Journal:

Article Title: Vascular Endothelial Growth Factor-C (VEGF-C/VEGF-2) Promotes Angiogenesis in the Setting of Tissue Ischemia

doi:

Figure Lengend Snippet: Histological sections retrieved at day 30 from ischemic hindlimb muscle (musculus adductor) and stained with alkaline phosphatase (counterstained with eosin). A: RSA control; B: rhVEGF-C/VEGF-2 protein; C: pGSVLacZ control plasmid; D: pcVEGF-C/VEGF-2 plasmid-treated rabbit. Administration of VEGF-C as recombinant protein or plasmid resulted in increased capillary density. Dots, which appear dark blue, indicate capillaries.

Article Snippet: E. coli -derived rhVEGF-A appeared as a single protein band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a molecular weight of 39.8 kd under nonreducing conditions. rhVEGF-C (VEGF-2) was expressed in the BaculoGold Virus Expression Vector System (Pharmingen) and purified by ion exchange and hydrophobic interaction chromatography as described before.

Techniques: Staining, Plasmid Preparation, Recombinant

Journal:

Article Title: Vascular Endothelial Growth Factor-C (VEGF-C/VEGF-2) Promotes Angiogenesis in the Setting of Tissue Ischemia

doi:

Figure Lengend Snippet: Proliferative response of HUVECs (A) or HMECs (B) to rhVEGF-A/VEGF-1 or VEGF-C/VEGF-2 protein. Cells (5 × 103) were seeded per well, and increase in cell number was assessed 48 hours after addition of growth factors using the MTS-colorimetric assay. Data are means (bars, SEM) of parallel samples. *P < 0.05 VEGF-A/VEGF-1 versus VEGF-C/VEGF-2 protein.

Article Snippet: E. coli -derived rhVEGF-A appeared as a single protein band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a molecular weight of 39.8 kd under nonreducing conditions. rhVEGF-C (VEGF-2) was expressed in the BaculoGold Virus Expression Vector System (Pharmingen) and purified by ion exchange and hydrophobic interaction chromatography as described before.

Techniques: Colorimetric Assay

Journal:

Article Title: Vascular Endothelial Growth Factor-C (VEGF-C/VEGF-2) Promotes Angiogenesis in the Setting of Tissue Ischemia

doi:

Figure Lengend Snippet: Migratory response of HUVECs (A) or HMECs (B) to rhVEGF-A/VEGF-1 or VEGF-C/VEGF-2 protein. Cells (2.5 × 104) were seeded in the upper wells of a 48-well microchemotaxis Boyden chamber and incubated for 4 hours at 37°C in medium 199 supplemented with 1% FBS. The lower wells contained different concentrations of growth factor. Cells migrating through a polycarbonate membrane with a pore size of 8 μm were quantified by staining the cells at the lower side of the membrane with Giemsa solution and counting three high-power fields (×100). Each condition was done in quadruplicate. Data are means (bars, SEM) of parallel samples. *P < 0.05 VEGF-A/VEGF-1 versus VEGF-C/VEGF-2 protein.

Article Snippet: E. coli -derived rhVEGF-A appeared as a single protein band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a molecular weight of 39.8 kd under nonreducing conditions. rhVEGF-C (VEGF-2) was expressed in the BaculoGold Virus Expression Vector System (Pharmingen) and purified by ion exchange and hydrophobic interaction chromatography as described before.

Techniques: Incubation, Staining

Journal:

Article Title: Vascular Endothelial Growth Factor-C (VEGF-C/VEGF-2) Promotes Angiogenesis in the Setting of Tissue Ischemia

doi:

Figure Lengend Snippet: Induction of NO release from HUVECs by VEGF-C/VEGF-2, measured with an NO-specific polarographic electrode connected to an NO meter. HUVECs in six-well plates were bathed with Krebs-Henseleit solution before addition of reagents. Baseline is defined as baseline fluctuation over a period of 5 minutes of NO production by cells not exposed to agonist. The NO donor S-nitroso-N-acetyl-penicillamine (0.1 mmol/L) and VEGF-A/VEGF-1 (100 ng/ml) were used as positive controls. Administration of the NO inhibitor l-NAME abrogated the effect of VEGF-C and served as a negative control. Data shown are means (bars, SEM) of four to six measurements in each group. *P < 0.05 versus 100 ng/ml VEGF-C.

Article Snippet: E. coli -derived rhVEGF-A appeared as a single protein band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a molecular weight of 39.8 kd under nonreducing conditions. rhVEGF-C (VEGF-2) was expressed in the BaculoGold Virus Expression Vector System (Pharmingen) and purified by ion exchange and hydrophobic interaction chromatography as described before.

Techniques: Negative Control

Journal:

Article Title: Vascular Endothelial Growth Factor-C (VEGF-C/VEGF-2) Promotes Angiogenesis in the Setting of Tissue Ischemia

doi:

Figure Lengend Snippet: A: Ratio of hindlimb perfusion pressures at day 0 (immediately before treatment) and day 30. The blood pressure ratio was defined for each rabbit as the ratio of systolic pressure of the ischemic limb to systolic pressure of the normal limb (n = 8 each). At day 0, no differences were observed among the groups. At day 30, the blood pressure ratio is significantly greater in rabbits receiving rhVEGF-C/VEGF-2 protein versus RSA controls and in rabbits receiving pcVEGF-C/VEGF-2 plasmid versus pGSVLacZ controls. B: Angiographic score, derived by quantitative analysis of angiographically demonstrable vessels in the medial thigh of the ischemic hindlimb, at days 0 and 30. At day 0, there are no differences among the groups. At day 30, the number of vessels is significantly greater in the VEGF-C/VEGF-2 protein- and plasmid-treated groups compared with controls. Administration of plasmid DNA appeared to yield a more pronounced effect than use of recombinant protein (n = 8 each). C: Iliac blood flow reserve (ratio between blood flow at rest and maximal flow induced by nitroprusside) measured from the internal iliac artery of the ischemic limb at day 30, using intra-arterial Doppler wire. Both VEGF-C/VEGF-2 protein and plasmid significantly increased iliac flow reserve (n = 8 each). D: Capillary density evaluated at day 30 in histological sections harvested from the medial thigh muscles of the nonischemic and ischemic limbs. In the nonischemic limb, capillary density was not different among groups (not shown). In the ischemic limb, capillary density is increased significantly by VEGF-C/VEGF-2 protein and plasmid (n = 8 each). NS, not significant.

Article Snippet: E. coli -derived rhVEGF-A appeared as a single protein band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a molecular weight of 39.8 kd under nonreducing conditions. rhVEGF-C (VEGF-2) was expressed in the BaculoGold Virus Expression Vector System (Pharmingen) and purified by ion exchange and hydrophobic interaction chromatography as described before.

Techniques: Plasmid Preparation, Derivative Assay, Recombinant