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SRP149081
Mus musculus
95 Downloadable Samples
NextSeq 500
Description
Purpose: Obesity is a global health issue. To investigate if protein and fat contents of the diets had effects on energy balance via the canonical hunger signaling pathways in the hypothalamus, RNAseq was performed on RNA extracted from the hypothalami of mice exposed to the different diets. A suggested mechanism by which animals may avoid obesity is by burning off excess energy via upregulation of white adipose tissue (WAT) browning. To investigate if protein and fat content of the diet had effects on energy balance via the browning related signaling pathways in the WAT, RNAseq was performed on RNA extracted from the subcutaneous WAT (sWAT) and epididymal WAT (eWAT) of mice exposed to the different diets. Methods: C57BL/6 male mice were used in this work. All mice were fed a standard diet with 10% fat and 20% protein (D12450B, Research Diets Ltd) for 2 weeks as the baseline period. Following 2 weeks of baseline monitoring (at age 12 weeks), all mice were randomly allocated to different groups and switched to the experimental diets for 12 weeks. After 12 weeks all mice were sacrificed and dissected. Methods: In total, mice were fed on 4 diet series, each series consisting of 6 different diets (total = 24 diets). In the first two series (Series 1: D14071601–D14071606 and series 2: D14071607 – D14071612) we fixed the level of fat by energy, and varied the protein content. The protein source was casein. The balance was made up by carbohydrate (roughly equal mix of corn starch and maltodextrose). The source of fat was a mix of cocoa butter, coconut oil, menhaden oil, palm oil and sunflower oil. This mix was designed to match the balance of saturated, mono-unsaturated and polyunsaturated fats (ratio 47.5: 36.8: 15.8) and the n-6: n-3 ratio (14.7: 1) in the typical western diet. The proportions of the different fat constituents and hence fatty acid distributions did not change as the total fat content changed. Sucrose and cellulose were both fixed 5% by energy and weight respectively, and all diets were supplemented with a standard vitamin and mineral mix. In the second two series of diets (series 3: D14071613 – D14071618 and series 4: D14071619 – D14071624) we fixed the level of protein by energy and then allowed the fat content to vary. In these diets the sucrose, cellulose and vitamin and mineral contents were the same as the diets in series 1 and 2. All these diets can be ordered direct from research diets (www.researchdiets.com) using the diet codes provided. Methods: From each diet group, the hypothalami of 8/20 individuals were collected. The left halves of two, and the right halves of another two, were pooled together as one sample, and the same was performed with the other 4 hypothalami, resulting in each diet group having 2 pooled samples of 4 hypothalami (n = 48 samples in total across 24 diets). From each diet group, the sWAT and eWAT of 12/20 individuals were also collected. A small piece from each of six sWAT collections were pooled together as one sample, and the same was performed with the other six eWAT collections. In this way each diet group had one pooled sWAT sample and one pooled eWAT sample (also n = 48 across 24 diets). Methods: The total RNA of the hypothalamus and WAT was isolated using the RNeasy Mini Kit (QIAGEN, 74104) according to manufacturer''s protocol. All sequencing was carried out using the Illumina NextSeq 500 sequencer. RNA fragments were sequenced by 75 bp long reads from paired ends (PE 2 x 75 bp, 150 bp per fragment). Quality control checks for raw data FASTQ files were done by using FASTQC (a quality control tool for high throughput sequence data; http://www.bioinformatics.babraham.ac.uk/projects/fastqc/). Paired-end reads were mapped to the Mus musculus genome (GRCm38) using Bowtie 2-2.1.0, TopHat-2.0.10, and Samtools-0.1.19; uniquely mapped reads for each gene were counted against the GTF file of GRCm38 provided by Ensembl (release 83) using HTSeq-0.6.1p1 using the strand = reverse; after read count data were obtained from the TopHat-HTSeq pipeline, counts per million (CPM) value for each gene was calculated by using the R package 'edgeR' (version 3.12.0, R version 3.2.2) to normalize the count data by the size of the library of each sample. Genes with the CPM value = 1 in at least one of the 24 diets group were retained (Anders et al., 2013). Generalized Linear Modelling (GLM) was applied by R (version 3.2.2). The GLM model used here was: ~p+f+p:f, which regresses gene expression (CPM) against the protein (p) and fat contents (f) of diets, as well as their interaction (p:f). However, when the effect of the interaction was not significant (p value = 0.05), the interaction term was dropped and a revised model (~p+f) was utilized. Results: With TopHat-HTSeq pipeline, reads of each sample were mapped to 46,078 genes. In hypothalamus there were 15,371 genes with the counts per million (CPM) value = 1 in at least one of the 24 diets group; in white adipose tissue there were 18,202 genes with the CPM value = 1 in at least one of the 24 diets group. No major changes in hypothalamic gene expression levels were found in relation to different dietary protein levels at fixed fat contents, however hypothalamic gene expression showed increase in expression of genes in reward pathways in relation to dietary fat, while Agrp and Npy were both downregulated in relation to dietary fat levels. WAT gene expression showed decrease in expression of general thermogenic related genes and WAT browning related genes in relation to both dietary protein and dietary fat, while Tgfb1, Pdk4 and Fgf1 were all upregulated in relation to dietary fat levels. Conclusions: Significant positive associations were evident between the fat levels of the diet and the main hedonic signaling systems linked to food intake. Significant negative associations were found between both protein and fat levels of the diet and WAT browning or general thermogenic signalings linked to energy expenditure. Overall design: In total 96 samples are analyzed. From each diet group, the hypothalami of 8/20 individuals were collected. The left halves of two, and the right halves of another two, were pooled together as one sample, and the same was performed with the other 4 hypothalami, resulting in each diet group having 2 pooled samples of 4 hypothalami (n = 48 samples in total across 24 diets). From each diet group, the sWAT and eWAT of 12/20 individuals were also collected. A small piece from each of six sWAT collections were pooled together as one sample, and the same was performed with the other six eWAT collections. In this way each diet group had one pooled sWAT sample and one pooled eWAT sample (also n = 48 across 24 diets)
Publication Title
Dietary Fat, but Not Protein or Carbohydrate, Regulates Energy Intake and Causes Adiposity in Mice.
Sample Metadata Fields
Sex, Age, Specimen part, Cell line, Subject
View Samples- Mus musculus
- 18 Downloadable Samples
Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
In the present study, we hypothesized that C/EBPa (CCAAT/enhancer-binding protein alpha) plays a role in cell regeneration in response to bronchiolar epithelial cell injury. C/EBPa mediated ciliated cell regeneration after naphthalene bronchiolar epithelial cell injury in vivo. Furthermore, we demonstrated that C/EBPa regulates protease/anti-protease balance after lung injury, and intratracheal treatment with anti-protease (BPTI) restored ciliated cell regeneration after naphthalene injury in CebpaD/D mice.
Publication Title
CCAAT/enhancer binding protein-α regulates the protease/antiprotease balance required for bronchiolar epithelium regeneration.
Sample Metadata Fields
Specimen part, Treatment
View Samples- Mus musculus
- 6 Downloadable Samples
- Affymetrix Mouse Genome 430 2.0 Array (mouse4302)
Description
Deletion of Stat3 induced genes influencing protein metabolism, transport, chemotaxis and apoptosis and decreased the expression of genes mediating lipid synthesis and metabolism. Srebf1 and 2, key regulators of fatty acid and steroid biosynthesis, were decreased in Stat3D/D mice. Stat3 influenced both pro- and anti-apoptotic pathways, regulating and maintaining the balance between a subset of pro- and anti-apoptotic genes that determine cell death or survival. Akt, a known target of Stat3, participates in many Stat3 mediated pathways including Jak-Stat signaling, apoptosis, the MAPK signaling, cholesterol and fatty acid biosynthesis. Deletion of Stat3 from type II epithelial cells altered the expression of genes regulating diverse cellular processes, including cell growth and apoptosis, lipid biosynthesis and metabolism. Stat3 regulates cell formation through a complex regulatory network that likely enhances alveolar epithelial cell survival and surfactant/lipid synthesis, necessary for the protection of the lung during injury.
Publication Title
Gene expression and biological processes influenced by deletion of Stat3 in pulmonary type II epithelial cells.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 18 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
We have previously demonstrated that deletion of the Cebpa gene in the developing fetal mouse lung caused death soon after birth from the failure of lung maturation. Many of the transcriptional pathways regulating morphogenesis of the fetal lung are induced postnatally and mediate repair of the injured lung. We hypothesized that C/EBPa plays a role in protection of the alveolar epithelium following hyperoxia injury of the mature lung. Transgenic Cebpa/ mice in which Cebpa was conditionally deleted from Clara cells (from early gestation) and type II cells (from near-term) were developed. Cebpa/ mice grow normally without any pulmonary abnormalities. Cebpa/ mice were highly susceptible to hyperoxia. Cebpa/ mice died within 4d after hyperoxia associated with severe lung inflammation and altered surfactant components at a time when all control mice survived. Microarrays were analyzed on isolated type II cells at an early stage (24h) of hyperoxia exposure to detect the primary genes influenced by deletion of Cebpa. The associated network analysis revealed the reduced expression of key genes related to surfactant lipid and protein homeostasis, such as Srebf, Scap, Lpcat1, Abca3, Sftpb, and Napsa. Genes for the cell signaling, immune response, and protective antioxidants, including GSH and Vnn-1,3, were decreased in the Cebpa/ mice lung. C/EBPa did not play a critical role in postnatal pulmonary function under normal conditions. In contrast, in the absence of C/EBPa, exposure to hyperoxia caused respiratory failure, supporting the concept that C/EBPa plays an important role in enhancing epithelial cell survival, surfactant lipid homeostasis, and maturation of SP-B from pro-SP-B.
Publication Title
C/EBP{alpha} is required for pulmonary cytoprotection during hyperoxia.
Sample Metadata Fields
Specimen part
View Samples- Mus musculus, Homo sapiens
- 8 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
This SuperSeries is composed of the SubSeries listed below.
Publication Title
Hippo/Yap signaling controls epithelial progenitor cell proliferation and differentiation in the embryonic and adult lung.
Sample Metadata Fields
Specimen part
View Samples- Homo sapiens
- 4 Downloadable Samples
- IlluminaHiSeq2000
Description
Primary human bronchial epithelial cells were transduced with control or hYAP(S127A) lentivirus in sphere forming conditions. Bronchospheres were harvested on day 18-20 for RNAseq analysis Overall design: Passage 1 Primary HBECs from 2 independent donors were transduced with control or hYAP lentivirus. 48 hours post infection, cells were plated on transwell inserts in a 50-50 mixture of ALI medium-Cultrex BME reduced growth factor (RGF) to form spheres. Well differentiated bronchospheres were harvested for RNA-seq analysis on day 18-20 by combining 3 wells of each group for each donor.
Publication Title
Hippo/Yap signaling controls epithelial progenitor cell proliferation and differentiation in the embryonic and adult lung.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 2 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
ShhCre;Mst1/2flx/flx (Mst1/2 D/D) mice were generated to conditionally delete Mst1 and Mst2 from epithelial progenitors during lung morphogenesis. Lungs from E18.5 control and Mst1/2 D/D mice were mechanically and enzymatically dissociated to generate single cell suspension. Epcam(+) cells were isolated using magnetic microbeads.
Publication Title
Hippo/Yap signaling controls epithelial progenitor cell proliferation and differentiation in the embryonic and adult lung.
Sample Metadata Fields
Specimen part
View Samples- Mus musculus
- 2 Downloadable Samples
- Illumina HiSeq 2000
Description
Mst1 and Mst2 were conditionally deleted from non-ciliated bronchiolar epithelial cells in the mature lung. Bronchiolar epithelial cells from control and Mst1/2 deleted mice were isolated by cell sorting and used for RNA-seq analysis. Overall design: Scgb1a1-rtTA/tetO-Cre/Mst1;2-flx/flx (Mst1/2 D/D) mice were generated to conditionally delete Mst1 and Mst2 from non-ciliated, secretory bronchiolar epithelial cells. Adult mice were maintained on doxycycline food for 16 days to induce deletion of Mst1/2. Lin-/CD326+/CD24-intermediate cells were isolated by fluorescence cell sorting to enrich for the targeted airway epithelial cells. mRNA isolated from Lin-/CD326+/CD24-intermediate cells from control and Mst1/2 D/D mice was pooled and analyzed by RNA-seq to identify transcriptional changes following deletion of Mst1 and Mst2 from mature lung bronchiolar epithelial cells.
Publication Title
Hippo/Yap signaling controls epithelial progenitor cell proliferation and differentiation in the embryonic and adult lung.
Sample Metadata Fields
No sample metadata fields
View Samples- Mus musculus
- 6 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
Kruppel-like transcription factor 5 (Klf5) is expressed during late embryogenesis in the forming murine bladder urothelium. Targeted disruption of the Klf5flox alleles by the ShhGfpCre transgene resulted in failure of the bladder urothelium to mature accompanied by hydronephrosis, hydroureter, and vesicoureteric reflux in all E18.5 fetuses. The bladder urothelium did not stratify nor did it express terminal differentiation markers characteristic of basal, intermediate, and umbrella cells including keratins 20, 14, and 5, and uroplakins. At E18.5, an ectopic alpha smooth muscle actin positive layer of cells was identified subjacent to the undifferentiated Klf5-deficient urothelium. The effects of Klf5 deficiency were unique to the urothelium since maturation of the epithelium comprising the bladder neck and urethra were unaffected by the lack of KLF5. mRNA microarray analysis of whole E14.5 control and Klf5 deficient bladders identified Ppar-gamma and Grhl3 as putative downstream intermediary transcription factors that regulate urothelial maturation. Transient transfection assays demonstrated that KLF5 regulated expression of the mGrhl3 promoter. These observations show that alterations in maturation of the bladder urothelium alone are sufficient to induce bladder dysfunction leading to prenatal hydronephrosis.
Publication Title
Kruppel-like factor 5 is required for formation and differentiation of the bladder urothelium.
Sample Metadata Fields
Specimen part
View Samples- Mus musculus
- 24 Downloadable Samples
- Affymetrix Mouse Gene 1.0 ST Array (mogene10st)
Description
In this study, we studied the genomic responses of the Insig and Scap deletion from perinatal lung.
Publication Title
Epithelial SCAP/INSIG/SREBP signaling regulates multiple biological processes during perinatal lung maturation.
Sample Metadata Fields
Specimen part
View Samples