Homo sapiens
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ABSTRACT:
PROVIDER: PRJNA126095 | ENA |
REPOSITORIES: ENA
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Project description:Lipid rafts are cholesterol-rich cell signaling platforms and their physiological role can be explored by cholesterol depletion. To dress a global picture of transcriptional changes ongoing after lipid raft disruption, we performed whole-genome expression profiling in epidermal keratinocytes, a cell type which synthesizes its cholesterol in situ. We used microarrays to identify transcriptional changes in gene expression of cholesterol-depleted keratinocytes. Cholesterol depletion by methyl-beta-cyclodextrin disrupts the organization of lipid rafts, which are cholesterol- and sphingolipid-rich membrane microdomains. Transcript levels were measured in autocrine confluent cultures of normal human epidermal keratinocytes were either left untreated (Ctrl), cholesterol-depleted for 1h with 7.5mM methyl-beta-cyclodextrin (MBCD), or mock cholesterol-depleted for 1h with 7.5mM cholesterol-charged methyl-beta-cyclodextrin complexes (MBCD/chol) (Mock cholesterol depletion is a suppementary negative control as this treatment does not extract cholesterol from cell membranes). Samples are analysed either immediately after the treatment (R0h) or after recovery times of 1h (R1h) respectively 8h (R8h). in total 9 samples are analysed and no replicates are performed.
2010-04-17 | E-GEOD-21364 | biostudies-arrayexpress
Project description:Lipid rafts are cholesterol-rich cell signaling platforms and their physiological role can be explored by cholesterol depletion. To dress a global picture of transcriptional changes ongoing after lipid raft disruption, we performed whole-genome expression profiling in epidermal keratinocytes, a cell type which synthesizes its cholesterol in situ. We used microarrays to identify transcriptional changes in gene expression of cholesterol-depleted keratinocytes. Cholesterol depletion by methyl-beta-cyclodextrin disrupts the organization of lipid rafts, which are cholesterol- and sphingolipid-rich membrane microdomains.
2010-04-17 | GSE21364 | GEO
Project description:A possible explanation for chronic inflammation in HIV-infected individuals treated with anti-retroviral therapy is hyperreactivity of myeloid cells due to a phenomenon called ‘trained immunity’. Here, we demonstrate that human monocyte-derived macrophages originating from monocytes initially treated with extracellular vesicles containing HIV-1 protein Nef (exNef), but differentiating in the absence of exNef, released increased levels of pro-inflammatory cytokines after lipopolysaccharide stimulation. This effect was associated with epigenetic changes related to inflammation and cholesterol metabolism pathways, upregulation of the lipid rafts, and was blocked by methyl-beta-cyclodextrin, statin, and by an inhibitor of the lipid raft-associated receptor IGF1R. Bone marrow-derived macrophages from exNef-injected mice, as well as from mice transplanted with bone marrow from exNef-injected animals, produced elevated levels of TNFalpha upon stimulation. These phenomena are consistent with exNef-induced trained immunity that may contribute to persistent inflammation and associated co-morbidities in HIV-infected individuals with undetectable HIV load.
2022-10-09 | GSE214958 | GEO
Project description:A possible explanation for chronic inflammation in HIV-infected individuals treated with anti-retroviral therapy is hyperreactivity of myeloid cells due to a phenomenon called ‘trained immunity’. Here, we demonstrate that human monocyte-derived macrophages originating from monocytes initially treated with extracellular vesicles containing HIV-1 protein Nef (exNef), but differentiating in the absence of exNef, released increased levels of pro-inflammatory cytokines after lipopolysaccharide stimulation. This effect was associated with epigenetic changes related to inflammation and cholesterol metabolism pathways, upregulation of the lipid rafts, and was blocked by methyl-beta-cyclodextrin, statin, and by an inhibitor of the lipid raft-associated receptor IGF1R. Bone marrow-derived macrophages from exNef-injected mice, as well as from mice transplanted with bone marrow from exNef-injected animals, produced elevated levels of TNFalpha upon stimulation. These phenomena are consistent with exNef-induced trained immunity that may contribute to persistent inflammation and associated co-morbidities in HIV-infected individuals with undetectable HIV load.
2022-10-09 | GSE214957 | GEO
Project description:Hereditary elliptocytosis is a red blood cell (RBC) disease mainly caused by mutations in spectrin, leading to cytoskeletal destabilization. Although patients with heterozygous mutation in α-spectrin (SPTA1) are asymptomatic, morphological changes and hemolysis are observed upon reduced functional α-spectrin production. The molecular mechanism is unknown. We analyzed the consequences of a α-spectrin mutation in a patient almost exclusively expressing the Pro260 variant of SPTA1 (pEl). pEl RBCs showed decreased size and circularity and increased fragility. The pEl RBC proteome was globally preserved but spectrin density at the cell edges rised and the two membrane anchorage complexes were less segregated. Hence, the pEl membrane was strongly impaired. First, the lipidome was modified, showing decreased phosphatidylserine vs increased lysophosphatidylserine species. Second, although membrane transversal asymmetry was preserved, curvature at the RBC edges and rigidity were increased. Third, sphingomyelin-enriched domains were altered in abundance, membrane:cytoskeleton anchorage and cholesterol content were targeted by the plasmatic acid sphingomyelinase (aSMase). Fourth, membrane calcium exchanges through the mechanosensitive channel PIEZO1 and the efflux pump PMCA were impaired, leading to increased intracellular calcium and ROS, lipid peroxidation and methemoglobin. Mecanistically, increased curvature through lysophosphatidylserine membrane insertion in healthy RBCs abrogated calcium influx. Cholesterol depletion of sphingomyelin-enriched domains by methyl-beta-cyclodextrin in pEl RBCs worsened calcium accumulation whereas aSMase inhibition by amitriptyline did the opposite. Those data indicated that α-spectrin tetramerization defect, lysophosphatidylserine, cholesterol domain depletion and aSMase cooperate to alter membrane properties and calcium exchanges, leading to calcium accumulation and oxidative stress. It could help develop novel therapies
2020-08-04 | PXD019059 | Pride
Project description:Deciphering signaling mechanisms critical for the extended pluripotent stem cell (EPSC) state and primed pluripotency is necessary for understanding embryonic development. Here, we identify a membrane protein, Podocalyxin-like protein 1 (PODXL) as being essential for extended and primed pluripotency. Alteration of PODXL expression levels affects self-renewal, protein expression of c-MYC and telomerase, and induced pluripotent stems cell (iPSC) and EPSC colony formation. PODXL is the first membrane protein reported to regulate de novo cholesterol biosynthesis, and human pluripotent stem cells (hPSCs) are more sensitive to cholesterol depletion than fibroblasts. The addition of exogenous cholesterol fully restores PODXL knockdown-mediated loss of pluripotency. PODXL affects lipid raft dynamics via the regulation of cholesterol. PODXL recruits the RAC1/CDC42/actin network to regulate SREBP1 and SREBP2 maturation and lipid raft dynamics. Single-cell RNA sequencing reveals PODXL overexpression enhanced chimerism between human cells in mouse host embryos (hEPSCs 57%). Interestingly, in the human-mouse chimeras, laminin and collagen signaling-related pathways are dominant in PODXL overexpressing cells. We conclude that cholesterol regulation via PODXL signaling is critical for ESC/EPSC.
2022-09-22 | GSE213618 | GEO
Project description:Eukaryotic lipid rafts are membrane microdomains rich in cholesterol that contain a selective set of proteins, and have been associated with multiple biological functions. The Lyme disease agent, Borrelia burgdorferi, is one of an increasing number of bacterial pathogens that incorporates cholesterol onto its membrane, and form cholesterol glycolipid domains that possess all the hallmarks of eukaryotic lipid rafts. In this study, we isolated lipid rafts from cultured B. burgdorferi as a detergent resistant membrane (DRM) fraction on density gradients, and characterized those molecules that partitioned exclusively or are highly enriched in these domains. Cholesterol glycolipids, the previously known raft-associated lipoproteins OspA and OpsB, and cholera toxin partitioned into the lipid rafts fraction indicating compatibility with components of the DRM. The proteome of lipid rafts was analyzed by a combination of LC-MS/MS or MudPIT. Identified proteins were analyzed in silico for parameters that included localization, isoelectric point, molecular mass and biological function. The proteome provided a consistent pattern of lipoproteins, proteases and their substrates, sensing molecules, and prokaryotic homologs of eukaryotic lipid rafts. This study provides the first analysis of a prokaryotic lipid raft and has relevance for the biology of Borrelia, other pathogenic bacteria, as well as for the evolution of these structures.
2015-09-21 | PXD002365 | Pride
Project description:Complement inhibitor C4b-binding protein (C4BP) is synthesized in liver and pancreas and composed of 7 identical alpha chains and one unique beta chain. We showed previously that C4BP binds islet amyloid polypeptide (IAPP) and affects fibril formation in vitro. Now we found that polymeric C4BP inhibited lysis of human erythrocytes incubated with monomeric IAPP while no erythrocyte lysis was observed after incubation with preformed IAPP fibrils. In contrast, monomeric alpha chain of C4BP had significantly reduced activity. Further, addition of monomeric IAPP to a rat insulinoma cell line (INS-1) resulted in decreased cell viability, which was restored in the presence of physiological concentrations of C4BP. Accordingly, addition of C4BP rescued the ability of INS-1 cells and isolated rat islets to respond to glucose stimulation with insulin secretion, which was impaired in the presence of IAPP alone. C4BP was internalized together with IAPP into INS-1 cells and therefore we aimed to study its effect on gene expression. Pathway analyses of mRNA expression microarray data indicated that cells exposed to C4BP and IAPP in comparison to IAPP alone increased expression of genes involved in cholesterol synthesis. Depletion of cholesterol through methyl-β-cyclodextrin or cholesterol oxidase abolished the protective effect of C4BP on IAPP cytotoxicity of INS-1 cells. Also, inhibition of phosphoinositide 3-kinase but not NF-κB had a similar effect. Taken together, one of the mechanisms by which C4BP protects beta-cells from IAPP cytotoxicity is by enhancing cholesterol synthesis.
2016-08-25 | GSE79224 | GEO
Project description:Complement inhibitor C4b-binding protein (C4BP) is synthesized in liver and pancreas and composed of 7 identical alpha chains and one unique beta chain. We showed previously that C4BP binds islet amyloid polypeptide (IAPP) and affects fibril formation in vitro. Now we found that polymeric C4BP inhibited lysis of human erythrocytes incubated with monomeric IAPP while no erythrocyte lysis was observed after incubation with preformed IAPP fibrils. In contrast, monomeric alpha chain of C4BP had significantly reduced activity. Further, addition of monomeric IAPP to a rat insulinoma cell line (INS-1) resulted in decreased cell viability, which was restored in the presence of physiological concentrations of C4BP. Accordingly, addition of C4BP rescued the ability of INS-1 cells and isolated rat islets to respond to glucose stimulation with insulin secretion, which was impaired in the presence of IAPP alone. C4BP was internalized together with IAPP into INS-1 cells and therefore we aimed to study its effect on gene expression. Pathway analyses of mRNA expression microarray data indicated that cells exposed to C4BP and IAPP in comparison to IAPP alone increased expression of genes involved in cholesterol synthesis. Depletion of cholesterol through methyl-β-cyclodextrin or cholesterol oxidase abolished the protective effect of C4BP on IAPP cytotoxicity of INS-1 cells. Also, inhibition of phosphoinositide 3-kinase but not NF-κB had a similar effect. Taken together, one of the mechanisms by which C4BP protects beta-cells from IAPP cytotoxicity is by enhancing cholesterol synthesis. The INS-1 cells were grown as 5 separate clones for 10 passages before plating in a 12-well plate (Nunc) at 100.000 cells per well and grown in complete RPMI 1640 medium to 70% confluency for approximately 48 h. The cells were then challenged by adding 77 μM monomeric IAPP alone or together with C4BP (0.6 μM). DMSO (1%) used as solvent for IAPP as well as C4BP (0.6 μM) alone were used as controls. RNA was extracted after 10h incubation and analysis carried out using Rat Gene 2.0 array chip (Affymetrix).
2016-08-25 | E-GEOD-79224 | biostudies-arrayexpress
Project description:The aim of the experiment was to identify novel genes with relevance to cellular cholesterol metabolism. HeLa cells were cultivated while sterol was depleted from the medium by applying 2-Hydroxypropyl-cyclodextrin. RNA for expression profiling was isolated 3h, 4.5h and 6h after depletion.
2008-12-17 | E-TABM-599 | biostudies-arrayexpress