Figure 6.

Altered growth factor signaling by Ndst1-deficient microvascular endothelia. (a) TUNEL assay was performed on mutant and wild-type endothelial cells undergoing sprouting on Matrigel. In these experiments, cells derived from Ndst1 ^f/f TekCre⁻ mice were infected with Ad-Cre (Ndst1 ^f/f Ad-Cre) or Ad-GFP virus as a control (Ndst1 ^f/f Ad-GFP). After infection with Ad-Cre, numerous apoptotic bodies were observed within the cell clusters (bottom, arrows). Ad-GFP–infected cells formed extensive sprouting networks that were relatively free of apoptotic bodies, and the few nonsprouting clusters that were present also were devoid of apoptotic bodies (top right, inset). Bars, 100 μm. (b) Phosphorylation of Erk (p44/p42) in response to the growth factors FGF-2 or VEGF₁₆₄ was measured by Western blotting. The bands were scanned, the intensity of the phosphorylated protein was normalized to the intensity of the nonphosphorylated protein, and the ratio was scaled to the value obtained at t = 0 (values in parentheses). Total Erk was similar in mutant and wild-type cells. No significant variation in signal intensity >60 min was observed in the absence of growth factor in mutant and wild-type endothelia. (c) Phosphorylation of Akt was measured in wild-type and mutant cells.