Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of mouse spleen after immunized using LPS vs MPL as adjuvant.

ABSTRACT: An unresolved issue in immunology is the extent to which inflammatory effects are needed for robust T cell responses. In this study, mice were immunized by iv injection using either high toxicity lipopolysaccharide (LPS) or low toxicity monophosphoryl lipid A (MPL) as adjuvant. Six hours after iv immunization, whole spleens were harvested and gene expression was measured in unfractionated splenic populations of cells. The analysis indicated that the low toxicity adjuvanticity of MPL was associated with TLR4-mediated signaling that was biased to the TRIF branch of TLR4, while LPS generated balanced MyD88 and TRIF-associated outcomes. Experiment Overall Design: B6.SJL mice adoptively transferred with T cells from B6.OTI and OTII transgenic TCR transgenic were immunized via intravenous injection with ova peptides alone ("Ova"), or with LPS ("OvaLPS") or MPL as adjuvant ("OvaMPL"). Six hours after immunization, the mice were euthanized by cervical dislocation, the spleens were removed and immediately lysed in guanidium chloride, and kept frozen until being used to extract total cellular RNA. Three mice each were given the indicated treatments, with independent processing and analysis of nine samples total.

ORGANISM(S): Mus musculus

SUBMITTER: Thomas Clark Mitchell

PROVIDER: E-GEOD-7768 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

The vaccine adjuvant monophosphoryl lipid A as a TRIF-biased agonist of TLR4.

Mata-Haro Verónica V Cekic Caglar C Martin Michael M Chilton Paula M PM Casella Carolyn R CR Mitchell Thomas C TC

Science (New York, N.Y.) 20070601 5831

The inflammatory toxicity of lipopolysaccharide (LPS), a component of bacterial cell walls, is driven by the adaptor proteins myeloid differentiation factor 88 (MyD88) and Toll-interleukin 1 receptor domain-containing adapter inducing interferon-beta (TRIF), which together mediate signaling by the endotoxin receptor Toll-like receptor 4 (TLR4). Monophosphoryl lipid A (MPLA) is a low-toxicity derivative of LPS with useful immunostimulatory properties, which is nearing regulatory approval for use ...[more]

PMID: 17569868

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Project description:Background: Toll-like family of receptors recognizes pathogen-associated molecular patterns (PAMPs) from different organisms. TLR4 is the receptor for bacterial lipopolysaccharide (LPS), dsRNA viral mimic poly(I:C) binds to TLR3, and bacterial CpG DNA signals through TLR9. TLR4 signaling is mediated by adaptor molecules Myd88 and TRIF while TLR3 pathway involves only the TRIF adaptor and TLR9 signals solely through Myd88. Methods: To identify genes other than those in TLR pathways that mediate macrophage response to different PAMPs, RAW264.7 cells were stimulated with LPS, poly(I:C), or CpG DNA, and RNA was profiled on microarrays 6 hrs and 24 hrs post-treatment. Gene expression data were analyzed to determine genes, pathways and transcriptional networks that are in common and unique to each of the three TLR stimuli. Potentially novel candidates revealed by this analysis were tested for their role in innate immunity using RNA interference. Results: Many genes are differentially regulated by LPS and poly(I:C) at both 6 hrs and 24 hrs following treatment, while CpG DNA elicits a much less pronounced transcriptional response. By analyzing gene expression data for networks and pathways, we prioritized differentially expressed genes that are in common to all three PAMPs as well as those shared by LPS and poly(I:C). Knockdown by RNA interference of two genes, Plec1 and TPST1, inhibited production of IL-6 in response to LPS in cultured macrophages. Conclusions: We have identified novel innate immunity genes that may be important in macrophage response to LPS, poly(I:C), and CpG DNA stimuli. Our results provide potential biomarkers and therapeutic targets that should be further investigated in mice and human populations. Keywords: time course For each treatment (Sigma LPS, LIST LPS, and media only), three biological replicates (separate macrophage cultures and RNA isolations) were profiled. Each sample was labeled with Cy3 and Cy5 and co-hybridized with Stratagene Universal Mouse Reference (dye flip techical replicates). Expression at 2 timepoints (6 and 24 hours post-treatment) was assessed.

Project description:Molecular Pathways and Transcriptional Networks Involved in the Macrophage Response to LPS, poly(I:C) and CpG DNA Stimulation Background: Toll-like family of receptors recognizes pathogen-associated molecular patterns (PAMPs) from different organisms. TLR4 is the receptor for bacterial lipopolysaccharide (LPS), dsRNA viral mimic poly(I:C) binds to TLR3, and bacterial CpG DNA signals through TLR9. TLR4 signaling is mediated by adaptor molecules Myd88 and TRIF while TLR3 pathway involves only the TRIF adaptor and TLR9 signals solely through Myd88. Methods: To identify genes other than those in TLR pathways that mediate macrophage response to different PAMPs, RAW264.7 cells were stimulated with LPS, poly(I:C), or CpG DNA, and RNA was profiled on microarrays 6 hrs and 24 hrs post-treatment. Gene expression data were analyzed to determine genes, pathways and transcriptional networks that are in common and unique to each of the three TLR stimuli. Potentially novel candidates revealed by this analysis were tested for their role in innate immunity using RNA interference. Results: Many genes are differentially regulated by LPS and poly(I:C) at both 6 hrs and 24 hrs following treatment, while CpG DNA elicits a much less pronounced transcriptional response. By analyzing gene expression data for networks and pathways, we prioritized differentially expressed genes that are in common to all three PAMPs as well as those shared by LPS and poly(I:C). Knockdown by RNA interference of two genes, Plec1 and TPST1, inhibited production of IL-6 in response to LPS in cultured macrophages. Conclusions: We have identified novel innate immunity genes that may be important in macrophage response to LPS, poly(I:C), and CpG DNA stimuli. Our results provide potential biomarkers and therapeutic targets that should be further investigated in mice and human populations. For each treatment (LPS, polyIC, CpG DNA, media only), three biological replicates (separate macrophage cultures and RNA isolations) were profiled. Each sample was labeled with Cy3 and Cy5 and co-hybridized with Stratagene Universal Mouse Reference (dye flip techical replicates). Expression at 2 timepoints (6 and 24 hours post-treatment) was assessed.

Dataset Information

Transcription profiling of mouse spleen after immunized using LPS vs MPL as adjuvant.

Publications

The vaccine adjuvant monophosphoryl lipid A as a TRIF-biased agonist of TLR4.

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