Gene expression profiling of Huh7 and THP1 cells after siRNA knockdown of C5/TRAF1 intergenic transcript. Homo sapiens

The C5/TRAF1 locus is a well validated genetic risk factor for RA. Upon further investigation of this region we recently discovered that the intergenic region between these two immune-related genes is activelly transcribed in a variety of tissues and is up-regulated in PBMCs, monocytes, B- and T-cells upon immune stimulation. The present study aimed to characterize the biological function of this newly identified C5/TRAF1 intergenic transcript, by analysing the effect of its knockdow on the gene expression profiles of Huh7 and THP1 cells. Overall design: siRNA transfections with the intergenic specific siRNA and 2 negative control siRNAs were performed in duplo and independently repeated 4 times per cell line, with different cell culture passages to account for the biological variation. The 4 independent experiments per cell line were divided in two design phases: the Discovery Phase (experiments 1 and 2) and the Validation Phase (experiments 3 and 4).

C5/TRAF1基因座是经过充分验证的类风湿关节炎（Rheumatoid Arthritis, RA）遗传风险因子。我们近期对该区域展开深入研究后发现，上述两个免疫相关基因之间的基因间区可在多种组织中被活跃转录，且在免疫刺激条件下，其在外周血单个核细胞（Peripheral Blood Mononuclear Cells, PBMCs）、单核细胞、B细胞及T细胞中的表达水平显著上调。本研究旨在对这一新发现的C5/TRAF1基因间区转录本的生物学功能进行系统表征，具体通过分析其基因敲低（knockdown）对Huh7细胞与THP1细胞基因表达谱的影响。总体实验设计：针对基因间区特异性小干扰RNA（small interfering RNA, siRNA）以及2种阴性对照siRNA的转染实验均一式两份开展，且每个细胞系独立重复实验4次，通过使用不同传代批次的细胞以控制生物学变异。每个细胞系的4次独立实验被划分为两个实验阶段：发现阶段（实验1与实验2）与验证阶段（实验3与实验4）。