Effect of hypoxia and AdCA5 on invasion and tube formation by endothelial cells. (A) Matrigel invasion by ECs exposed to hypoxia or AdCA5. ECs were incubated for 24 hours in the presence or absence of adenovirus (5 pfu/cell), transferred to a Matrigel-coated membrane in a Boyden chamber, and exposed to 20% or 1% O2 for 24 hours, and the number of cells that had invaded through the Matrigel to the underside of the filter was counted. Three independent experiments were performed and the results for each condition were normalized to the result obtained for untreated cells (20% O2). P values were determined from the mean and standard deviation (shown) using Student t test. (B) Tube formation by ECs exposed to hypoxia or AdCA5. ECs were plated on Matrigel in the absence or presence (5 pfu/cell) of adenovirus (AdLacZ or AdCA5) that was added to the medium at the time of plating. Adenovirus-infected cells and one set of uninfected cells were incubated under nonhypoxic conditions (20% O2), whereas another set of uninfected cells was incubated under hypoxic conditions (1% O2). The cultures were analyzed 4, 8, and 24 hours after plating by phase-contrast microscopy. Each condition was performed in triplicate in 2 independent experiments and representative fields are shown.
