Abstract 16325: Endothelial Microparticle Uptake in Target Cells is Annexin I/PSR Dependent and Prevents Apoptosis
Objective: Endothelial microparticles (EMP) are released from activated or apoptotic cells, but their effect on target cells and the exact way of incorporation are largely unknown. We sought to determine the uptake mechanism and the biological effect of EMP on endothelial and endothelial regenerating cells.
Methods and results: EMP were generated from starved endothelial cells and isolated by ultracentrifugation. Caspase 3 activity assay showed that EMP (2000 annexin V+/µl) protect target endothelial cells against camptothecin-induced apoptosis in a dose-dependent way. TUNEL assay further confirmed the beneficial effect of EMP against apoptosis in target cells (89.4±4.3% vs. 65.9±8.6%, p<0.001, n=5-6). Proteomic analysis was performed to identify molecules contained in EMP, which might be involved in EMP uptake. Abundant expression of annexin I in EMP was found and confirmed by western blot, while the corresponding receptor phosphatidylserine receptor (PSR) of phosphatidylserine was present on endothelial target cells. Silencing either annexin I on EMP or PSR on target cells using siRNA showed that the uptake of EMP by endothelial cells is annexin I/PSR dependent as demonstrated by immunofluorescence staining and ELISA. Annexin I-downregulated EMP abrogated the EMP-mediated protection against apoptosis of endothelial and endothelial regenerating target cells (p<0.05, n=4-6). p38-activation was found to mediate camptothecin-induced apoptosis. Finally, HCAEC pretreated with EMP inhibited camptothecin-induced p38 activation by restoring MKP-1 expression.
Conclusion: EMP are incorporated by endothelial cells in an annexin I/PSR dependent way and protect target cells against apoptosis. Inhibition of p38 activity is involved in EMP-mediated protection against apoptosis.
- © 2012 by American Heart Association, Inc.