Project description:An 8X15k oligonucleotide microarray was developed consisting of 2334 E. glacialis probes and 2166 Tursiops truncatus probes and used to measure the transcriptome level effects of right whale kidney fibroblast cells exposed to cadmium. Cells were exposed to three concentrations of cadmium chloride (CdCl2) for three exposure times. Cells exposed to 10-6M CdCl2 for 4 hours and 24 hours showed upregulated genes involved in protection from metal toxicity, oxidative stress, protein renaturation, apoptosis inhibition, and several regulators of cellular processes. Downregulated genes represented a suite of functions including cell proliferation, transcription regulation, actin polymerization, and stress fiber synthesis. The collection of differentially expressed genes in this study support proposed mechanisms of cadmium-induced apoptosis such as mitochondrial membrane potential collapse, reactive oxygen species (ROS) influx, and cell cycle arrest. The results confirm the right whale microarray as a reproducible tool in measuring differentiated gene expression and should be a valuable asset for transcriptome analysis of other baleen whales and potential health assessment protocols.
Project description:An 8X15k oligonucleotide microarray was developed consisting of 2334 E. glacialis probes and 2166 Tursiops truncatus probes and used to measure the transcriptome level effects of right whale kidney fibroblast cells exposed to cadmium. Cells were exposed to three concentrations of cadmium chloride (CdCl2) for three exposure times. Cells exposed to 10-6M CdCl2 for 4 hours and 24 hours showed upregulated genes involved in protection from metal toxicity, oxidative stress, protein renaturation, apoptosis inhibition, and several regulators of cellular processes. Downregulated genes represented a suite of functions including cell proliferation, transcription regulation, actin polymerization, and stress fiber synthesis. The collection of differentially expressed genes in this study support proposed mechanisms of cadmium-induced apoptosis such as mitochondrial membrane potential collapse, reactive oxygen species (ROS) influx, and cell cycle arrest. The results confirm the right whale microarray as a reproducible tool in measuring differentiated gene expression and should be a valuable asset for transcriptome analysis of other baleen whales and potential health assessment protocols. 35 Samples were analyzed representing 3 biological replicates for each treatment and corresponding controls except treatment 4hour_0.01uMCdCl2 had only two biological replicates due to spot intensity values. There are 3 spot replicates/probe on the array, along with Agilent control grid specific for 8X15K arrays, and 4 Karenia brevis sequences used as a negative control (K.brevis.chlorophyllA/Bbindingprotein, K.brevis.flavodoxin,K.brevis.photolyase,K.brevis.photosystemcoreprotein).
Project description:Basque whalers were active in the North Atlantic between the 11th and 18th. In the 16th and 17th c., they focused their attention to the coasts of Labrador and the Gulf of St. Lawrence, establishing shore stations from where they launched boats for chasing whales. On shore, they proceeded to render the blubber into oil by boiling it in large trypots. The residual blubber and remaining tissues were then used as fuel to boil more blubber. When the fire pit was full, the cinders were shoveled out, and the process began anew with new materials. Fist-sized lumps of cinder found at Bonne Espérance-4 (EiBk-61), a 16th Basque whaling site on the Quebec Lower North Shore, were sampled for proteomics analysis, to detect potential remains of whale tissues in the cinder. A simple protocol was employed for rapidly processing samples for nanoLC-MS/MS analysis. Out of 10 spots sampled on two lumps, materials recovered from one successfully yielded whale proteins. The study confirmed the presence of blubber and muscle remains (42 protein groups, including proteins such as myosin, myoglobin and hemoglobin) as well as baleen remains identified by cuticular keratins (12 protein groups, and up to 46 % protein coverage on type I keratin). Baleen, abundantly found at the site, was likely also used as fuel; based on keratin markers, the baleen belonged to a Balaenidae species. The processing of bowhead whale tissue was substantiated by specific peptides from myoglobin and obscurin, a result consistent with the targeting of bowhead whale by Basque whalers.
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.
Project description:Cadmium accumulation in kidney results in an irreversible chronic toxicity, but the underlying mechanisms are not clear. Transcriptomics assay may provide insight for the involved complex molecular networks. We used Affymetrix RTA arrays to detail the global gene expression profile of kidney tissues of SD rats with chronic exposure to Cadmium, and identified distinct classes of cadmium exposure related mRNA and pathways.
Project description:To understand the bioenergetics in cadmium-exposed macrophages, RNA sequencing (RNA-seq) was performed in lung macrophages isolated from vehicle- and cadmium-exposed mice.
Project description:Fish gills are not only the respiratory organ, but also essential for ion-regulation, acid-base control, detoxification, waste excretion and host defense. Multifactorial gill diseases are common in farmed Atlantic salmon, and still poorly understood. Understanding gill pathophysiology is of paramount importance, but the sacrifice of large numbers of experimental animals for this purpose should be avoided. Therefore, in vitro models, such as cell lines, are urgently required to replace fish trials. An Atlantic salmon gill epithelial cell line, ASG-10, was established at the Norwegian Veterinary institute in 2018. This cell line forms a monolayer expressing cytokeratin, e-cadherin and desmosomes, hallmarks of a functional epithelial barrier. To determine the value of ASG-10 for comparative studies of gill functions, the characterization of ASG-10 was taken one step further by performing functional assays and comparing the cell proteome and transcriptome with those of gills from juvenile freshwater Atlantic salmon. The ASG-10 cell line appear to be a homogenous cell line consisting of epithelial cells, which express tight junction proteins. We demonstrated that ASG-10 forms a barrier, both alone and in co-culture with the Atlantic salmon gill fibroblast cell line ASG- 13. ASG-10 cells can phagocytose and express several ATP-binding cassette transport proteins. Additionally, ASG-10 expresses genes involved in biotransformation of xenobiotics and immune responses. Taken together, this study provides an overview of functions that can be studied using ASG-10, which will be an important contribution to in vitro gill epithelial research of Atlantic salmon.
Project description:The association between kidney stone disease and renal fibrosis has been widely explored in recent years but its underlying mechanisms remain far from complete understanding. Using label-free quantitative proteomics (nanoLC-ESI-LTQ-Orbitrap MS/MS), this study identified 23 significantly altered secreted proteins from calcium oxalate monohydrate (COM)-exposed macrophages (COM-MP) compared with control macrophages (Ctrl-MP) secretome. Functional annotation and protein-protein interactions network analysis revealed that these altered secreted proteins were involved mainly in inflammatory response and fibroblast activation. BHK-21 renal fibroblasts treated with COM-MP secretome had more spindle-shaped morphology with greater spindle index. Immunofluorescence study and gelatin zymography revealed increased levels of fibroblast activation markers (α-smooth muscle actin and F-actin) and fibrotic factors (fibronectin and matrix metalloproteinase-9 and -2) in the COM-MP secretome-treated fibroblasts. Our findings indicate that proteins secreted from macrophages exposed to COM crystals induce renal fibroblast activation and may play important roles in renal fibrogenesis in kidney stone disease.