RNA-seq for U937 cells with or without 3 day differentiation with PMA and recovery
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE107566 | GEO | 2018/08/03
REPOSITORIES: GEO
Similar Datasets
Project description:RNA-seq for U937 cells with or without 3 day differentiation with PMA and recovery
| PRJNA420647 | ENA
Project description:Complete, reproducible extraction of protein material is essential for comprehensive and unbiased proteome analyses. A current gold standard is single-pot, solid-phase-enhanced sample preparation (SP3), in which organic solvent and magnetic beads are used to denature and capture proteins, with subsequently washes allowing contaminant removal. However, SP3 is dependent on effective protein immobilisation onto beads, risks losses during wash steps, and experiences a drop-off in protein recovery at higher protein inputs. Magnetic beads may also contaminate samples and instruments, and become costly for larger scale protein preparations. Here, we propose solvent precipitation SP3 (SP4) as an alternative to SP3, omitting magnetic beads and employing brief centrifugation—either with or without low-cost inert glass beads—as the means of aggregated protein capture. SP4 recovered equivalent or greater protein yields for 1 — 5000 µg preparations and improved reproducibility (median protein R2 0.99 (SP4) vs. 0.97 (SP3)). Deep proteome profiling (n=9,076) also demonstrated improved recovery by SP4 and a significant enrichment of membrane and low-solubility proteins vs. SP3. The effectiveness of SP4 was verified in three other labs, each confirming equivalent or improved proteome characterisation over SP3. This work suggests that protein precipitation is the primary mechanism of SP3, and reliance on magnetic beads presents protein losses, especially at higher concentrations and amongst hydrophobic proteins. SP4 represents an efficient and effective alternative to SP3, provides the option to omit beads entirely, and offers virtually unlimited scalability of input and volume—all whilst retaining the speed and universality of SP3.
2022-08-03 | PXD028768 | Pride
Project description:Complete, reproducible extraction of protein material is essential for comprehensive and unbiased proteome analyses. A current gold standard is single-pot, solid-phase-enhanced sample preparation (SP3), in which organic solvent and magnetic beads are used to denature and capture proteins, with subsequently washes allowing contaminant removal. However, SP3 is dependent on effective protein immobilisation onto beads, risks losses during wash steps, and experiences a drop-off in protein recovery at higher protein inputs. Magnetic beads may also contaminate samples and instruments, and become costly for larger scale protein preparations. Here, we propose solvent precipitation SP3 (SP4) as an alternative to SP3, omitting magnetic beads and employing brief centrifugation—either with or without low-cost inert glass beads—as the means of aggregated protein capture. SP4 recovered equivalent or greater protein yields for 1 — 5000 µg preparations and improved reproducibility (median protein R2 0.99 (SP4) vs. 0.97 (SP3)). Deep proteome profiling (n=9,076) also demonstrated improved recovery by SP4 and a significant enrichment of membrane and low-solubility proteins vs. SP3. The effectiveness of SP4 was verified in three other labs, each confirming equivalent or improved proteome characterisation over SP3. This work suggests that protein precipitation is the primary mechanism of SP3, and reliance on magnetic beads presents protein losses, especially at higher concentrations and amongst hydrophobic proteins. SP4 represents an efficient and effective alternative to SP3, provides the option to omit beads entirely, and offers virtually unlimited scalability of input and volume—all whilst retaining the speed and universality of SP3.
2022-08-04 | PXD028736 | Pride
Project description:Complete, reproducible extraction of protein material is essential for comprehensive and unbiased proteome analyses. A current gold standard is single-pot, solid-phase-enhanced sample preparation (SP3), in which organic solvent and magnetic beads are used to denature and capture protein aggregates, with subsequent washes removing contaminants. However, SP3 is dependent on effective protein immobilisation onto beads, risks losses during wash steps, and experiences a drop-off in recovery at higher protein inputs. Magnetic beads may also contaminate samples and instruments, and become costly for larger-scale protein preparations. Here, we propose solvent precipitation SP3 (SP4) as an alternative to SP3, omitting magnetic beads and employing brief (5-minute) centrifugation—either with or without low-cost inert glass beads—as a means of acetonitrile-induced aggregated protein capture. SP4 recovered equivalent or greater protein yields for 1–5000µg preparations and improved reproducibility (median protein R2 0.99 (SP4) vs. 0.97 (SP3)). Deep proteome profiling revealed SP4 yielded greater recovery of low-solubility and transmembrane proteins than SP3, benefits to aggregating protein using 80% vs. 50% organic solvent, and equivalent recovery by SP4 and S-Trap. SP4 was verified in three other labs, across eight sample types, and five lysis buffers—all confirming equivalent or improved proteome characterisation vs. SP3. With near-identical recovery, this work further illustrates protein precipitation as the primary mechanism of SP3 protein clean-up, and identifies that magnetic capture risks losses, especially at higher protein concentrations and amongst more hydrophobic proteins. SP4 offers a minimalistic approach to protein clean-up that provides cost-effective input scalability, the option to omit beads entirely, and suggests important considerations for SP3 applications—all whilst retaining the speed and compatibility of SP3.
2022-08-01 | PXD032095 | Pride
Project description:Sequencing data related to the manuscript entitled, "CD22 blockade restores homeostatic microglial phagocytosis in the aging brain."
2019-03-01 | GSE127543 | GEO
Project description:Sequencing data related to the manuscript entitled, "CD22 blockade restores homeostatic microglial phagocytosis in the aging brain."
2019-03-01 | GSE127542 | GEO
Project description:Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes rash, fever and severe polyarthritis that can last for years in humans. Murine models display inflammation and macrophage infiltration only in the adjacent tissues at the site of inoculation, showing no signs of systemic polyarthritis. Monocyte-derived macrophages are one cell type suspected to contribute to a systemic CHIKV infection. The purpose of this study was to analyze differences in CHIKV infection in two different cell lines, human U937 and murine RAW264.7 monocyte derived macrophages. PMA-differentiated U937 and RAW264.7 macrophages were infected with CHIKV, and infectious virus production was measured by plaque assay and by reverse transcriptase quantitative PCR at various time points. Secreted cytokines in the supernatants were measured using cytometric bead arrays. Cytokine mRNA levels were also measured to supplement expression data. Here we show that CHIKV replicates more efficiently in human macrophages compared to murine macrophages. In addition, infected human macrophages produced around 10-fold higher levels of infectious virus when compared to murine macrophages. Cytokine induction by CHIKV infection differed between human and murine macrophages; IL-1, IL-6, IFN-γ, and TNF were significantly upregulated in human macrophages. This evidence suggests that CHIKV replicates more efficiently and induces a much greater pro-inflammatory cytokine profile in human macrophages, when compared to murine macrophages. This may shed light on the critical role that macrophages play in the CHIKV inflammatory response.
| S-EPMC7067478 | biostudies-literature
Project description:The venom of Apis mellifera (honey bee) has been reported to play a role in immunotherapy, but existing evidence to support its immuno-modulatory claims is insufficient. Four fractions from whole bee venom (BV) were separated using medium pressure liquid chromatography. Their ability to induce the production of cytokines TNF?, IL-1? and IL-6 in phorbol-12-myristate-13-acetate (PMA)-treated U937 cells was assessed. The levels of the three cytokines produced by stimulation with the four fractions and crude BV without LPS were not significantly different from negative control values. However, co-stimulation of the cells with LPS and Fraction 4 (F-4) induced a 1.6-fold increase in TNF-? level (p < 0.05) compared to LPS alone. Likewise, LPS-induced IL-1? production was significantly synergised in the presence of F-1 (nine-fold), F-2 (six-fold), F-3 (four-fold) and F-4 (two-fold) fractions, but was only slightly enhanced with crude BV (1.5-fold) relative to LPS. Furthermore, the LPS-stimulated production of IL-6 was not significantly increased in cells co-treated with F-2 and F-3, but the organic fraction (F-4) showed an inhibitory effect (p < 0.05) on IL-6 production. The latter was elucidated by NMR spectroscopy and found to contain(Z)-9-eicosen-1-ol. The effects observed with the purified BV fractions were more marked than those obtained with the crude sample.
| S-EPMC4931628 | biostudies-literature
Project description:ZXDC1 augments the expression of various markers of monocyte/macrophage differentiation when over-expressed in the U937 cell line treated with the phorbol ester PMA. Likewise, knockdown of ZXDC1 restricts the induced expression of these markers. We sought to identify specfic gene targets of ZXDC1 during the process of monocyte/macrophage differentiation in U937 by performing gene expression profiling in cells exhibiting reduced expression of ZXDC1 compared to controls. We used microarray to identify genes regulated by ZXDC during PMA-induced differentiation of U937 monoblasts towards a monocyte/macrophage phenotype
2013-10-01 | GSE45417 | GEO
Project description:Human histomonocytic U937 cells exhibit macrophage-like properties after phorbol ester stimulation. Accumulating evidence demonstrated that remarkable number of transcripts changed in their amount at total RNA levels. However, post-transcriptional regulation has not been fully elucidated in this model. Thus, we compared expression profiles between polysomal and cytoplasmic RNAs before and after phorbol 12-myristate 13-acetate stimulation (48h, 32nM). Table 1: Detailed description of the expression data of human histomonocytic cell line U937 cells. Data of total cytoplasmic and polysomal fractions before and after PMA stimulation is shown. This table contains all spot data except: 1) saturated spots, 2) undetectable spots, 3) stained spots, and 4) spots only detected in less than one samples. Column A: Gene symbol (“I_xxxxxx” is corresponding to EST clones) Column B: Genbank accession no. Column C: Description Column D: Average value of expression levels in total cytoplasmic fraction in the absence of PMA Column E: Average value of expression levels in total cytoplasmic fraction in the presence of PMA (32nM, 48h) Column F: Average value of expression levels in polysomal fraction in the absence of PMA Column G: Average value of expression levels in polysomal fraction in the presence of PMA Column H: Expression ratio of polysome, PMA (-) to cytoplasm, PMA(-) Column I: Expression ratio of polysome, PMA (+) to cytoplasm, PMA(+) Column J: Expression ratio of cytoplasm, PMA (+) to cytoplasm, PMA(-) Column K: Expression ratio of polysome, PMA (+) to polysome, PMA(-) Column L: Different expression between polysome, PMA (-) and cytoplasm, PMA(-): different=1 Column M: Different expression between polysome, PMA (+) and cytoplasm, PMA(+): different=1 Column N: Different expression between cytoplasm, PMA (+) and cytoplasm, PMA(-): different=1 Column O: Different expression between polysome, PMA (+) and polysome, PMA(-): different=1 Table 2: Candidates post-transcriptionally regulated by PMA. We extracted 105 transcripts whose expression levels were altered only in either fraction accompanied by changes in polysome/cytoplasm expression ratio. Column A: Gene symbol (“I_xxxxxx” is corresponding to EST clones) Column B: Genbank accession no. Column C: Description Column D: Average value of expression levels in total cytoplasmic fraction in the absence of PMA Column E Average value of expression levels in total cytoplasmic fraction in the presence of PMA Column F: Average value of expression levels in polysomal fraction in the absence of PMA Column G: Average value of expression levels in polysomal fraction in the presence of PMA Column H: Expression ratio of polysome, PMA (-) to cytoplasm, PMA(-) Column I: Expression ratio of polysome, PMA (+) to cytoplasm, PMA(+) Column J: Expression ratio of cytoplasm, PMA (+) to cytoplasm, PMA(-) Column K: Expression ratio of polysome, PMA (+) to polysome, PMA(-) Keywords = phorbol ester Keywords = macrophage Keywords = polysome Keywords = post-transcriptional regulation Keywords: parallel sample
2004-11-01 | GSE1783 | GEO