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Blood, Vol. 113, Issue 3, 744-754, January 15, 2009
CYP1B1 expression promotes the proangiogenic phenotype of endothelium through decreased intracellular oxidative stress and thrombospondin-2 expression
Blood Tang et al.
113: 744
Supplemental materials for: Tang et al
Files in this Data Supplement:
- Document 1. Supplemental materials and methods (PDF, 82.4 KB)
- Table S1. Mouse specific siRNAs (PDF, 39.3 KB)
- Figure S1. Retinal endothelial cells (EC) prepared from CYP1B1+∕+ and CYP1B1−∕− immorto mice (JPG, 101 KB)
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(A) Morphology of mouse retinal EC cultured on gelatin-coated plates. CYP1B1+∕+ and CYP1B1−∕− EC were cultured on gelatin-coated plates and photographed using a Nikon phase microscope in digital format. (B) Expression of cell-specific markers in EC. FACS analysis of PECAM-1 and VE-cadherin in CYP1B1+∕+ and CYP1B1−∕− EC.
- Figure S2. Migration of CYP1B1+∕+ and CYP1B1−∕− EC (JPG, 72.9 KB)
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(A) Scratch wound assays of EC. Confluent monolayers of CYP1B1+∕+ or CYP1B1−∕− EC were wounded and wound closure was monitored every 24 h and photographed in digital format (×40). A reprehensive experiment is shown. The quantitative assessment of the data is shown in (B) Data in each bar are the percent of wound closed (error bar indicate standard deviation). Please note that the CYP1B1−∕− EC migrated significantly slower than CYP1B1+∕+ EC (n=3, *P< 0.05). The transwell migration of CYP1B1+∕+ and CYP1B1−∕− EC is shown in (C). The mean number of migrated cells was determined by counting 10 high-power fields (×400; error bars indicate the standard deviation), following fixation and staining of membranes. Please note that the number of CYP1B1−∕− retinal EC which migrated through the transwell membrane were significantly lower compared with CYP1B1+∕+ retinal EC (n=3, *P< 0.05).These experiments were repeated with two different isolation of EC with similar results.
- Figure S3. Adhesion of CYP1B1+∕+ and CYP1B1−∕− EC to various matrix proteins (JPG, 54 KB)
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CYP1B1+∕+ and CYP1B1−∕− EC were plated on different concentrations of fibronectin (FN), vitronectin (VN), collagen (COL), or laminin (LN) and allowed to adhere. The number of adherent cells was determined by measuring intracellular acid phosphatase levels as described in Methods. Please note a significant decrease in the adhesion of CYP1B1−∕− retinal EC to fibronectin and vitronectin. These experiments were repeated with two different preparations of EC with similar results.
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