B组(127.6±14.1)相比差异有统计学意义(P<0.01);C组单核细胞TLR4阳性表达率为(62.1±3.3)%,与B组的(47.6±4.7)%差异有统计学意义(P<0.01).结论:LPS可促进结核病患者外周血单核细胞TLR4表达及细胞内ROS产生水平.