Background: The forkhead transcription factor, FOXO3a, is known to induce apoptosis in endothelial cells (EC). However, its effects on extracellular matrices (ECM), which are important in EC survival, are remained unknown. Here, we evaluated the role of FOXO3a on EC-ECM interaction.
Methods and Results:
1) Constitutively active, triple-mutant (TM) FOXO3a was transduced to human umbilical vein endothelial cells by adenoviral vector. Transduction with TM-FOXO3a led to dehiscence of EC from the fibronectin-coated plate, resulting apoptosis, which was significantly reversed by matrix metalloproteinase (MMP) inhibitor, GM6001.
2) In real time RT-PCR and Western blot analysis, FOXO3a increased the expression of MMP-3 and MMP-7, whereas decreased tissue inhibitors of metalloproteinases-1 (TIMP-1). In casein and gelatin zymography, FOXO3a increased overall MMP enzymatic activity.
3) FOXO3a-transduced ECs showed higher activity of fibronectin degradation leading to impaired adhesion to matrix. Moreover, FOXO3a transduction decreased the expression of VE-cadherin and 棺-catenin in pericellular matrix, leading to destruction of cell-cell contact. Such disturbances of cell-matrix or cell-cell interaction by FOXO3a were reversed by MMP inhibitor, GM6001, suggesting that MMPs play important roles in apoptosis of ECs by FOXO3a.
4) In vivo experiments, FOXO3a gene transfer to vascular wall induced EC denudation, leading to loss of barrier function of endothelium as well as loss of endothelium-dependent relaxation.
Conclusion: Activation of MMP and disruption of cell-to-cell or cell-to-matrix interaction represent novel mechanisms of FOXO3a-mediated apoptosis in EC.