Transcriptome of the liver in 15-day-old C3H/HeJ mice
Ontology highlight
ABSTRACT: Male C3H/HeOuJ mice are susceptible to spontaneous liver neoplasms. We collected liver samples from young, tumour-free mice for control experiments. Liver tissue samples were snap frozen in liquid nitrogen and total RNA was extracted using the AllPrep 96 DNA/RNA Kit (Qiagen) according to the manufacturer’s instructions. Libraries were prepared using the TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Gold (Illumina) and sequenced in an Illumna HiSeq4000 to produce 150bp paired-end reads.
Project description:Male C3H/HeOuJ and CAST/EiJ mice were treated with diethylnitrosamine to induce liver tumours. We collected liver samples from untreated P15 mice for control experiments. Liver tissue samples were snap frozen in liquid nitrogen and total RNA was extracted using the AllPrep 96 DNA/RNA Kit (Qiagen) according to the manufacturer’s instructions. Libraries were prepared using the TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Gold (Illumina) and sequenced on an Illumna HiSeq4000 to produce 150bp paired-end reads.
Project description:Male C3H/HeOuJ mice are susceptible to spontaneous liver neoplasms. We aged a cohort of mice for up to 76 weeks and isolated liver tumours. Tumours were bisected: one half was snap frozen in liquid nitrogen for RNA extraction and the other half of the tumour was processed for histological examination. Total RNA was extracted using the AllPrep 96 DNA/RNA Kit (Qiagen) according to the manufacturer’s instructions. Libraries were prepared using the TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Gold (Illumina) and sequenced in an Illumina HiSeq4000 to produce 50bp paired-end reads.
Project description:This study aims to investigate the interactions of mutagenic lesions from diethylnitrosamine (DEN) treatment of mouse livers with such processes as replication, transcription, and interaction of DNA with proteins. Liver samples of 15-day old (P15) untreated C3H/HeOuJ mice were isolated and flash-frozen. ATAC-seq was performed to identify the regions of open chromatin and sites likely to be occupied by transcription factors.
Project description:This study aims to investigate the interactions of mutagenic lesions from diethylnitrosamine (DEN) treatment of mouse livers with such processes as replication, transcription, and interaction of DNA with proteins. Liver samples of 15-day old (P15) untreated C3H/HeOuJ mice were isolated and flash-frozen. ChIP-seq was performed to identify CTCF binding sites in livers of ten pooled individuals. The experiment was done with five biological replicates with a matched input library.
Project description:CTCF is a highly conserved and ubiquitously expressed protein involved in several fundamental processes such as fine-tuning gene expression, imprinting, X-chromosome inactivation and 3D chromatin organisation. To understand the impact of differences in the concentration of CTCF abundance on these processes, we exploit a CTCF hemizygous mouse model with a stable reduction in the concentration of this protein. We derived twelve independent primary lines of mouse embryonic fibroblasts (MEFs) from six wildtype and six CTCF-hemizygous mouse E13.5 embryos. Total RNA from each MEF line was purified using QIAzol Lysis Reagent (Qiagen); DNase treatment and removal was performed using the TURBO DNA-freeTM Kit (Ambion, Life Technologies). Libraries were prepared using the TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Gold (Illumina) and sequenced in an Illumna HiSeq4000 to produce 150bp paired-end reads. On the same MEF lines we have performed ChIPseq for CTCF, H3K4me3 and H3K27ac and HiC.
Project description:Alopecia areata (AA), a cell mediated autoimmune disease, is the second most common form of hair loss in humans. While the autoimmune disease is responsible for the underlying pathogenesis, the alopecia phenotype is ultimately due to hair shaft fragility and breakage associated with structural deficits. Quantitative trait genetic analyses using the C3H/HeJ mouse AA model identified cysteine-rich secretory protein 1 (Crisp1), a hair shaft structural protein, as a candidate gene within the major AA locus. Crisp1 transcripts in the skin at various times during disease development were barely detectable. In situ hybridization identified Crisp1 expression within the medulla of hair shafts from clinically normal strains of mice but not C3H/HeJ mice with AA. Follow-up work with 5-day-old C3H/HeJ mice with normal hair also had essentially no expression of Crisp1. Other non-inflammatory based follicular dystrophy mouse models with similar hair shaft abnormalities also have little or no Crisp1 expression. Shotgun proteomics, used to determine strain difference in hair proteins, confirmed that there was very little CRISP1 within normal C3H/HeJ mouse hair in comparison to 11 other strains. However, mutant mice with hair medulla defects also had undetectable levels of CRISP1 in their hair. Crisp1 null mice had normal skin, hair follicles, and hair shafts indicating that the lack of the CRISP1 protein does not translate directly into defects in the hair shaft or hair follicle. These results suggest that CRISP1 may be an important structural component of mouse hair and that its strain-specific dysregulation may indicate a predisposition to hair shaft disease such as AA.
Project description:Previously the effect of the pruritogens, such as histamine and chloroquine, was tested in 11 inbred mouse strains, and this study aimed to identify resistant and sensitive strains, consistent with the observation that underlies the large variability in human populations. In the present study, we used the low responder C3H/HeJ (C3H) and the more sensitive C57BL/6J (C57) strain to find out if resistance and sensitivity to develop pruritus is restricted to only histamine and chloroquine or extends to other known pruritogens as well. We tested five additional commonly known pruritogens. We established dose-response relationships by injecting four concentrations of the pruritogens in the range of 0.3, 1, 3, and ten-fold in the nuchal fold. Then we assessed the scratching behavior for 30 min after injection with an automated custom-designed device based on the bilateral implantation of mini-magnets in the hind paws and on single cages placed within a magnetic coil. We found that the resistance to pruritogens is a general phenotype of the C3H strain and extends to all pruritogens tested, including not only histamine and chloroquine, but also endothelin, trypsin, 5-HT (serotonin), the short peptide SLIGRL, and Lysophosphatidic acid (LPA). C57 was more sensitive to all pruritogens and, in contrast to C3H, dose-response relationships were evident for some of the pruritogens. In general, comparable peak scratch responses were observed for the 0.3-fold concentrations of the pruritogens in C57 whereas C3H required at least the ten-fold concentration and still displayed only between 5 and 33% of the scratch responses observed in C57 for the respective pruritogen. The general resistance to pruritogens and the low level of scratching behavior found in the C3H strain is an interesting trait and represents a model for the study of the heritability of itch. It is accompanied in C3H with a higher sensitivity in assays of nociception.
Project description:Alopecia areata (AA) is an autoimmune non-scarring hair loss disease. Recently, several reports have suggested that innate immune systems such as interferon-α (IFN-α)-producing plasmacytoid dendritic cells and NOD-like receptor family pyrin domain-containing protein 3 (NLRP3) inflammasomes play a role in the pathogenesis of AA. However, critical studies about their involvement in the initiation of AA have not yet been reported. Therefore, we investigated the expression of innate immune cytokines in serum and skin, and examined the effect of a selective NLRP3 inhibitor, MCC950, on AA in C3H/HeJ mice, induced by transferring cultured skin-draining lymph node cells. IFN-α production was upregulated in lesions of AA-affected mice, and interleukin-1β in serum and skin was highly expressed before onset as well as postonset. Furthermore, MCC950 treatment prevented AA development and promoted hair growth in AA mouse models by reducing NLRP3 signalling and Th1/Tc1 chemokines and cytokines in the skin. These results suggest that NLRP3 inflammasome contributes to AA onset and chronicity, and NLRP3 inhibitor may be a potential therapeutic agent for AA.
Project description:The exact global impact of leptospirosis is unknown due to inadequate surveillance systems in place in most low-income countries. In this study, we analyzed the differences in mouse inflammatory signatures involved in pathogenic versus non-pathogenic Leptospira recognition at 24h and 72h post infection. Injection of C3H-HeJ mice with non-pathogenic L. biflexa increased circulation of a few chemokines (5/21, 24%) without secretion of cytokines in blood that resulted in engagement of resident macrophages, dendritic cells, neutrophils and NK cells without engagement of T cells. In contrast, pathogenic L. interrogans induced circulation of a much higher panel of chemokines (18/21, 86%) and pro- and anti-inflammatory cytokines (11/19, 58%) in blood with a resulting signaling cascade leading to engagement of macrophages, dendritic cells, monocytes, NK cells and T cells without engagement of neutrophils. Although neutrophils do not appear to be engaged, a considerable number of chemokines that recruit other granulocytes such as eosinophils and basophils were also increased at 72h post infection with L. interrogans. Overall, the data suggest that prevention of dissemination of L. biflexa is associated with an early engagement of the innate immune response characterized by upregulation of a few chemokines that results in an efficacious phagocytic response without an overwhelming increase of pro-inflammatory cytokines. However, when macrophages fail to clear a pathogenic serovar such as L. interrogans, the adaptive response (T cells) is engaged to help out, but the resulting chemo-cytokine storm mediates a robust but non-resolving inflammatory response to pathogenic Leptospira that results in dissemination, kidney colonization, pathology and disease.