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Angiogenesis interactome and time course microarray data reveal the distinct activation patterns in endothelial cells.

Chu LH, Lee E, Bader JS, Popel AS - PLoS ONE (2014)

Bottom Line: We used the Short Time-series Expression Miner (STEM) to identify significant temporal gene expression profiles.The statistically significant patterns between 2D fibronectin and 3D type I collagen substrates for telomerase-immortalized EC (TIME) show that different substrates could influence the temporal gene activation patterns in the same cell line.This computational methodology can be extended to investigate other molecules or biological processes such as cell cycle.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, School of Medicine, Johns Hopkins University, Baltimore, Maryland, United States of America.

ABSTRACT
Angiogenesis involves stimulation of endothelial cells (EC) by various cytokines and growth factors, but the signaling mechanisms are not completely understood. Combining dynamic gene expression time-course data for stimulated EC with protein-protein interactions associated with angiogenesis (the "angiome") could reveal how different stimuli result in different patterns of network activation and could implicate signaling intermediates as points for control or intervention. We constructed the protein-protein interaction networks of positive and negative regulation of angiogenesis comprising 367 and 245 proteins, respectively. We used five published gene expression datasets derived from in vitro assays using different types of blood endothelial cells stimulated by VEGFA (vascular endothelial growth factor A). We used the Short Time-series Expression Miner (STEM) to identify significant temporal gene expression profiles. The statistically significant patterns between 2D fibronectin and 3D type I collagen substrates for telomerase-immortalized EC (TIME) show that different substrates could influence the temporal gene activation patterns in the same cell line. We investigated the different activation patterns among 18 transmembrane tyrosine kinase receptors, and experimentally measured the protein level of the tyrosine-kinase receptors VEGFR1, VEGFR2 and VEGFR3 in human umbilical vein EC (HUVEC) and human microvascular EC (MEC). The results show that VEGFR1-VEGFR2 levels are more closely coupled than VEGFR1-VEGFR3 or VEGFR2-VEGFR3 in HUVEC and MEC. This computational methodology can be extended to investigate other molecules or biological processes such as cell cycle.

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Experiments of VEGFR1, VEGFR2 and VEGFR3 for MEC and HUVEC.
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pone-0110871-g004: Experiments of VEGFR1, VEGFR2 and VEGFR3 for MEC and HUVEC.

Mentions: We explored time-dependent VEGF receptor expression in blood endothelial cells after VEGF treatment, as the VEGF-VEGFR axis is pivotal in endothelial cell growth and maintenance. The experimental results for FLT1 (VEGFR1), KDR (VEGFR2) and FLT4 (VEGFR3) are shown in Figure 4. Briefly, one million of HUVEC or MEC were starved overnight, after which we treated the cells with 50 ng/ml of human VEGF165 and incubated them for 0, 1, 3, 6, 12 and 24 hr at 37°C. Total protein levels of VEGFR1/2/3 and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were obtained for normalization in data analyses. The number of pixels of each western band was analyzed using ImageJ (NIH, Bethesda). In VEGFR1 and VEGFR3 analyses, the full length VEGFR1 (180 kDa) and the full length VEGFR3 (195 kDa) in MEC and HUVEC were analyzed, as HUVEC do not express some isoforms. Each band was finally normalized by the GAPDH level.


Angiogenesis interactome and time course microarray data reveal the distinct activation patterns in endothelial cells.

Chu LH, Lee E, Bader JS, Popel AS - PLoS ONE (2014)

Experiments of VEGFR1, VEGFR2 and VEGFR3 for MEC and HUVEC.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4199761&req=5

pone-0110871-g004: Experiments of VEGFR1, VEGFR2 and VEGFR3 for MEC and HUVEC.
Mentions: We explored time-dependent VEGF receptor expression in blood endothelial cells after VEGF treatment, as the VEGF-VEGFR axis is pivotal in endothelial cell growth and maintenance. The experimental results for FLT1 (VEGFR1), KDR (VEGFR2) and FLT4 (VEGFR3) are shown in Figure 4. Briefly, one million of HUVEC or MEC were starved overnight, after which we treated the cells with 50 ng/ml of human VEGF165 and incubated them for 0, 1, 3, 6, 12 and 24 hr at 37°C. Total protein levels of VEGFR1/2/3 and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) were obtained for normalization in data analyses. The number of pixels of each western band was analyzed using ImageJ (NIH, Bethesda). In VEGFR1 and VEGFR3 analyses, the full length VEGFR1 (180 kDa) and the full length VEGFR3 (195 kDa) in MEC and HUVEC were analyzed, as HUVEC do not express some isoforms. Each band was finally normalized by the GAPDH level.

Bottom Line: We used the Short Time-series Expression Miner (STEM) to identify significant temporal gene expression profiles.The statistically significant patterns between 2D fibronectin and 3D type I collagen substrates for telomerase-immortalized EC (TIME) show that different substrates could influence the temporal gene activation patterns in the same cell line.This computational methodology can be extended to investigate other molecules or biological processes such as cell cycle.

View Article: PubMed Central - PubMed

Affiliation: Department of Biomedical Engineering, School of Medicine, Johns Hopkins University, Baltimore, Maryland, United States of America.

ABSTRACT
Angiogenesis involves stimulation of endothelial cells (EC) by various cytokines and growth factors, but the signaling mechanisms are not completely understood. Combining dynamic gene expression time-course data for stimulated EC with protein-protein interactions associated with angiogenesis (the "angiome") could reveal how different stimuli result in different patterns of network activation and could implicate signaling intermediates as points for control or intervention. We constructed the protein-protein interaction networks of positive and negative regulation of angiogenesis comprising 367 and 245 proteins, respectively. We used five published gene expression datasets derived from in vitro assays using different types of blood endothelial cells stimulated by VEGFA (vascular endothelial growth factor A). We used the Short Time-series Expression Miner (STEM) to identify significant temporal gene expression profiles. The statistically significant patterns between 2D fibronectin and 3D type I collagen substrates for telomerase-immortalized EC (TIME) show that different substrates could influence the temporal gene activation patterns in the same cell line. We investigated the different activation patterns among 18 transmembrane tyrosine kinase receptors, and experimentally measured the protein level of the tyrosine-kinase receptors VEGFR1, VEGFR2 and VEGFR3 in human umbilical vein EC (HUVEC) and human microvascular EC (MEC). The results show that VEGFR1-VEGFR2 levels are more closely coupled than VEGFR1-VEGFR3 or VEGFR2-VEGFR3 in HUVEC and MEC. This computational methodology can be extended to investigate other molecules or biological processes such as cell cycle.

Show MeSH
Related in: MedlinePlus