A20 protects from CD40-CD40 ligand-mediated endothelial cell activation and apoptosis.
BACKGROUND
CD40/CD40 ligand (CD40L) signaling is a potent activator of endothelial cells (ECs) and promoter of atherosclerosis. In this study, we investigate whether A20 (a gene we have shown to be antiinflammatory and antiapoptotic in ECs) can protect from CD40/CD40L-mediated EC activation.


RESULTS
Overexpression of CD40, in a transient transfection system, activates the transcription factor NF-kappaB and upregulates IkappaBalpha, E-selectin, and tissue factor (TF) reporter activity. Coexpression of A20 inhibits NF-kappaB and upregulation of IkappaBalpha and E-Selectin but not TF, suggesting that CD40 induces TF in a non-NF-kappaB-dependent manner. In human coronary artery ECs (HCAECs), adenovirus-mediated overexpression of A20 blocks physiological, CD40-induced activation of NF-kappaB, upstream of IkappaBalpha degradation (Western blot) and subsequently upregulation of ICAM-1, VCAM-1, and E-selectin (flow cytometry). Although A20 does not block TF transcription its expression in HCAECs inhibits TF induction (colorimetric assay and RT-PCR) by blunting CD40 upregulation. We demonstrate that CD40 signaling induces apoptosis in a proinflammatory microenvironment. A20 overexpression protects from CD40-mediated EC apoptosis (DNA content analysis and trypan blue exclusion). We also demonstrate that signaling through CD40L activates NF-kappaB and induces apoptosis in ECs, both of which are inhibited by A20 overexpression.


CONCLUSIONS
A20 works at multiple levels to protect ECs from CD40/CD40L mediated activation and apoptosis. A20-based therapy could be beneficial for the treatment of vascular diseases such as atherosclerosis and transplant-associated vasculopathy.
Longo
Christopher R
CR
Immunobiology Research Center, Department of Surgery and Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Mass 02215, USA.
Arvelo
Maria B
MB
Patel
Virendra I
VI
Daniel
Soizic
S
Mahiou
Jerome
J
Grey
Shane T
ST
Ferran
Christiane
C
D000818:Animals; D017209:Apoptosis; D019013:CD40 Antigens / Q000235:genetics / Q000378:metabolism*; D023201:CD40 Ligand / Q000494:pharmacology*; D002417:Cattle; D002478:Cells, Cultured; D016207:Cytokines / Q000037:antagonists & inhibitors; D019610:Cytoprotection; D004268:DNA-Binding Proteins; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D016328:NF-kappa B / Q000378:metabolism; D009687:Nuclear Proteins; D011506:Proteins / Q000235:genetics / Q000502:physiology*; D015398:Signal Transduction; D013925:Thromboplastin / Q000096:biosynthesis / Q000235:genetics; D015533:Transcriptional Activation; D014162:Transfection; D000072598:Tumor Necrosis Factor alpha-Induced Protein 3; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D020298:U937 Cells; D015854:Up-Regulation
D019013:CD40 Antigens; D016207:Cytokines; D004268:DNA-Binding Proteins; D047908:Intracellular Signaling Peptides and Proteins; D016328:NF-kappa B; D009687:Nuclear Proteins; D011506:Proteins; D014409:Tumor Necrosis Factor-alpha; D023201:CD40 Ligand; D013925:Thromboplastin; C076529:TNFAIP3 protein, human; D000072598:Tumor Necrosis Factor alpha-Induced Protein 3