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Blood, Vol. 111, Issue 8, 4145-4154, April 15, 2008
Angiopoietin-1 promotes endothelial cell proliferation and migration through AP-1–dependent autocrine production of interleukin-8
Blood Abdel-Malak et al.
111: 4145
Supplemental materials for: Abdel-Malak et al
Files in this Data Supplement:
- Document 1. Supplemental materials and methods (PDF, 85 KB)
- Table S1. Primer used for real-time PCR experiments (PDF, 68.9 KB)
- Figure S1. Effects of anti-IL-8 antibody on IL-8-induced proliferation and migration of HUVECs (JPG, 31.4 KB)
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For BrdU measurements, cells were exposed to recombinant IL-8 for a total of 48h. For migration assay, IL-8 (with or without neutralizing antibody) was placed into the lower chamber and migration was measured after 5 h. *P<0.05 compared with control.
- Figure S2. Effects of inhibition of Erk1/2 (30 µM PD98059) and p38 (10 µM PD169316) MAPKs on Ang-1-induced phosphorylation of SAPK/JNK proteins (Thr183/Tyr185) (JPG, 22.2 KB)
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Note that inhibition of Erk1/2 and p38 MAPK pathways had no effect on Ang-1-induced SAPK/JNK phosphorylation.
- Figure S3. Changes in mRNA expression (measured with real-time PCR) of c-Jun, JunB, JunD, c-Fos, FosB, Fra1 and Fra2 measured after 1 h of treatment of HUVECs with 300 ng/ml of Ang-1 (JPG, 36.9 KB)
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Data are means ± SEM (n=6) and are expressed as fold changes from HUVECs treated with Ang-1 vehicle. Note that Ang-1 exposure had no significant influence on the expression of these proteins.
- Figure S4 (JPG, 48.6 KB)
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(A) Changes in total I B- and p65 NFB phosphorylation (Ser536) in response to Ang-1. Actin used as loading control and TNF- treatment (10 min exposure) as positive control. (B) Mobilization of p50 and p65 NFB subunits in response to Ang-1 treatment. Cytosolic (C) and nuclear (N) fractions were analyzed with immunoblotting. (C) Binding of nuclear extracts from Ang-1 treated HUVECs to specific NFB DNA probes. Nuclear extracts provided by kit were used as positive control. Competition with the cold probe performed as specificity control. (D) HUVECs transfected with adenoviruses expressing GFP, dominant negative IKK- or IKK- proteins. Cells then exposed for 12h to Ang-1, IL-8 protein then detected in culture media. Data are expressed as mean ± SEM (n=6).
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