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Gene expression pattern in swine neutrophils after lipopolysaccharide exposure: a time course comparison.

Sanz-Santos G, Jiménez-Marín A, Bautista R, Fernández N, Claros GM, Garrido JJ - BMC Proc (2011)

Bottom Line: 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.Many genes controlling biological functions were altered with time including those controlling metabolism and cell organization, ubiquitination, adhesion, movement or inflammatory response.

View Article: PubMed Central - HTML - PubMed

Affiliation: 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. ge1gapaj@uco.es.

ABSTRACT

Background: 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.

Methods: 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).

Results: 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.

Conclusions: 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.

No MeSH data available.


Related in: MedlinePlus

Differentially expressed genes grouped into three different clusters. Cluster 1 contains 8 genes with up-regulation tendency through the time course. 747 genes belonging the cluster 2, with a down-regulation tendency at 9 hr. Opposite tendency can be observed in the cluster 3, where 335 genes show an up-regulation at 9 hr and down-regulation at 18 hr.
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Figure 2: Differentially expressed genes grouped into three different clusters. Cluster 1 contains 8 genes with up-regulation tendency through the time course. 747 genes belonging the cluster 2, with a down-regulation tendency at 9 hr. Opposite tendency can be observed in the cluster 3, where 335 genes show an up-regulation at 9 hr and down-regulation at 18 hr.

Mentions: In this study, we performed microarray analysis in order to elucidate the response of the porcine neutrophils to LPS from S. Typhimuriun along a time course of 6, 9 and 18 hr in vitro stimulation. To achieve that, ANOVA adapted for time course experiments was used, in order to identify genes whose temporal expression patterns differ between control and LPS-treated samples. As a result, 1090 genes were differentially expressed along the time course and the resulting hierarchical clustering is shown in Figure 1. Moreover, genes were clustered according to idealized expression patterns. The query gene is assigned to a cluster designated by the idealized pattern that has the maximal correlation with that gene. Figure 2 shows the three gene expression clusters obtained in this analysis. This criterion for classifying genes according to their similarity to an idealized expression pattern allows us to determine the gene expression tendency through the time course. Clustering results reveal distinct temporal patterns and the most significant transcriptional changes happened after 9 hr post LPS exposure. Individual ANOVA tests for each time point confirm this finding as shown in Figure 3. Thus, at 6 and 18 hr, there are 125 and 108 DEG, respectively, whereas at 9 hr, there are 1494 DEG.


Gene expression pattern in swine neutrophils after lipopolysaccharide exposure: a time course comparison.

Sanz-Santos G, Jiménez-Marín A, Bautista R, Fernández N, Claros GM, Garrido JJ - BMC Proc (2011)

Differentially expressed genes grouped into three different clusters. Cluster 1 contains 8 genes with up-regulation tendency through the time course. 747 genes belonging the cluster 2, with a down-regulation tendency at 9 hr. Opposite tendency can be observed in the cluster 3, where 335 genes show an up-regulation at 9 hr and down-regulation at 18 hr.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3108205&req=5

Figure 2: Differentially expressed genes grouped into three different clusters. Cluster 1 contains 8 genes with up-regulation tendency through the time course. 747 genes belonging the cluster 2, with a down-regulation tendency at 9 hr. Opposite tendency can be observed in the cluster 3, where 335 genes show an up-regulation at 9 hr and down-regulation at 18 hr.
Mentions: In this study, we performed microarray analysis in order to elucidate the response of the porcine neutrophils to LPS from S. Typhimuriun along a time course of 6, 9 and 18 hr in vitro stimulation. To achieve that, ANOVA adapted for time course experiments was used, in order to identify genes whose temporal expression patterns differ between control and LPS-treated samples. As a result, 1090 genes were differentially expressed along the time course and the resulting hierarchical clustering is shown in Figure 1. Moreover, genes were clustered according to idealized expression patterns. The query gene is assigned to a cluster designated by the idealized pattern that has the maximal correlation with that gene. Figure 2 shows the three gene expression clusters obtained in this analysis. This criterion for classifying genes according to their similarity to an idealized expression pattern allows us to determine the gene expression tendency through the time course. Clustering results reveal distinct temporal patterns and the most significant transcriptional changes happened after 9 hr post LPS exposure. Individual ANOVA tests for each time point confirm this finding as shown in Figure 3. Thus, at 6 and 18 hr, there are 125 and 108 DEG, respectively, whereas at 9 hr, there are 1494 DEG.

Bottom Line: 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.Many genes controlling biological functions were altered with time including those controlling metabolism and cell organization, ubiquitination, adhesion, movement or inflammatory response.

View Article: PubMed Central - HTML - PubMed

Affiliation: 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. ge1gapaj@uco.es.

ABSTRACT

Background: 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.

Methods: 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).

Results: 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.

Conclusions: 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.

No MeSH data available.


Related in: MedlinePlus