NR4A orphan nuclear receptor family members, NR4A2 and NR4A3, regulate neutrophil number and survival

Defects in neutrophil number and survival are common to both hematologic disorders and chronic inflammatory diseases. At sites of inflammation, neutrophils respond to multiple signals that activate protein kinase A (PKA) signalling, which positively regulates neutrophil survival. We aimed to study the transcriptional responses to several stimuli in human neutrophils. We used microarrays to detail the global programme of gene expression and identified distinct classes of up-regulated genes, which pointing to a key role of NR4A receptors in regulation of neutrophil lifespan and homeostasis. Homo sapiens neutrophil total RNA extracts were produced after 4h incubation in each specific treatment (i.e. GMCSF, hypoxia, TLR4 agonist, PKA agonist and negative control LPS media, from the latter we also collected supernatant). These were converted to cDNA and cDNA from 5 donors was pooled prior hybridization on Affymetrix GeneChip Human Genome U133 Plus 2.0 Array.

中性粒细胞数量与存活异常常见于血液系统疾病与慢性炎症性疾病中。在炎症部位，中性粒细胞会响应多种激活蛋白激酶A（PKA）信号通路的信号，该通路可正向调控中性粒细胞的存活。本研究旨在探究人类中性粒细胞对多种刺激物的转录应答反应。我们通过基因芯片技术解析全局基因表达谱，并鉴定出多类不同的上调基因，结果表明NR4A受体在中性粒细胞寿命与稳态调控中发挥关键作用。将人类中性粒细胞在各特定处理条件（即粒细胞-巨噬细胞集落刺激因子（GMCSF）、缺氧、Toll样受体4（TLR4）激动剂、蛋白激酶A（PKA）激动剂，以及阴性对照脂多糖（LPS）培养基，同时从阴性对照培养基中收集上清液）中孵育4小时后提取总RNA。将上述总RNA反转录为互补DNA（cDNA），并将来自5名供体的cDNA混合后，在Affymetrix GeneChip人类基因组U133 Plus 2.0阵列上进行杂交实验。