sterile hood and left exposed to air for 10 minutes for each sample, then transferred to a

24 well plate.

Cell Culture Techniques

 PVECs were supplied by Bert Herrara from Dr. Edward Block's lab weekly as

suspensions in 12 mL of media. All cells received from Dr. Block's lab were between

passage 2 and 5. Cells were expanded by diluting the suspension to 20 mL with fresh

media and subsequently transferred as 10 mL aliquots to a 75 cm2 angled neck, vented

tissue culture flask. Flasks were incubated at 370C and 5% CO2 for 48 hours, and then

existing media was exchanged for fresh. Media was changed every 72 hours after that.

Typically, the PVECs formed a confluent monolayer on the culture flask within 72 hours

from initial plating.

Cell passage procedure

 To passage the cells, confluent flasks were washed 3X with Hank's BSS. A few

mL of trypsin/EDTA 1X solution was poured in the flask and swirled to counteract any

remaining serum proteins and the remaining liquid was poured off. A small amount (-1-

2 mL) of fresh trypsin/EDTA solution was added again, with just enough to coat the

bottom of the flask. The flask was then placed in the incubator at 370C for 5 minutes,

and then checked with the inverted microscope to determine the appearance of the cells.

Once the cells became rounded, the sides of the flasks were struck on each side to

dislodge the remaining adherent cells. Serum containing media was added to counteract

the trypsin, and the suspension was mixed by aspiration with a 10 mL pipet. The

suspension was split into 3 flasks from each original flask and left to incubate as before.