SMAD1/5 binding regions and expression data of human umbilical vein endothelial cells (HUVECs) and pulmonary arterial smooth muscle cells (PASMCs) treated with BMPs
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ABSTRACT: This SuperSeries is composed of the SubSeries listed below.
Project description:Smad1/5 are transcription factors that engage in BMP-induced transcription. We determined and analyzed Smad1/5 binding sites by ChIP-sequencing. We used expression microarrays to compare the Smad1/5 binding sites identified by ChIP-seq to BMP-induced gene expressions.
Project description:Smad1/5 binding regions were identified by ChIP-seq. HUVECs were treated with BMP for 1.5 h and anti-SMAD1/5 ChIP-seq analyses were performed using Illumina genome analyzer.
Project description:Human umbilical cords were obtained from the Lucille Packard Children Hospital. In this study, we devised a rapid isolation scheme to preserve the in vivo phenotype of each endothelial subtype. ECs were first isolated from umbilical cords by collagenase perfusion through the vein or artery as described. Cells were further purified using a Percoll density gradient (Amersham Biosciences, Piscataway, NJ) to remove residual erythrocytes and platelets. 1-5 x 106 ECs were then cultured overnight on gelatin-coated T12.5 flasks in Clonetics EGM-2 media (Cambrex Bio Science, Walkersville, MD) in which the ECs generally reached confluence the next morning. The confluent EC were trypsinized and subjected to two rounds of immuno-magnetic beads purification, adapted from a published purification protocol. There was a negative selection step using CD14, CD45 and CD64 to remove residual contaminating leukocytes, followed by positive selection using a mouse anti-endothelial cell monoclonal antibody (anti-CD146/clone P1H12 purchased from Chemicon, Temecula, CA). Total processing time was limited to 20 to 24 hours. The homogeneous, viable, primary ECs were used immediately to construct the library. The construction of SAGE libraries was performed with the I-SAGE kit (Invitrogen, Carlsbad, CA) per manufacturer’s instructions Keywords: other
Project description:Human umbilical cords were obtained from the Lucille Packard Children Hospital. In this study, we devised a rapid isolation scheme to preserve the in vivo phenotype of each endothelial subtype. ECs were first isolated from umbilical cords by collagenase perfusion through the vein or artery as described. Cells were further purified using a Percoll density gradient (Amersham Biosciences, Piscataway, NJ) to remove residual erythrocytes and platelets. 1-5 x 106 ECs were then cultured overnight on gelatin-coated T12.5 flasks in Clonetics EGM-2 media (Cambrex Bio Science, Walkersville, MD) in which the ECs generally reached confluence the next morning. The confluent EC were trypsinized and subjected to two rounds of immuno-magnetic beads purification, adapted from a published purification protocol. There was a negative selection step using CD14, CD45 and CD64 to remove residual contaminating leukocytes, followed by positive selection using a mouse anti-endothelial cell monoclonal antibody (anti-CD146/clone P1H12 purchased from Chemicon, Temecula, CA). Total processing time was limited to 20 to 24 hours. The homogeneous, viable, primary ECs were used immediately to construct the library. The construction of SAGE libraries was performed with the I-SAGE kit (Invitrogen, Carlsbad, CA) per manufacturer’s instructions Keywords: other
Project description:Smad1/5 are transcription factors that engage in BMP-induced transcription. We determined and analyzed Smad1/5 binding sites by ChIP-sequencing. We used expression microarrays to compare the Smad1/5 binding sites identified by ChIP-seq to BMP-induced gene expressions. PASMCs were treated with 50 ng/ml BMP-4 for 0, 2, or 24 hrs. RNA was extracted and hybridized on Affymetrix microarrays.
Project description:This SuperSeries is composed of the following subset Series: GSE15482: The effect of PPARa siRNA on endothelial cells treated with fenofibrate GSE15483: The effect of fenofibrate on endothelial cells Refer to individual Series