Sanz-Santos Gema, Jiménez-Marín Angeles, Bautista Rocío, Fernández Noé, Claros Gonzalo M, Garrido Juan J
Grupo de Genómica y Mejora Animal, Departamento de Genética, Facultad de Veterinaria, Universidad de Córdoba, Campus de Rabanales, Edificio Gregor Mendel C5, 14071 Córdoba, Spain.
BMC Proc. 2011 Jun 3;5 Suppl 4(Suppl 4):S11. doi: 10.1186/1753-6561-5-S4-S11.
Experimental exposure of swine neutrophils to bacterial lipopolysaccharide (LPS) represents a model to study the innate immune response during bacterial infection. Neutrophils can effectively limit the infection by secreting lipid mediators, antimicrobial molecules and a combination of reactive oxygen species (ROS) without new synthesis of proteins. However, it is known that neutrophils can modify the gene expression after LPS exposure. We performed microarray gene expression analysis in order to elucidate the less known transcriptional response of neutrophils during infection.
Blood samples were collected from four healthy Iberian pigs and neutrophils were isolated and incubated during 6, 9 and 18 hrs in presence or absence of lipopolysaccharide (LPS) from Salmonella enterica serovar Typhimurium. RNA was isolated and hybridized to Affymetrix Porcine GeneChip®. Microarray data were normalized using Robust Microarray Analysis (RMA) and then, differential expression was obtained by an analysis of variance (ANOVA).
ANOVA data analysis showed that the number of differentially expressed genes (DEG) after LPS treatment vary with time. The highest transcriptional response occurred at 9 hr post LPS stimulation with 1494 DEG whereas at 6 and 18 hr showed 125 and 108 DEG, respectively. Three different gene expression tendencies were observed: genes in cluster 1 showed a tendency toward up-regulation; cluster 2 genes showing a tendency for down-regulation at 9 hr; and cluster 3 genes were up-regulated at 9 hr post LPS stimulation. Ingenuity Pathway Analysis revealed a delay of neutrophil apoptosis at 9 hr. Many genes controlling biological functions were altered with time including those controlling metabolism and cell organization, ubiquitination, adhesion, movement or inflammatory response.
LPS stimulation alters the transcriptional pattern in neutrophils and the present results show that the robust transcriptional potential of neutrophils under infection conditions, indicating that active regulation of gene expression plays a major role in the neutrophil-mediated- innate immune response.
将猪中性粒细胞实验性暴露于细菌脂多糖（LPS）是研究细菌感染期间固有免疫反应的一种模型。中性粒细胞可通过分泌脂质介质、抗菌分子以及活性氧（ROS）的组合而无需新合成蛋白质来有效限制感染。然而，已知中性粒细胞在LPS暴露后可改变基因表达。我们进行了微阵列基因表达分析，以阐明感染期间中性粒细胞鲜为人知的转录反应。
从4头健康的伊比利亚猪采集血样，分离中性粒细胞，并在有或无鼠伤寒沙门氏菌血清型鼠伤寒脂多糖（LPS）的情况下孵育6、9和18小时。分离RNA并与Affymetrix猪基因芯片®杂交。微阵列数据使用稳健微阵列分析（RMA）进行标准化，然后通过方差分析（ANOVA）获得差异表达。
ANOVA数据分析表明，LPS处理后差异表达基因（DEG）的数量随时间变化。LPS刺激后9小时转录反应最高，有1494个DEG，而在6小时和18小时分别有125个和108个DEG。观察到三种不同的基因表达趋势：聚类1中的基因显示出上调趋势；聚类2中的基因在9小时显示出下调趋势；聚类3中的基因在LPS刺激后9小时上调。 Ingenuity通路分析显示9小时时中性粒细胞凋亡延迟。许多控制生物学功能的基因随时间发生改变，包括那些控制代谢、细胞组织、泛素化、粘附、运动或炎症反应的基因。
LPS刺激改变了中性粒细胞的转录模式，目前的结果表明在感染条件下中性粒细胞具有强大的转录潜力，表明基因表达的主动调节在中性粒细胞介导的固有免疫反应中起主要作用。
