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RNA-seq of co-culture experiment between human HUVEC endothelial cells and mouse glioma GL126 cells

ABSTRACT: Increased awareness for the role for angiocrine factors in stem cell biology, in general, has suggested a role for angiocrine factors in the maintenance of GSC stemness and also there are reciprocal bi-directional signals from endothelial cells (ECs) to glioma (Gl261) and vice versa. To extend, refine and elucidate perfusion-independent modes of GBM-EC communication (i.e., distinguishing contact dependent or contact independent routes), we carried-out the following experiments: ECs from umbilical veins tagged with RFP (HUVEC-RFP) were cultured with mouse glioma line Gl261 fluorescently labelled with GFP (Gl261-GFP) until reaching confluency by 72 h post culturing. Cells were than harvested, RNA extracted, and the combined co-culture transcriptomes were sequenced from a generated cDNA library without prior separation of the respective cell types (sample named as Co). The Co transcriptome was compared to that of an artificial mixtures of the two cell types (Gl261-GFP and HUVEC-RFP) in the exact cells ratio attained by the end of the co-cluttering aided by FACS-based determination of the GFP/RFP ratio (Gl261:HUVEC - 3:1) attained by the end of the co-cluttering aided by FACS-based determination of the GFP/RFP ratio (sample named as Sep). In this procedure advantage was taken on the different species from which GBM and EC are derived and specialized bioinformatics tools allowing to unambiguously assign the transcripts to their respective GBM (mouse) or EC (human) origins, thereby circumventing confounding factors associated with cell purification

INSTRUMENT(S): NextSeq 500

ORGANISM(S): Mus musculus

SUBMITTER: Saran Kumar

PROVIDER: E-MTAB-9610 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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RNA-seq of co-culture experiment between human endothelial cells (HUVECs) and mouse glioma (Gl261) with or without Notch Inhibitor Gamma Secretase Inhibitor (GSI)

Project description:Increased awareness for the role for angiocrine factors in stem cell biology, in general, has suggested a role for angiocrine factors in the maintenance of GSC stemness and also there are reciprocal bi-directional signals from endothelial cells (ECs) to glioma (Gl261) and vice versa. To extend, refine and elucidate perfusion-independent modes of GBM-EC communication (i.e., distinguishing contact dependent or contact independent routes), we carried-out the following experiments: ECs from umbilical veins tagged with RFP (HUVEC-RFP) were cultured with mouse glioma line Gl261 fluorescently labelled with GFP (Gl261-GFP) until reaching confluency by 72 h post culturing. Cells were than harvested, RNA extracted, and the combined co-culture transcriptomes were sequenced from a generated cDNA library without prior separation of the respective cell types (sample named as Co). The Co transcriptome was compared to that of an artificial mixtures of the two cell types (Gl261-GFP and HUVEC-RFP) in the exact cells ratio attained by the end of the co-cluttering aided by FACS-based determination of the GFP/RFP ratio (sample named as Sep). In this procedure advantage was taken on the different species from which GBM and ECs are derived and specialized bioinformatics tools allowing to unambiguously assign the transcripts to their respective GBM (mouse) or EC (human) origins, thereby circumventing confounding factors associated with cell purification. Considering that contact-dependent EC signaling is primarily regulated by canonical Notch pathway and that the Notch pathway was reported to play a role in self-renewal of GSCs, we also treated GBM-EC co-cultures with the Notch pathway inhibitor Gamma secretase inhibitor (GSI) (or with the DMSO vehicle alone) for 24 h before harvesting.

2022-01-01 | E-MTAB-9604 | biostudies-arrayexpress

Transcription profiling of human melanoma-endothelial cells co-cultured vs controls to investigate cellular communication

Project description:Average life expectancy for patients with metastatic melanoma is less than 6 months, and only a handful of treatment options are available. If the disease can be stopped before it spreads to other organs, life expectancy is greatly increased. The goal of this project is to identify possible regulators of melanoma metastasis by determining genes whose expression is modulated when the cells are grown in contact with endothelial cells. Identification of genes involved in this cell-cell communication could have therapeutic implications. Experiment Overall Design: Using a lentiviral system, melanoma cell line 1205Lu was transduced with GFP, and HUVEC (human umbillical vein endothelial cells) cells at passage 2 were transduced with RFP. These labeled cells were plated at near confluency either alone (controls) or mixed at a 1:1 ratio (co-cultures) for 48 hours in endothelial growth medium (EGM-2, Cambrex). The labeled cells were then sorted by FACS and RNA was collected from all 4 cultures. There are a total of four samples: GFP-1205Lu cells grown alone, GFP-1205Lu cells separated from co-culture with RFP-HUVEC cells, RFP-HUVEC cells grown alone, and RFP-HUVEC cells separated from co-culture with GFP-1205Lu cells. The JHMI Microarray Core was responsible for sample processing and initial data processing using Affymetrix-recommended protocols (Affymetrix GeneChip Expression Analysis Technical Manual, 2004).

2008-10-18 | E-GEOD-8699 | biostudies-arrayexpress

Expression profile of human umbilical venous endothelial cells and human pluripotent stem cell-derived endothelial cells

Project description:We report that HUVEC-derived angiocrine factors amplify erythroblast. We took hPSC-EC as a reference

2019-09-28 | GSE138104 | GEO

Endothelial-to-osteoblast transition in normal mouse bone development and postnatal growth

Project description:To investigate the transcriptome of EC-OSB using Col1a1-GFP/Tie2-cre/Rosa-tdTomatodouble reporter mice, we sorted RFP+/GFP+ cells (EC-OSB) and RFP+ (non-EC-OSB cells) for RNA-seq analysis.

2022-12-01 | GSE207839 | GEO

Gene array analysis of Human Umbilical Vein Endothelial Cells (HUVEC) transduced with lentivirus containing cDNA of NKX2,5 or GATA4 or GFP.

Project description:The aim of this study was to induce cardiomyogenic conversion of Human Umbilical Vein Endothelial Cells (HUVEC ) by triggering ectopic expression of the cardiac transcription factors Nkx2.5 and GATA4. Cells were cultured without cell myocardial co-culture system and transduced with lentivirus containing coding sequence of Nkx2.5, GATA4 or GFP as control. Cells were cultured in the EBM-2/ EGM-2 medium, 12 days before microarray analysis. We report in this study that over-expression of NKX2.5 and GATA4 in HUVECs stimulates the expression of some specific genes important in the cardiomyogenic differentiation process, leading to partial switch of these cells from the endothelial to the myocardial course. 12 arrays were analysed. Three competitive hybridizations to the arrays were performed in duplicate using each time, two dye-swap following this experimental design: AWA065 : CY5 (HUVEC+NKX2,5) / CY3 (HUVEC) AWA067 : CY5 (HUVEC) / CY3 (HUVEC+NKX2,5) AWA075 : CY5 (HUVEC+NKX2,5) / CY3 (HUVEC) AWA076 : CY5 (HUVEC) / CY3 (HUVEC+NKX2,5) AWA068 : CY5 ((HUVEC) / CY3 (HUVEC+GATA4) AWA070 : CY5 (HUVEC+GATA4) / CY3 ((HUVEC) AWA073 : CY5 (HUVEC+GATA4) / CY3 ((HUVEC) AWA002 : CY5 ((HUVEC) / CY3 (HUVEC+GATA4) AWA066 : CY5 (HUVEC) / CY3 (HUVEC+NKX2.5+GATA4) AWA069 : CY5 (HUVEC+NKX2.5+GATA4) / CY3 (HUVEC) AWA071 : CY5 (HUVEC+NKX2.5+GATA4) / CY3 (HUVEC) AWA074 ; CY5 (HUVEC) / CY3 (HUVEC+NKX2.5+GATA4)

2012-04-20 | E-GEOD-37459 | biostudies-arrayexpress

Expression profiling of Endothelial (EC) and Non-endothelial Cells

Project description:Gene expression profiling of HUVEC (human umbilical vein EC cell; Lonza), HAEC (human aortic EC cells), HCAEC (human coronary artery EC cells), HPAEC (human pulmonary artery EC cells), HMVEC (human microvascular (dermal) , HASMC ( Human Aortic Smooth Muscle Cells), T cells and Bcells. Gene expression profiling of Endothelial cells and Non-endothelial cells in order to identify the genes with preferntial expression to endothelial cells. The experiments are performed in duplicate on both the HT Human Genome U133A and U133B arrays.

2011-04-04 | E-GEOD-21212 | biostudies-arrayexpress

Sorted MDMs with RFP+GFP+ or RFP+GFP- Mtb

Project description:Sorted MDMs with RFP+GFP+ or RFP+GFP- Mtb

| PRJNA542934 | ENA

Gene expression profiling of Human Umbilical Vein Endothelial Cells (HUVEC) after treatment with Erg or control antisense (GeneBloc)

Project description:The endothelial transcription factor Erg (Ets Related Gene) plays an important role in homeostasis and angiogenesis by regulating many endothelial functions including survival and junction stability. Here we show that Erg regulates endothelial cell migration. Transcriptome profiling of Erg-deficient endothelial cells (EC) identified 80 genes involved in cell migration as candidate Erg targets, including regulators of the Rho GTPases. Inhibition of Erg expression in human umbilical vein endothelial cells (HUVEC) resulted in decreased migration in vitro, whilst Erg over-expression using adenovirus caused increased migration. Live-cell imaging of Erg-deficient HUVEC showed a reduction in lamellipodia, in line with decreased motility. Both actin and tubulin cytoskeletons were disrupted in Erg-deficient EC, with a dramatic increase in tubulin acetylation. Amongst the most significant microarray hit was the cytosolic histone deacetylase (HDAC)-6, a regulator of cell migration. Rescue experiments confirmed that HDAC6 mediates the Erg-dependent regulation of tubulin acetylation and actin localization. The functional role of Erg in EC was studied by gene expressing profiling using three separate HUVEC isolates treated with either control antisense or Erg-specific antisense for 24 or 48 hours.

2012-02-27 | E-GEOD-32984 | biostudies-arrayexpress

Human Toll-Like Receptors Pathway Array Data

Project description:Endothelial cell (EC) Toll-like receptor 2 (TLR2) activation upregulates the expression of inflammatory mediators and of TLR2 itself, and modulates important endothelial functions, including coagulation and permeability. We defined TLR2 signaling pathways in ECs, and tested the hypothesis that TLR2 signaling differs in ECs and monocytes. We found that ERK5, heretofore unrecognized as mediating TLR2 activation in any cell type, is a central mediator of TLR2-dependent inflammatory signaling in HUVEC, primary human lung microvascular ECs and human monocytes. Additionally, we observed that whereas MEK1 negatively regulates TLR2 signaling in EC, MEK1 promotes TLR2 signaling in monocytes. We also noted that activation of TLR2 led to the upregulation of intracellularly expressed TLR2 and inflammatory mediators via NF-M-NM-:B, JNK and p38-MAPK. Finally, we found that p38-MAPK, JNK, ERK5 and NF-M-NM-:B promote the attachment of human neutrophils to lung microvascular EC that were pretreated with TLR2 agonists. This study newly identifies ERK5 as a key regulator of TLR2 signaling in ECs and monocytes, and indicates that there are fundamental differences in TLR signaling pathways between EC and monocytes. qPCR gene expression profiling. HUVEC monolayers were grown to confluency in EGM-2 media. THP1 suspension cells were grown RPMI 1640 supplemented with 10% FBS, L-glutamine, and antibiotics. Neither cell line was treated with any agonists. RQ values were calculated using the 2-M-NM-^TM-NM-^TCt method. Two biological replicates and one technical replicate were analyzed for both cell lines.

2011-11-04 | E-GEOD-33449 | biostudies-arrayexpress

Angiocrine Bmp2 signaling in murine liver controls normal iron homeostasis

Project description:Microvascular endothelial cells (EC) display a high degree of phenotypic and functional heterogeneity among different organs. Organ-specific EC control their tissue microenvironment by angiocrine factors in health and disease. Liver sinusoidal EC (LSEC) are uniquely differentiated to fulfil important organ-specific functions in development, under homeostatic conditions, and in regeneration and liver pathology. Recently, Bmp2 has been identified by us as an organ-specific angiokine derived from LSEC. To study angiocrine Bmp2 signaling in the liver, we conditionally deleted Bmp2 in LSEC using EC subtype-specific Stab2-Cre mice. Genetic inactivation of hepatic angiocrine Bmp2 signaling in Stab2-Cre;Bmp2fl/fl (Bmp2LSECKO) mice caused massive iron overload in the liver, and increased serum iron levels and iron deposition in several organs similar to classic hereditary hemochromatosis. Iron overload was mediated by decreased hepatic expression of hepcidin, a key regulator of iron homeostasis. Thus, angiocrine Bmp2 signaling within the hepatic vascular niche represents a constitutive pathway indispensable for iron homeostasis in vivo that is non-redundant with Bmp6. Notably, we demonstrate that organ-specific angiocrine signaling is essential not only for the homeostasis of the respective organ, but also for the homeostasis of the whole organism.

2017-01-31 | GSE90506 | GEO

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