微小RNA-155在类风湿关节炎外周血单个核细胞中的表达及意义

Altered expression of miR-155 in peripheral blood mononuclear cells of rheumatoid arthritis patients

目的了解RA患者PBMCs中差异表达的微小RNA（miRNAs，miR），探讨RA患者PBMCs中miR-155表达与RA的相关性，并了解炎症因子对miR-155表达水平的作用。方法①提取RA患者及健康对照各5例PBMCs总RNA。通过微阵列芯片技术分析miRNAs表达谱。②在芯片检测结果基础上，选择与免疫调节功能密切相关的miR-155，利用实时荧光定量（RT）-PCR测定26例RA患者和23名健康对照者的PBMC中miR-155的表达水平。③评价miR-155的表达水平与RA患者临床及实验室指标的相关性。④利用RT-PCR测定在TNF-α，IFN-γ和脂多糖刺激下miR-155表达水平的变化。采用独立样本t检验，配对t检验，F检验以及Spearman相关分析进行统计学处理。结果①miRNAs微阵列芯片表达谱提示RA患者PBMC中有14种miRNAs信号强度差别在2倍以上，其中miR-155高表达（t=9．218，P=0．001）。②miR-155的表达与血浆CRP的水平呈正相关（r=0．57，P=0．002）。③在TNF．d刺激RA患者PBMCs12、24、48h时，miR-155的表达上调，分别为11．1±4．0、2．5±1．6（t=-8．53，P〈0．01）；9．5±3．6、2．5±1．6（t=-8．042，P〈0．001）；8．3±3．0、2．5±1．6（t=-8．416，P〈0．001）。IFN-γ刺激12、24、48h时，miR-155的表达上调，分别为27．5±15．0、4．6±2．8（t=-4．613，P=0．006）；23．8±11．1、4．6±2．8（t=-5．678，P=0．002）；19．8±10．0、4．6±2．8（t=-5．118，P=0．004）。脂多糖刺激12、24、48h时，miR-155表达水平亦显著上调，分别为11．4±6．3、3．8±2．1（t=-4．285，P=0．008）；9．6±5．0、3．8±2．1（t=-4．495，P=0．006）；7．9±4．0、3．8±2．1（t=-4．771，P=0．005）。结论RA患者PBMCs中miR-155的表达上调与炎症因子诱导相关，可能参与了RA发病过程的调节。

Objective （1） To Screen for the miRNAs differently expressed in the peripheral blood mono-nuclear ceils （PBMCs） of rheumatoid arthritis （RA） by microarray experiments. （2） To further evaluate the expression of miR-155 in PBMCs of RA. （3） To determine the relevance between the expression of miR-155 and clinical as well as laboratory features. （4） To test whether inflammatory mediators can induce miR-155 expression in PBMCs of RA. Methods （1） Total RNA was isolated from peripheral blood mononuclear cells obtained from 5 patients of RA and 5 normal controls. Expression profiling of miRNAs was performed in a microarray analysis. （2） MiR-155 was identified for further study by stem-loop real-time RT-PCR based on SYBR-Green. PBMC from 26 patients of RA and 23 normal controls were collected. （3） Association between miR-155 and the clinical and laboratory features of RA was evaluated. （4）Induction of miR-155 following stimulation with TNF-α, IFN-γ and LPS of cultures of RA PBMCs was examined by real-time RT-PCR. Statistical analysis was done with student＇s t test, paired t test, and ANOVA, Spearman correlation. Results （1） Expression profiling of miRNAs revealed significant differential expression of 14 miRNAs, of which signal intensity changed over two times. MiR-155 was up-regulated in PBMCs of RA than in normal controls （t=9.218, P=0.001）. （2） The expression level of miR-155 had a positive correlation with serum CRP level （r=0.57, P= 0.002）. （3） Expression of miR-155 was markedly up-regulated in PBMCs of RA after stimulated with TNF-α, IFN-γ and LPS, especially with TNF-α. Conclusions The expression of miR-155 is induced by stimulating with TNF-α, IFN-γand LPS. MiR-155 may be a regulator in RA pathogenesis. Further studies are required to elucidate the function of miR-155.