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Transmigration across activated endothelium induces transcriptional changes, inhibits apoptosis, and decreases antimicrobial protein expression in human monocytes.

ABSTRACT: We investigated the hypothesis that transmigration drives monocyte transcriptional changes. Using Agilent whole human genome microarrays, we identified over 692 differentially expressed genes (2x, P<0.05) in freshly isolated human monocytes following 1.5 h of transmigration across IL-1beta-stimulated ECs compared with untreated monocytes. Genes up-regulated by monocyte transmigration belong to a number of over-represented functional groups including immune response and inhibition of apoptosis. qRT-PCR confirmed increased expression of MCP-1 and -3 and of NAIP following monocyte transmigration. Additionally, quantification of Annexin V binding revealed a reduction in apoptosis following monocyte transmigration. Comparison of gene expression in transmigrated monocytes with additional controls (monocytes that failed to transmigrate and monocytes incubated beneath stimulated ECs) revealed 89 differentially expressed genes, which were controlled by the process of diapedesis. Functional annotation of these genes showed down-regulation of antimicrobial genes (e.g., alpha-defensin down 50x, cathelicidin down 9x, and CTSG down 3x). qRT-PCR confirmed down-regulation of these genes. Immunoblots confirmed that monocyte diapedesis down-regulates alpha-defensin protein expression. However, transmigrated monocytes were functional and retained intact cytokine and chemokine release upon TLR ligand exposure. Overall, these data indicate that the process of monocyte transmigration across stimulated ECs promotes further monocyte recruitment and inhibits monocyte apoptosis. Unexpectedly, following transmigration, monocytes displayed reduced antimicrobial protein expression.

SUBMITTER: Williams MR

PROVIDER: S-EPMC2780914 | biostudies-literature | 2009 Dec

REPOSITORIES: biostudies-literature

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Transmigration across activated endothelium induces transcriptional changes, inhibits apoptosis, and decreases antimicrobial protein expression in human monocytes.

Williams Marcie R MR Sakurai Yumiko Y Zughaier Susu M SM Eskin Suzanne G SG McIntire Larry V LV

Journal of leukocyte biology 20090825 6

We investigated the hypothesis that transmigration drives monocyte transcriptional changes. Using Agilent whole human genome microarrays, we identified over 692 differentially expressed genes (2x, P<0.05) in freshly isolated human monocytes following 1.5 h of transmigration across IL-1beta-stimulated ECs compared with untreated monocytes. Genes up-regulated by monocyte transmigration belong to a number of over-represented functional groups including immune response and inhibition of apoptosis. q ...[more]

PMID: 19706840

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Project description:BackgroundAnthrax lethal toxin (LT), produced by the Gram-positive bacterium Bacillus anthracis, is a highly effective zinc dependent metalloprotease that cleaves the N-terminus of mitogen-activated protein kinase kinases (MAPKK or MEKs) and is known to play a role in impairing the host immune system during an inhalation anthrax infection. Here, we present the transcriptional responses of LT treated human monocytes in order to further elucidate the mechanisms of LT inhibition on the host immune system.ResultsWestern Blot analysis demonstrated cleavage of endogenous MEK1 and MEK3 when human monocytes were treated with 500 ng/mL LT for four hours, proving their susceptibility to anthrax lethal toxin. Furthermore, staining with annexin V and propidium iodide revealed that LT treatment did not induce human peripheral monocyte apoptosis or necrosis. Using Affymetrix Human Genome U133 Plus 2.0 Arrays, we identified over 820 probe sets differentially regulated after LT treatment at the p <0.001 significance level, interrupting the normal transduction of over 60 known pathways. As expected, the MAPKK signaling pathway was most drastically affected by LT, but numerous genes outside the well-recognized pathways were also influenced by LT including the IL-18 signaling pathway, Toll-like receptor pathway and the IFN alpha signaling pathway. Multiple genes involved in actin regulation, signal transduction, transcriptional regulation and cytokine signaling were identified after treatment with anthrax LT.ConclusionWe conclude LT directly targets human peripheral monocytes and causes multiple aberrant gene responses that would be expected to be associated with defects in human monocyte's normal signaling transduction pathways and function. This study provides further insights into the mechanisms associated with the host immune system collapse during an anthrax infection, and suggests that anthrax LT may have additional downstream targets outside the well-known MAPK pathway.

Dataset Information

Transmigration across activated endothelium induces transcriptional changes, inhibits apoptosis, and decreases antimicrobial protein expression in human monocytes.

Publications

Transmigration across activated endothelium induces transcriptional changes, inhibits apoptosis, and decreases antimicrobial protein expression in human monocytes.

Similar Datasets

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