Differential gene expression in preimplantation embryos has been documented, but few focused studies have been done to compare differential expression in human embryos after embryonic genome activation and specifically how they relate to blastocyst development. We hypothesized that blastocyst stage embryos would differentially express genes in pathways important in cell division, mobilization, and processes important in embryo implantation including endometrial apposition, adhesion, and invasion. We analyzed gene expression in 6 preimplantation human embryos.
Differentially expressed genes in preimplantation human embryos: potential candidate genes for blastocyst formation and implantation.
Specimen part
View SamplesAims/hypothesis: While lipid deposition in skeletal muscle is considered to be involved in obesity-associated insulin resistance, neutral intramyocellular lipid (IMCL) accumulation per se does not necessarily induce insulin resistance. We previously demonstrated that overexpression of the lipid droplet coat protein perilipin 2 augments intramyocellular lipid content while improving insulin sensitivity. Another member of the perilipin family, perilipin 5 (PLIN5), is predominantly expressed in oxidative tissues like skeletal muscle. Here we investigated the effects of PLIN5 overexpression in comparison with effects of PLIN2 on skeletal muscle lipid levels, gene expression profiles and insulin sensitivity. Methods: Gene electroporation was used to overexpress PLIN5 in tibialis anterior muscle of rats fed a high fat diet. Eight days after electroporation, insulin-mediated glucose uptake in skeletal muscle was measured by means of a hyperinsulinemic euglycemic clamp. Electron microscopy, fluorescence microscopy and lipid extractions were performed to investigate IMCL accumulation. Gene expression profiles were obtained using microarrays. Results: TAG storage and lipid droplet size increased upon PLIN5 overexpression. Despite the higher IMCL content, insulin sensitivity was not impaired and DAG and acylcarnitine levels were unaffected. In contrast to the effects of PLIN2 overexpression, microarray data analysis revealed a gene expression profile favoring FA oxidation and improved mitochondrial function. Conclusions/interpretation: Both PLIN2 and PLIN5 increase neutral IMCL content without impeding insulin-mediated glucose uptake. As opposed to the effects of PLIN2 overexpression, overexpression of PLIN5 in skeletal muscle promoted expression of a cluster of genes under control of PPAR and PGC1 involved in FA catabolism and mitochondrial oxidation.
Overexpression of PLIN5 in skeletal muscle promotes oxidative gene expression and intramyocellular lipid content without compromising insulin sensitivity.
Sex, Age, Specimen part, Treatment
View SamplesThis work examines sarcoma formation within discrete subsets of KRAS(G12V)-expressing p16p19null myogenic and mesenchymal cells found normally in skeletal muscle. We show that prospectively isolated skeletal muscle precursor cells (SMPs) within the satellite cell pool can serve as cancer cells-of-origin for mouse rhabdomyosarcomas (soft tissue sarcomas with features of myogenic differentiation). Alternatively, non-myogenic progenitors (ScaPCs) induce sarcomas lacking myogenic differentiation markers.
Sarcomas induced in discrete subsets of prospectively isolated skeletal muscle cells.
Specimen part
View SamplesPurpose: Identify whole lung gene expression patterns in a house dust mite model of mild/moderate asthma Methods: Lung gene expression profiles of 10 week old BALB/c female mice were generated by ribosome-depleted, 150 nt, paired-end, stranded RNA-seq with Illumina HiSeq v4. Sequence reads that passed quality filters after trimming were analyzed with Sailfish-cir to identify linear RNAs and circular RNAs. Differential expression of linear RNAs was assessed with Deseq2 . QRT–PCR validation was performed using TaqMan and SYBR Green methods. Results: 100 million sequence reads per sample were mapped to the mouse genome (build mm10) using Sailfish-cir to identify linear and circular RNA transcripts. Pathway analysis of differentially expressed genes identified upregulation of gene sets for human asthma, mouse lung allergic inflammation, Muc5ac regulated genes and smooth muscle genes after allergic sensitization. Gene level exppression in each asthma-related pathway was reduced by the miR-145 antagonist. The miR-145 antagonist and several nontargeting oligos also upregulated interferon signaling pathways suggesting a general antiinflammatory effect of LNA/DNA oligos in the lung. Conclusions: Lung-directed delivery of LNA/DNA oligonucleotides with cationic lipid nanoparticles is an efffective means to prevent inflammatory gene expression in a house dust mite model of mild/moderate asthma. Overall design: Linear and circular RNA transcript expression was compared in whole lung tissue from unsensitized, house dust mite sensitzed, antimiR-145 treated treated mice
Nanoparticle Delivery of Anti-inflammatory LNA Oligonucleotides Prevents Airway Inflammation in a HDM Model of Asthma.
Age, Specimen part, Cell line, Treatment, Subject
View SamplesPurpose: Identify whole lung gene expression patterns modified by nanoparticle delivery of an antisense LNA/DNA oligonucleotide targeting mmu-miR145a-5p and nontargeting oligonucleotides Methods: Lung gene expression profiles of 10 week old BALB/c female mice were generated by polyA RNA-seq with Illumina HiSeq v4. Sequence reads that passed quality filters after timming were analyzed at the gene level with RNA STAR, featureCounts and Deseq2 . qRT–PCR validation was performed using TaqMan and SYBR Green methods. Results: 10-15 million sequence reads per sample were mapped to the mouse genome (build mm10). Pathway analysis of differentially expressed genes identified upregulation of gene sets for human asthma, mouse lung allergic inflammation, Muc5ac regulated genes and smooth muscle genes after allergic sensitization. Gene level exppression in each asthma-related pathway was reduced by the miR-145 antagonist. The miR-145 antagonist and several nontargeting oligos also upregulated interferon signaling pathways suggesting a general antiinflammatory effect of LNA/DNA oligos in the lung. Conclusions: Lung-directed delivery of LNA/DNA oligonucleotides with cationic lipid nanoparticles is an efffective means to prevent inflammatory gene expression in a house dust mite model of asthma Overall design: Lung gene expression in unsensitized, house dust mite sensitized, antimiR-145 treated and nontargeting oligonucleotide treated mice
Nanoparticle Delivery of Anti-inflammatory LNA Oligonucleotides Prevents Airway Inflammation in a HDM Model of Asthma.
Age, Specimen part, Cell line, Subject
View SamplesType 2 diabetes is characterized by excessive lipid storage in skeletal muscle. Excessive intramyocellular lipid storage exceeds intracellular needs and induces lipotoxic events ultimately contributing to the development of insulin resistance. Lipid droplet (LD)-coating proteins may control proper lipid storage in skeletal muscle. Perilipin 2 (PLIN2/ADRP) is one of the most abundantly expressed LD-coating proteins in skeletal muscle. Here we examined the role of PLIN2 in myocellular lipid handling and insulin sensitivity by investigating the effects of in vitro PLIN2 knockdown and in vitro and in vivo overexpression. PLIN2 knockdown decreased LD formation and triacylglycerol storage, marginally increased FA oxidation, and increased incorporation of palmitate into diacylglycerols and phospholipids. PLIN2 overexpression in vitro increased intramyocellular TAG storage paralleled with improved insulin sensitivity. In vivo muscle-specific PLIN2 overexpression resulted in increased LD accumulation and blunted the high-fat diet-induced increase of OXPHOS protein content. Diacylglycerol levels were unchanged, while ceramide levels were increased. Despite the increased intramyocellular lipid accumulation, PLIN2 overexpression improved skeletal muscle insulin sensitivity. We conclude that PLIN2 is essential for lipid storage in skeletal muscle by enhancing the partitioning of excess FAs towards triacylglycerol storage in LDs thereby blunting lipotoxicity-associated insulin resistance.
Perilipin 2 improves insulin sensitivity in skeletal muscle despite elevated intramuscular lipid levels.
Cell line
View SamplesBrown fat generates heat via the mitochondrial uncoupling protein UCP1, defending against hypothermia and obesity. Recent data suggest that there are two distinct types of brown fat: classical brown fat derived from a myf-5 cellular lineage and UCP1-positive cells that emerge in white fat from a non-myf-5 lineage. Here, we report the isolation of beige cells from murine white fat depots.
Beige adipocytes are a distinct type of thermogenic fat cell in mouse and human.
Cell line
View SamplesWe used microarrays to explore the expression profile from cells expressing wild type and UHRF1 S674A mutant. HeLa cells expressing UHRF1 WT and S674A mutant showed similar gene expression pattern without significant affecting the transcription of DNA repari genes.
A cell cycle-dependent BRCA1-UHRF1 cascade regulates DNA double-strand break repair pathway choice.
Specimen part
View SamplesPurpose: Osteoblast cells mature from a mesenchymal stem cell pool to become cells capable of forming bone matrix and mineralizing this matrix. The goal of this study was to characterize temporal changes in the transcriptome across osteoblast maturation, starting with committed mesenchymal stem cell/ early pre-osteoblast stage through to mature osteoblasts capable of matrix mineralization. Methods: Enriched populations of pre-osteoblast like cells were obtained from neonatal calvaria from C57BL/6J mice expressing CFP under the control of the Col3.6 promoter. These cells were placed into culture for 4 days, removed from culture and subjected FACS sorting based on the presence/absence of CFP expression. Cells expressing CFP were returned to culture, subjected to an osteoblast differentiation cocktail and RNA was collected at 2, 4, 6, 8, 10, 12, 14, 16 and 18 days post differentiation. Methods II: mRNA profiles for each time point were generated by next generation RNA sequencing, using an Illumina HiSeq 2000. Three technical replicates per samples were sequenced. The alignments for abundance estimation of transcripts was conducted using Bowtie version 0.12.9, using the NCBIm37 reference genome. Expression level per gene was calculated using RSEM version 1.2.0 with the parameters of --fragment-length-mean 280 and --fragment-length-sd 50, and the expression level for each sample was normalized relative to the per sample upper quartile. Overall design: Gene expression in calvarial osteoblasts from neonatal C57BL/6J-Col3.6 CFP mice at 9 time points post differentiation
Identification of 153 new loci associated with heel bone mineral density and functional involvement of GPC6 in osteoporosis.
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The mutation spectrum revealed by paired genome sequences from a lung cancer patient.
Specimen part
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