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SRP042975
Danio rerio
4 Downloadable Samples
IlluminaHiSeq2000
Description
Scavenger receptors on the cell surface of macrophages play an important role in host defence through their ability to bind microbial ligands and induce phagocytosis. Concurrently, signal transduction pathways are initiated that aid in defence mechanisms against the invading microbe. Here we report on the function of scavenger receptor Marco (macrophage receptor with collagenous structure) during infection of zebrafish embryos with Mycobacterium marinum, a close relative of Mycobacterium tuberculosis. Morpholino knockdown demonstrates that Marco is required for the rapid phagocytosis of M. marinum following intravenous infection. Furthermore, gene expression analysis shows that Marco controls the initial transient pro-inflammatory response to M. marinum and remains a determining factor for the immune response signature at later stages of infection. Increased bacterial burden following marco knockdown indicates that this scavenger receptor is important for control of M. marinum growth, likely due to delayed phagocytosis and reduced pro-inflammatory signalling observed under conditions of Marco deficiency Overall design: Embryos were injected at the one cell stage with a morpholino targeting marco, or with the standard control morpholino from GeneTools for comparison. Subsequently, at 24 hours post fertilization (hpf) the morphants and their controls were manually dechorionated at 24 hpf and at 28 hpf they were infected by injecting 200 colony forming units of M. marinum Mma20 into the caudal vein, or mock-injected with PBS/2%PVP. After injections embryos were transferred into fresh egg water containing 0.003% 1-phenyl-2-thiourea (Sigma-Aldrich) to prevent melanization and incubated for 4 days at 28°C. After the incubation period, infected and uninfected morphants, mutants and their controls were imaged and groups of 30 embryos were snap-frozen in liquid nitrogen and RNA was isolated for Illumina RNAseq analysis.
Publication Title
Phagocytosis of mycobacteria by zebrafish macrophages is dependent on the scavenger receptor Marco, a key control factor of pro-inflammatory signalling.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 155 Downloadable Samples
Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
This SuperSeries is composed of the SubSeries listed below.
Publication Title
Genomic profiling of CHEK2*1100delC-mutated breast carcinomas.
Sample Metadata Fields
Specimen part
View Samples- Danio rerio
- 72 Downloadable Samples
- IlluminaHiSeq2000
Description
Both embryonic and adult zebrafish Mycobacterium marinum infection studies have contributed to our knowledge of the development and function of tuberculous granulomas, which are typical for mycobacterial pathogenesis. In this review we discuss how transcriptome profiling studies have helped to characterize this infection process and we include new RNA sequencing (RNA-Seq) data that reveals three main phases in the host response to M. marinum during the early stages of granuloma development in zebrafish embryos and larvae. The late-phase response shares common components with the strong and acute host transcriptome response that has previously been reported for S. typhimurium infection in zebrafish embryos. In contrast, the early/mid-phase response to M. marinum infection, characterized by suppressed pro-inflammatory signaling, is strikingly different from the acute response to S. typhimurium infection. Furthermore, M. marinum infection shows a collective and strongly fluctuating regulation of lipoproteins, while S. typhimurium infection has pronounced effects on amino acid metabolism and glycolysis. Overall design: Embryos were infected at 28 hpf by injecting 250 colony forming units of M. marinum Mma20 in 2%PVP into the caudal vein, or mock-injected with PBS/2%PVP. After injections, embryos were transferred into fresh egg water containing 0.003% 1-phenyl-2-thiourea (Sigma-Aldrich) to prevent melanization and incubated at 28°C. After the incubation period, infected and uninfected groups of 30 embryos were snap-frozen in liquid nitrogen and RNA was isolated for Illumina RNAseq analysis. Samples were taken at the following timepoints: 2, 4, 6, 8 hpi and 1, 2, 3, 4, 5 dpi.
Publication Title
Transcriptomic Approaches in the Zebrafish Model for Tuberculosis-Insights Into Host- and Pathogen-specific Determinants of the Innate Immune Response.
Sample Metadata Fields
No sample metadata fields
View Samples- Homo sapiens
- 20 Downloadable Samples
- Affymetrix Human Genome U133 Plus 2.0 Array (hgu133plus2)
Description
To study differentially expressed genes in neuro-ectodermal cell lines
Publication Title
Downregulation of Axl in non-MYCN amplified neuroblastoma cell lines reduces migration.
Sample Metadata Fields
Sex, Specimen part
View Samples- Danio rerio
- 12 Downloadable Samples
- IlluminaHiSeq2500
Description
We use the Tlr2 mutant of zebrafish embryos model to study the transcriptome response to Mycobacterium marinum infection. We injected M.marinum into the caudal vein at 28 hours post fertilization and took samples at 4 days post infection. Overall design: This deep sequence study was designed to determine the gene expression profile in the Tlr2 mutant and heterozygote by M.marinum infection. RNA was isolated at 4 days post infection. Tlr2 mutants and heterozygotes zebrafish embryos were micro-injected into the caudal vein with 150CFU M.marinum, or PBS as a control at 28 hours post fertilization. After injections embryso were transerred into fresh egg water and incubated at 28 degree. At 4 days post infection triplicateds of 10 embryos per condition were snapfrozen in liquid nitogen, and total RNA was isolated using TRIZOL reagent.
Publication Title
Infection and RNA-seq analysis of a zebrafish tlr2 mutant shows a broad function of this toll-like receptor in transcriptional and metabolic control and defense to Mycobacterium marinum infection.
Sample Metadata Fields
No sample metadata fields
View Samples- Danio rerio
- 9 Downloadable Samples
- Affymetrix Zebrafish Genome Array (zebrafish)
Description
In this study, zebrafish ZF4 and PAC2 cells were seeded in 0.5% or 1% FCS, respectively, and grown to 85% confluence and subsequently cultured for 24 hours without serum. Then they were treated with either medium without serum or medium with serum (ZF4 in 10% FCS and PAC2 in 15% FCS).After 6 hours, RNA was extracted from the cells and analyzed using the Affymetrix GeneChip Zebrafish Genome Array (GeneChip 430). There are 15502 oligonucleotide sets on each Affymetrix chip, 14895 of which can be linked to a UniGene assignment (Unigene data set 06-12-2005).
Publication Title
Genetic and transcriptome characterization of model zebrafish cell lines.
Sample Metadata Fields
Cell line, Compound
View Samples- Danio rerio
- 4 Downloadable Samples
- IlluminaGenomeAnalyzerII
Description
Transcriptional profiling of the zebrafish embryonic host response to a systemic bacterial infection with Salmonella typhimurium (strain SL1027); comparison between traf6 knock-down and control morpholino treated embryos. Overall design: All infection experiments were performed using mixed egg clutches of ABxTL strain zebrafish. Embryos injected with traf6 morpholino or a 5bp mismatch control morpholino were staged at 27 hours post fertilization (hpf) by morphological criteria and approximately 250 cfu of DsRed expressing Salmonella bacteria were injected into the caudal vein close to the urogenital opening. As a control an equal volume of PBS was likewise injected. Pools of 20-40 infected and control embryos were collected 8 hours post infection (hpi). The whole procedure was preformed in triplicate on separate days. Total RNA of the biological triplicates was pooled using equal amounts of RNA prior to RNAseq library preparation.
Publication Title
Transcriptome analysis of Traf6 function in the innate immune response of zebrafish embryos.
Sample Metadata Fields
No sample metadata fields
View Samples- Danio rerio
- 24 Downloadable Samples
- Illumina HiSeq 2500
Description
Glucocorticoid drugs are widely used to treat immune-related diseases, but their use is limited by side effects and by resistance, which especially occurs in macrophage-dominated diseases. In order to improve glucocorticoid therapies, more research is required into the mechanisms of glucocorticoid action. In the present study, we have used a zebrafish model for inflammation to study glucocorticoid effects on the innate immune response. In zebrafish larvae, the migration of neutrophils towards a site of injury is inhibited by the synthetic glucocorticoid beclomethasone, while migration of macrophages is glucocorticoid resistant. RNA sequencing was done on on Fluorescence-Activated Cell Sorting (FACS)-sorted macrophages.The results show that the vast majority of the wounding-induced transcriptional changes in these cells are inhibited by beclomethasone, whereas a small subset is glucocorticoid-insensitive. As a result, beclomethasone decreases the number of macrophages that differentiate towards a pro-inflammatory (M1) phenotype, which we demonstrated using a tnfa:eGFP-F reporter line and analysis of macrophage morphology. We conclude that the glucocorticoid resistance of the wounding-induced macrophage migration is due to the insensitivity of the induction of macrophage-specific chemoattractants to glucocorticoid inhibition, which may explain the relative resistance of macrophage-dominated diseases to glucocorticoid therapy. However, the induction of pro-inflammatory genes in macrophages is strongly attenuated, which inhibits their differentiation to an M1 phenotype. Overall design: After anesthesia with 0.02% aminobenzoic acid ethyl ester (tricaine, Sigma Aldrich), the tails of 3 days post fertilization (dpf) embryos were partially amputated with a 1mm sapphire blade (World Precision Instruments) on 2% agarose-coated Petri dishes under a Leica M165C stereomicroscope (Chatzopoulou et al., 2016). Amputated and non-amputated (control) embryos were pretreated for 2 hours with 25 µM beclomethasone (Sigma Aldrich) or vehicle (0.05% dimethyl sulfoxide (DMSO)) in egg water prior to amputation and received the same treatment after the amputation. Macrophages were sorted from Tg(mpeg1.4:mCherry-F) embryos as previously described (Rougeot et al., 2014; Zakrzewska et al., 2010) at 4 hours post amputation (hpa). The sorted cells were collected in QIAzol lysis reagent (Qiagen) for RNA isolation. Extracted total RNA was amplified using the SMART-seq V4 kit (Clontech) for sequencing. The RNA seq libraries generated with the SMART-seq V4 kit were sequenced using an Illumina HiSeq 2500 instrument according to the manufacturer's instructions with a read length of 50 nucleotides.
Publication Title
Glucocorticoids inhibit macrophage differentiation towards a pro-inflammatory phenotype upon wounding without affecting their migration.
Sample Metadata Fields
Treatment, Subject
View Samples- Danio rerio
- 12 Downloadable Samples
- IlluminaHiSeq2000
Description
Macrophage expressed gene 1 (MPEG1) encodes an evolutionary conserved protein with a predicted Membrane Attack Complex/Perforin domain associated with host defence against invading pathogens. In vertebrates, MPEG1 is an integral membrane protein of macrophages, but how it contributes to the macrophage defence mechanisms remains unknown. Zebrafish have three copies of MPEG1, two of which (mpeg1 and mpeg1.2) are expressed in macrophages whereas the third could be a pseudogene. The mpeg1 and mpeg1.2 genes show differential regulation during infection of zebrafish embryos with the bacterial pathogens, Mycobacterium marinum and Salmonella typhimurium. While mpeg1 is down-regulated during infection with both pathogens, mpeg1.2 is infection inducible. Up-regulation of mpeg1.2 is partially dependent on the presence of functional Mpeg1, and requires the Toll-like receptor adaptor molecule MyD88 and transcription factor NF?B. Knockdown of mpeg1 alters the immune response to M. marinum infection and results in increased bacterial burden. In S. typhimurium infection, both mpeg1 and mpeg1.2 knockdown increase bacterial burdens, but mpeg1 morphants show an increased survival rate. The combined results of these two in vivo infection models support the anti-bacterial function of the Mpeg1 family and indicate that the intricate cross-regulation of the two mpeg1 copies aids the zebrafish host in combatting infection Overall design: Embryos were injected at the one cell stage with a morpholino targeting mpeg1, or with the standard control morpholino from GeneTools, or with a morpholino targeting ptpn6 (Kanwal et al., 2013, J. Immunol 190:1631-45) for comparison. Subsequently, at 24 hours post fertilisation (hpf) the morphants and their controls were manually dechorionated at 24 hpf and at 28 hpf they were infected by injecting 200 colony forming units of M. marinum Mma20 into the caudal vein, or mock-injected with PBS/2%PVP. After injections embryos were transferred into fresh egg water containing 0.003% 1-phenyl-2-thiourea (Sigma-Aldrich) to prevent melanisation and incubated for 4 days at 28°C. After the incubation period, infected and uninfected morphants, mutants and their controls were imaged and groups of 30 embryos were snap-frozen in liquid nitrogen and RNA was isolated for Illumina RNAseq analysis.
Publication Title
Macrophage-expressed perforins mpeg1 and mpeg1.2 have an anti-bacterial function in zebrafish.
Sample Metadata Fields
No sample metadata fields
View Samples- Danio rerio
- 11 Downloadable Samples
- IlluminaHiSeq2000
Description
MyD88 is an adaptor protein in Toll-like receptor and interleukin 1 receptor mediated signaling pathways that plays an essential role in activation of immune responses following pathogen recognition. We investigate that role in the zebrafish embryo model by using a zebrafish mutant line that contains a premature stop condon in the gene encoding MyD88, leading to a truncated protein that lacks domains important for its normal function. We infected these MyD88 mutants and wildtype individuals with Mycobacterium marinum to compare the resulting immune response by transcriptome profiling on total RNA isolated from single embryos. Autophagy regulator dram1 was identified as one of the MyD88-dependent genes. Overall design: This RNAseq analysis was used to determine the effect of a truncation of the MyD88 protein on the innate immune response of zebrafish embryos during infection with Mycobacterium marinum. Myd88 mutant and wild type embryos were derived by incrossing homozygous myd88 mutant parents (allele hu3568, van der Vaart et al., 2013, Disease models & mechanisms 6, 841-854) or their wildtype siblings. RNA was isolated from pools of 20 embryos at 4 days post infection (4 dpi). The following treatment groups were used: homozygous mutants mock-injected with PBS/2%PVP 4 dpi, (2) wildtype siblings mock-injected with PBS/2%PVP 4dpi, (3) M. marinum-infected homozygous mutants 4dpi, (4) M. marinum-infected wildtype siblings 4dpi. Embryos were grown at 28.5–30°C in egg water and manually dechorionated at 24 hours post fertilization (hpf). Subsequently, embryos were infected at 28 hpf by micro-injecting 200 colony forming units (CFU) of Mycobacterium marinum Mma20 bacteria into the caudal vein, or were mock-injected with buffer (PBS/2%PVP) as a control. After injections embryos were transferred into fresh egg water and incubated for 4 days at 28°C. After the incubation period, single embryos were snap-frozen in liquid nitrogen and RNA was isolated for RNAseq analysis.
Publication Title
Macrophage-expressed perforins mpeg1 and mpeg1.2 have an anti-bacterial function in zebrafish.
Sample Metadata Fields
No sample metadata fields
View Samples