Chacterization of Day 1 and Day 5 Atlantic salmon adherent head kidney leukocytes (HKLs)
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PROVIDER: PRJNA725828 | ENA |
REPOSITORIES: ENA
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Project description:Transcriptional profiling of Atlantic salmon adherent HKLs that were cultured for 1 day (24 h) and cultured for 5 days (120 h). Our previous data (RNA-seq, morphology and phagocytosis) indicate that Day 1 cells are predominatly monocytes while Day 5 cells are predominatly macrophages.
2021-04-29 | GSE173493 | GEO
Project description:Atlantic salmon (average 550g) were used in this study. The fish were starved for 24 h and were sacrificed with an overdose of MS-222. Then head kidney (HK) and distal intestine (DI) samples were collected. Immune cells from HK and DI were isolated and cultured at 12°C in Leibovitz’s L-15 Medium (L-15). Both HK and DI leukocytes were allowed to adhere on a petri dish with 2 mL L-15+ for 2 days at 12°C. After removing the supernatant containing non-adherent cells, the petri dish with the adherent cells was placed on ice for 10 min, and the cells were detached by washing three times with 1.5 mL ice-cold PBS supplemented with 5 mM EDTA. The cells obtained were centrifuged (500 × g, 5 min, 4°C) and re-suspended in 1 mL L-15+. Then, the cells were counted using a portable cell counter. Total RNA was extracted from adherent cells from HK or DI (< 500,000 cells). To compare the expression of marker genes between adherent cells from HK and DI, 12 libraries were prepared (6 replicates for each). Transcriptomic analysis revealed that adherent intestinal cells (AICs) expressed genes associated with macrophages, T cells, and endothelial cells. In addition, AIC had higher expression of TNF and TGFβ-related genes that can be linked to macrophages and T cells. Heatmap analysis of selected genes indicated that the adherent cells from the two organs had apparently higher expression of macrophage-related markers.
2020-11-10 | GSE154142 | GEO
Project description:Head kidney (HK) and distal intestine (DI) from Atlantic salmon (average 550g) were collected. Immune cells from DI and HK were extracted and grown at 12°C in Leibovitz’s L-15 Medium (L-15). The isolated DI or HK leukocytes were allowed to adhere on a petri dish with 2 mL L-15+ for 2 days at 12°C. After removing the supernatant containing non-adherent cells, the adherent cells on petri dish were detached by washing three times with 1.5 mL ice-cold PBS supplemented with 5 mM EDTA. The cells were centrifuged (500 × g, 5 min, 4°C) and re-suspended with 2 mL L-15+. Total RNA was extracted from adherent cells from HK or DI (< 500,000 cells). To understand the miRNA expression profiles of adherent intestinal cells (AICs) and to compare them with those of adherent head kidney cells (AKCs), twelve libraries (6 for AIC and 6 for AKC) were prepared. Small RNA-sequencing revealed 37 differentially expressed miRNAs; 20 were up-regulated and 17 were down-regulated. Of them, 12 miRNAs in AIC were found to be involved in regulating macrophage functions.
2021-12-22 | GSE154147 | GEO
Project description:We report the miRNA profile of extracellular vesicles released from Atlantic salmon head kidney white blood cells that have been cultured for 1 day or 5 days.
2020-10-26 | GSE143360 | GEO
Project description:The Atlantic salmon (Salmo salar) is an economically important fish, both in aquaculture and in the wild. In vertebrates, macrophages are some of the first cell types to respond to pathogen infection and disease. While macrophage biology has been characterized in mammals, less is known in fish. Our previous work identified changes in the morphology, phagocytic ability, and miRNA profile of Atlantic salmon adherent head kidney leukocytes (HKLs) from predominantly "monocyte-like" at Day 1 of in vitro culture to predominantly "macrophage-like" at Day 5 of culture. Therefore, to further characterize these two cell populations, we examined the mRNA transcriptome profile in Day 1 and Day 5 HKLs using a 44K oligonucleotide microarray. Large changes in the transcriptome were revealed, including changes in the expression of macrophage and immune-related transcripts (e.g. csf1r, arg1, tnfa, mx2), lipid-related transcripts (e.g. fasn, dhcr7, fabp6), and transcription factors involved in macrophage differentiation and function (e.g. klf2, klf9, irf7, irf8, stat1). The in silico target prediction analysis of differentially expressed genes (DEGs) using miRNAs known to change expression in Day 5 HKLs, followed by gene pathway enrichment analysis, supported that these miRNAs may be involved in macrophage maturation by targeting specific DEGs. Elucidating how immune cells, such as macrophages, develop and function is a key step in understanding the Atlantic salmon immune system. Overall, the results indicate that, without the addition of exogenous factors, the adherent HKL cell population differentiates in vitro to become macrophage-like.
| S-EPMC8415484 | biostudies-literature
Project description:Adherent Intestinal Cells from Atlantic Salmon Show Phagocytic Ability and Express Macrophage Markers
| PRJNA645192 | ENA
Project description:Lufenuron is among the insecticides in use to combat salmon lice infestation in marine aquaculture. This benzoylurea drug acts as a chitin synthesis inhibitor, and prevent molting in the developing salmon lice larvae. In this work we aimed to study the molecular response to lufenuron exposure in Atlantic salmon given a standard oral dose. Juvenile salmon weighing about 40 g were treated with the drug for one week, and transcriptional responses were studied in liver tissue one (day 8) and seven days (day 14) after the treatment period. Exposure gave a transient inhibition of transcription in the liver at day 8 (2437 significant DEGs), followed by a weaker compensatory response at day 14 (169 significant DEGs). Many pathways associated with RNA metabolism were impacted at day 8. The sumoylation pathway was most strongly affected at day 8. Potential hepatic biomarkers for lufenuron exposure in fish could be phactr3, mst1r and slc47a1. In conclusion, this study shows that a 7-day standard dose of lufenuron downregulates transcription in liver tissue. The inhibitory effect on transcription was transitory, with a much weaker response one week after the medication period.
2023-02-01 | GSE199382 | GEO
Project description:We used RNA-seq to characterize the transcriptomic changes induced by type I IFN treatment and salmon alphavirus subtype 3 (SAV-3) infection in TO-cells, a macrophage/dendritic like cell-line derived from Atlantic salmon (Salmo salar L) head kidney leukocytes.
2014-12-31 | GSE64095 | GEO
Project description:MicroPoly(A)Pure kits (Ambion) were used to prepare mRNA from P. salmonis-infected and control Atlantic salmon head kidney. Head kidney mRNA samples were prepared from pooled (n = 10) infected or control kidney. Approximately 200 ng of infected or control head kidney mRNA was used as template in aRNA synthesis reactions. Two infected head kidney samples were pooled to yield 8.9 ug of aRNA, and a single control head kidney sample contained 8.4 ug of aRNA. Infected and control head kidney aRNA samples were visualized on agarose gels to check quality and quantity (data not shown). Except for the amounts of aRNA and reverse transcription (RT) primer used, head kidney target labeling reactions and microarray hybridizations were identical. Head kidney target syntheses used 2 ug of aRNA and 2 ug of random primers (Roche). RT primers were mixed with aRNA samples, and nuclease-free H2O (Invitrogen) was added to make 10 ul total volume. Samples were incubated at 70 degrees C for 4 min, then placed on ice. RT reactions included 50 mM Tris HCl pH 8.3, 75 mM KCl, 3 mM MgCl2, 10 mM DTT, 0.5 mM each dATP, dCTP, and dGTP, 0.2 mM dTTP, 0.05 mM Cy3-dUTP or Cy5-dUTP (Amersham), 40U RNAguard ribonuclease inhibitor (Amersham), and 400U Superscript II Rnase H- reverse transcriptase (Invitrogen). RT reactions were incubated at 42 degrees C for 2.5 h, followed by addition of 0.025U RNase A (Sigma) and 1.5U RNase H (New England BioLabs) and incubation at 37 degrees C for 30 min. Labeled targets were cleaned using the Qiaquick PCR purification kit (Qiagen) and precipitated, with 300 mM sodium acetate, 1 ul of 20 ug/ul glycogen (Invitrogen), and 2.5 volumes of 100% ethanol, at -20 degrees C overnight. Each labeled target was recovered by centrifugation (14,000 rpm, 4 degrees C, 1 h), washed in 70% ethanol, air-dried, and resuspended in 60 ul of hybridization buffer: 50% deionized formamide, 5X SSC, 0.1% SDS, 5 ul of 5 ug/ul oligo dT (5’T18[Q-N]3’), 5 ul of 2 ug/ul BSA (Pierce), and 2 ul of 10 ug/ul sonicated human placental DNA (Sigma). Targets were incubated at 96 degrees C for 3 min, and then at 65 degrees C until applied to microarrays. Microarrays were prepared for hybridization by washing 2 X 5 min in 0.1% SDS, washing 5 X 1 min in MilliQ H2O, immersing 3 min in 95 degrees C MilliQ H2O, and drying by centrifugation (2000 rpm, 5 min, in 50 ml conical tube). Microarray hybridizations were run in the dark under HybriSlips hybridization covers (Grace Biolabs) in slide hybridization chambers (Corning) submerged in a 45 degrees C water bath for 16 h. Coverslips were floated off in 45 degrees C (1X SSC, 0.2% SDS), and arrays were washed 1 X 10 min in 45 degrees C (1X SSC, 0.2% SDS), 3 X 4 min in room temperature (0.1X SSC, 0.1% SDS), and 4 X 4 min in room temperature 0.1X SSC. Slides were dried by centrifugation as before. Fluorescent images of hybridized arrays were acquired immediately at 10 um resolution using ScanArray Express (PerkinElmer). The Cy3 and Cy5 cyanine fluors were excited at 543 nm and 633 nm respectively, and the same laser power (90%) and photomultiplier tube (PMT) settings were used for all slides in a study (PMT 75 Cy3, PMT 63 or 64 Cy5 for head kidney study). Fluorescent intensity data were extracted from TIF images using Imagene 5.5 software (Biodiscovery). Keywords: other
2004-02-11 | GSE1031 | GEO