Proliferative and antiapoptotic effects of EPC-derived MVs. (A,B) Eight thousand cells/well (A, HMECs; B, HUVECs) into 96-well plates were added with 10 μM BrdU, incubated for 72 hours in EBM-2 without FCS and growth factors in the presence of vehicle alone or of different doses of MVs (A, HMECs ■) or with 1 U/mL RNase-treated MVs (□). HUVECs (B) were incubated with vehicle alone (M199 medium) or vehicle plus 1 μg/mL of anti-α4 integrin– and anti-CD29–blocking Abs or with 10 μg/mL MVs alone or 10 μg/mL MVs preincubated (30 minutes, 37°C) with 1 μg/mL anti-α4 integrin or anti-CD29–blocking Ab or both or with 1 U/mL RNase or DNase (1 hour, 37°C). Cells were then fixed with 0.5 M ethanol/HCl and incubated with nuclease to digest the DNA. BrdU incorporated into the DNA was detected using an anti-BrdU peroxidase-conjugated mAb and visualized with a soluble chromogenic substrate. Optical density was measured with an ELISA reader at 405 nm. Results are expressed as mean (± 1 SD) of 3 experiments. Analysis of variance with Newmann-Keuls multicomparison test was performed; §P < .05 MVs versus vehicle alone; *P < .05 MV treatments versus MV alone. (C,D) The percentage of apoptotic cells after 48-hour serum withdrawal was evaluated by the TUNEL assay. HMECs (C) were incubated with vehicle alone or with different doses of MVs (■) or with 1 U/mL RNase-treated MV (▨). HUVECs (D) were incubated with vehicle alone or vehicle plus 1 μg/mL of anti-α4 integrin and anti-CD29–blocking Abs or with 10 μg/mL MV alone or 10 μg/mL MV preincubated (30 minutes, 37°C) with 1 μg/mL anti-α4 integrin or anti-CD29–blocking Ab or both or with 1 U/mL RNase (1 hour, 37°C). Results are expressed as mean (± 1 SD) of 3 experiments. Analysis of variance with Newmann-Keuls multicomparison test was performed; *P < .05 MVs versus vehicle alone; §P < .05 MV treatments versus MVs alone.