LCM-ATACseq on human lung macrophages
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ABSTRACT:
PROVIDER: EGAS00001006167 | EGA |
REPOSITORIES: EGA
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Project description:In the reported study, we employed Laser Capture Microdissection (LCM) for the transcriptome profiling of lung macrophages cells populations as a function of location within the healthy tissue. In detail, macrophage mini-bulks (100 cells each) were collected by LCM from 4 healthy human donors in 5 different locations of the airways (a total of 20 biopsies), including parenchyma (L1 – lower left lobe (LLL); L6 – 80% distance from LLL tip), trachea (L2), bronchi (L3 – 1st/2nd generation; L5 – 3rd/4th generation) and processed for RNA-seq.
| EGAS00001006168 | EGA
Project description:In the reported study, we employed Laser Capture Microdissection (LCM) for the transcriptome profiling of lung macrophages cells populations as a function of location within the healthy tissue. In detail, macrophage mini-bulks (100 cells each) were collected by LCM from 4 healthy human donors in 5 different locations of the airways (a total of 20 biopsies), including parenchyma (L1 – lower left lobe (LLL); L6 – 80% distance from LLL tip), trachea (L2), bronchi (L3 – 1st/2nd generation; L5 – 3rd/4th generation) and processed for RNA-seq.
| EGAD00001008694 | EGA
Project description:In this study we employed Laser Capture Microdissection (LCM) for the multimodal profiling of lung macrophages cell populations as a function of location within the healthy tissue. In detail, macrophage mini-bulks (~100 cells) were collected from 4 healthy human donors in 5 different locations of the airways (a total of 20 biopsies), including parenchyma (L1 – lower left lobe (LLL); L6 – 80% distance from LLL tip), trachea (L2), bronchi (L3 – 1st/2nd generation; L5 – 3rd/4th generation), and processed for ATAC-seq.
| EGAD00001008693 | EGA
Project description:The inflammatory functions of the cytokine tumor necrosis factor (TNF) rely on its ability to induce cytokine production and to induce cell death. Caspase dependent and independent pathways – apoptosis and necroptosis – respectively, regulate immunogenicity by the release of distinct sets of cellular proteins. To obtain an unbiased, systems-level understanding of this important process, we here applied mass spectrometry-based proteomics to dissect protein release during apoptosis and necroptosis. We report hundreds of proteins released from human myeloid cells in time-course experiments. Both cell death types induce receptor shedding, but only apoptotic cells released nucleosome components. Conversely, necroptotic cells release lysosomal components by activating lysosomal exocytosis at early stages of necroptosis- induced membrane permeabilisation and show reduced release of conventionally secreted cytokines.
2020-07-18 | PXD014966 | Pride
Project description:We performed transcriptional and metabolic profiling for yeast at early stages of senescence (4th, 7th and 11th generation), that is, for populations in which most cells are still alive. Transcriptional profiles showed up- and down-regulation for about 20% of the genes profiled after the first 4 generations, few further changes by the 7th generation, and an additional 12% of the genes were up- and down-regulated after 11 generations. Pathway analysis revealed that these 11th generation cells had accumulated transcripts coding for enzymes involved in sugar metabolism, the tricarboxylic acid (TCA) cycle and amino acid degradation, and showed decreased levels of mRNAs coding for enzymes involved in amino acid biosynthetic pathways. Stationary phase-induced genes were highly expressed after 11 generations even though the growth medium contained adequate levels of nutrients, indicating deterioration of the nutrient sensing and/or signaling pathways by the 11th generation. These changes are presumably early indications of replicative senescence. Cells of yeast strain BY4742 were labeled with biotin and cultivated in YPD medium at 30M-BM-0C. The labeled cells at early stages of senescence (4th, 7th and 11th generation) were isolated using Dynabeads Biotin Binder (Invitrogen, Carlsbad, CA, USA). Total RNA samples were prepared from the senescent cells. Antisense RNA was synthesized and labeled with Cy3 for the 4th, 7th, and 11th generation and with Cy5 for the 1st generation. Each mixture of Cy3- and Cy5-labeled antisense RNA was hybridized on a 3D-Gene Yeast Oligo Chip 6K (Toray, Kanagawa, Japan). The hybridized array was scanned with a 3D-Gene Scanner 3000 (Toray).
2014-07-27 | E-GEOD-59797 | biostudies-arrayexpress
Project description:Spatial tissue proteomics integrating whole-slide imaging, laser microdissection and ultrasensitive mass spectrometry is a powerful approach to link cellular phenotypes to functional proteome states in (patho)physiology. To be applicable to large patient cohorts and low sample input amounts, including single-cell applications, loss-minimized and streamlined end-to-end workflows are key. We here introduce an automated sample preparation protocol for laser microdissected samples utilizing the cellenONE® robotic system, which has the capacity to process 192 samples in three hours. Following laser microdissection collection directly into the proteoCHIP LF 48 or EVO 96 chip, our optimized protocol facilitates lysis, formalin de-crosslinking and tryptic digest of low-input archival tissue samples. The seamless integration with the Evosep ONE LC system by centrifugation allows ‘on-the-fly’ sample clean-up, particularly pertinent for laser microdissected workflows. We validate our method in human tonsil archival tissue, where we profile proteomes of spatially-defined B-cell, T-cell and epithelial microregions of 4,000 µm2 to a depth of ~2,000 proteins and with high cell type specificity. We finally provide detailed equipment templates and experimental guidelines for broad accessibility.
2024-03-22 | PXD047762 | Pride
Project description:Expression and differential expression analysis of milk samples from healthy and diseased diary cows. Diseases were grouped by their occurrence in the mammary gland or extra-mammary. Furthermore, the diseases were classified by their severity. All cows were examined thoroughly by the dairy herd manager, trained staff, or a veterinarian. Expression and differential expression was assessed by using the Affymetrix Bovine Genome Array (GPL2112). Control animals (2-4 years old, 1st to 3th lactation, one animal 4th and one 8th lactation) showed no clinical signs of disease and had no abnormalities in the udder or milk. Their somatic cell count (SCC) was less than 100,000 cells/ml milk. Most of the control samples were taken during early lactation (10-100 days post-partum, dpp). Diseased cows were in their 1st to 8th lactation within 10-220 dpp.
2018-06-29 | GSE93082 | GEO
Project description:Transcriptional analysis of sex-regulated genes from male and female dsxM-^VeGFP sexed larvae (1st instar, late 2nd /early 3rd instar and 4th instar), pupae and virgin non-blood-fed 3 day old adult A. gambiae mosquitoes
2011-05-01 | E-MEXP-3093 | biostudies-arrayexpress
Project description:Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study were to compare signals from competent and abnormal human embryos impacted differently on the expression of endometrial receptivity genes in mouse uteri using transcriptome profiling (RNA-seq). Methods: Immature female (25 d C57BL/6) mice were given a hormone treatment of a single dose 1 mg progesterone and 10 mg/kg/day ò-estradiol for a total of 3d to prime the uterus for embryo transfer. The uterine horns of control and study mice were injected with an equal volume (50 ül) of either unconditioned embryo culture medium (ECM), serving as controls, or pooled conditioned media from competent (n = 9) or arresting embryos (n = 18). Uterine mRNA profiles of 25-day-old wild-type (WT) were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000 platform. The sequence reads that passed quality filters were analyzed with the following methods: Bowtie Alignment followed by TopHat (splice juntions mapper) and Cufflinks (transcript abundance). qRTâÂÂPCR validation was performed using SYBR Green assays. Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (build mm9) and identified 16,014 transcripts in the uteri WT and NrlâÂÂ/â mice with BWA workflow and 34,115 transcripts with TopHat workflow. RNA-seq data confirmed stable expression of 25 known housekeeping genes, and 12 of these were validated with qRTâÂÂPCR. RNA-seq data had a linear relationship with qRTâÂÂPCR for more than four orders of magnitude and a goodness of fit (R2) of 0.8798. Approximately 10% of the transcripts showed differential expression between the WT and NrlâÂÂ/â retina, with a fold change âÂÂ¥1.5 and p value <0.05. Altered expression of 25 genes was confirmed with qRTâÂÂPCR, demonstrating the high degree of sensitivity of the RNA-seq method. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized genes that may contribute to retinal function. Data analysis with BWA and TopHat workflows revealed a significant overlap yet provided complementary insights in transcriptome profiling. Conclusions: Our study represents the first detailed analysis of maternal uterine reponse to signals from competent and abnormal human embryos, with biological replicates, generated by RNA-seq technology. Supernatant from competent and abnormal human embryos were flushed into the uterine horns of 25d old C57BL/6 female mice. Uterine mRNA profiles of 21-day old mice were generated by deep sequencing, in triplicate, using Illumina HiSeq 2000 platform (HiSeq).
2014-02-11 | E-GEOD-47019 | biostudies-arrayexpress
Project description:The initial site of smoking-induced lung disease is the small airway epithelium, which is difficult and time consuming to sample by fiberoptic bronchoscopy. We developed a rapid, office-based procedure to obtain trachea epithelium without conscious sedation from healthy nonsmokers (n=26) and healthy smokers (n=19, 27 ± 15 pack-yr). Gene expression differences [fold-change >1.5, p< 0.01, Benjamini-Hochberg correction] were assessed with Affymetrix microarrays. 1,057 probe sets were differentially expressed in healthy smokers vs nonsmokers, representing >500 genes. Trachea gene expression was compared to an independent group of small airway epithelial samples (n=23 healthy nonsmokers, n=19 healthy smokers, 25 ± 12 pack-yr). The trachea epithelium is more sensitive to smoking, responding with 3-fold more differentially-expressed genes than small airway epithelium. The trachea transcriptome paralleled the small airway epithelium, with 156 of 167 (93%) genes that are significantly up- and down-regulated by smoking in the small airway epithelium showing similar direction and magnitude of response to smoking in the trachea. Trachea epithelium can be obtained without conscious sedation, representing a less invasive surrogate “canary” for smoking-induced changes in the small airway epithelium. This should prove useful in epidemiologic studies correlating gene expression with clinical outcome in assessing smoking-induced lung disease.
2009-08-27 | GSE13933 | GEO