肥胖症患者血清对人源单核细胞细胞上Toll样受体4/转录因子-κB信号通路的影响

Serum of obesity induce the activation of TLR4/NF-κB signaling pathway on THP-1 cell line

目的 探讨肥胖症及伴相关代谢紊乱患者血清对人源单核细胞（THP-1）细胞表面Toll样受体4/转录因子-κB（TLR4/NF-κB）信号通路的活化作用.方法 分别用10例单纯性肥胖及伴不同代谢紊乱患者血清干预THP-1细胞株24、48 h后,测定THP-1细胞表面TLR4和细胞内NF-κB p65磷酸化的表达水平及其分泌白细胞介素（IL-1）β和肿瘤坏死因子（TNF）-α的能力.Western印迹检测TLR4蛋白和细胞内NF-κB p65磷酸化蛋白水平;RT-PCR检测TLR4 mRNA的表达水平;ELISA检测细胞上清液中IL-1 β和TNF-α的水平.结果 THP-1细胞表面TLR4、细胞内NF-κB p65磷酸化的蛋白及TLR4 mRNA表达水平以及分泌IL-1β和TNF-α的能力差异有统计学意义（P＜0.05）;THP-1细胞表面TLR4、细胞内NF-κB p65磷酸化蛋白及TLR4 mRNA表达水平以及分泌IL-1β和TNF-α的能力随肥胖症代谢紊乱组分的增加而增高,并随干预时间的延长而增高（P＜0.05）.正常对照组、单纯性肥胖组、肥胖伴高血糖组、肥胖伴血脂紊乱组以及肥胖伴3种代谢紊乱组血清孵育THP-1细胞48 h,TNF-α的浓度（ng/L）分别为（222±32）、（246±52）、（322±32）、（322±34）和（490±83）,IL-1β（ng/L）分别为（94±19）、（133±19）、（174±22）、（180±30）、（279±38）（P＜0.05）.结论 单纯性肥胖及肥胖伴不同组分代谢紊乱患者的血清可不同程度地诱导TLR4/NF-κB信号通路的活化,伴代谢紊乱组分越多的肥胖患者的血清诱导TLR4/NF-κB信号通路活化的能力越强.

Objective To investigate the activation of TLR4/NF-κB signaling pathway and the level of inflammation in volunteers with varying degrees of metabolic disorders and the effect of intervention by TLR4 monoclonal antibody. Methods THP-1 cell line was cultured with 10% serum of volunteers with various degrees of metabolic disorders（each 10 cases） for 24,48 h. TLR4 and NF-κB p65 phosphorylation protein were measured by Western blot. TLR4 mRNA was measured by RT-PCR. The expression of IL-1 β and TNF-α were detected by ELISA. Results The TLR4 mRNA and protein, the level of NF-κB p65 phosphorylation protein in THP-1 cell line and IL-1β and TNF-α expression in culture supernatant in normal, simple obesity, obesity with hyperglycemia, obesity with hyperlipidemia, obesity with three metabolic disorders groups had statistical significance（ P ＜ 0. 05 ）. The TLR4 mRNA and protein, the level of NF-κB p65 phosphorylation protein and the IL-1β and TNF-α expression in obesity with three metabolic disorders group were higher than those in other group, and those were time-dependent （ P ＜0. 05 ）. TNF-α expression in normal, simple obesity, obesity with hyperglycemia, obesity with hyperlipidemia, obesity with three metabolic disorders group at 48 h were （222 ± 32）, （246 ± 52 ）, （ 322 ± 32 ）, （ 322 ± 34） and （490 ±83） ng/L, respectively, IL-1β（ng/L）were （94 ± 19）, （ 133 ± 19）, （ 174 ±22）, （ 180 ±30）, （279 ±38） （P ＜0. 05）. Conclusions The activation of TLR4/NF-κB signaling pathway on THP-1 cell line which is cultured by serum with various degrees of metabolic disorders is different, with the increasing of severity of metabolic disorders, TLR4/NF-κB signaling pathway activation levels also incr.