| Title | Activation of intrinsic and extrinsic proapoptotic signaling pathways in interleukin-18-mediated human cardiac endothelial cell death. | | Author(s) | Chandrasekar B, Vemula K, Surabhi RM, Li-Weber M, Owen-Schaub LB, Jensen LE, Mummidi S | | Institution | Department of Medicine, University of Texas Health Science Center, 7703 Floyd Curl Drive, San Antonio, TX 78229, USA. chandraseka@uthscsa.edu | | Source | J Biol Chem 2004 May 7; 279(19):20221-33. | | MeSH | Apoptosis
Blotting, Western
Cell Death
Cell Separation
Cells, Cultured
DNA
Down-Regulation
Endothelium, Vascular
Enzyme Activation
Enzyme-Linked Immunosorbent Assay
Flow Cytometry
Genes, Dominant
Humans
Immunoblotting
Inflammation
Interleukin-1
Interleukin-18
Luciferases
Mitochondria
Myocardium
NF-kappa B
Oligonucleotides, Antisense
Protein Binding
Proto-Oncogene Proteins c-bcl-2
RNA, Messenger
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Research Support, U.S. Gov't, P.H.S.
Signal Transduction
Time Factors
Tumor Necrosis Factor-alpha
Up-Regulation
bcl-X Protein
| | Abstract | Endothelial cells are the primary targets of circulating immune and inflammatory mediators. We hypothesize that interleukin-18, a proinflammatory cytokine, induces endothelial cell apoptosis. Human cardiac microvascular endothelial cells (HCMEC) were treated with interleukin (IL) 18. mRNA expression was analyzed by ribonuclease protection assay, protein levels by immunoblotting, and cell death by enzyme-linked immunosorbent assay and fluorescence-activated cell sorter analysis. We also investigated the signal transduction pathways involved in IL-18-mediated cell death. Treatment of HCMEC with IL-18 increases 1) NF-kappaB DNA binding activity; 2) induces kappaB-driven luciferase activity; 3) induces IL-1beta and TNF-alpha expression via NF-kappaB activation; 4) inhibits antiapoptotic Bcl-2 and Bcl-X(L); 5) up-regulates proapoptotic Fas, Fas-L, and Bcl-X(S) expression; 6) induces fas and Fas-L promoter activities via NF-kappaB activation; 7) activates caspases-8, -3, -9, and BID; 8) induces cytochrome c release into the cytoplasm; 9) inhibits FLIP; and 10) induces HCME cell death by apoptosis as seen by increased annexin V staining and increased levels of mono- and oligonucleosomal fragmented DNA. Whereas overexpression of Bcl-2 significantly attenuated IL-18-induced endothelial cell apoptosis, Bcl-2/Bcl-X(L) chimeric phosphorothioated 2'-MOE-modified antisense oligonucleotides potentiated the proapoptotic effects of IL-18. Furthermore, caspase-8, IKK-alpha, and NF-kappaB p65 knockdown or dominant negative IkappaB-alpha and dominant negative IkappaB-beta or kinase dead IKK-beta significantly attenuated IL-18-induced HCME cell death. Effects of IL-18 on cell death are direct and are not mediated by intermediaries such as IL-1beta, tumor necrosis factor-alpha, or interferon-gamma. Taken together, our results indicate that IL-18 activates both intrinsic and extrinsic proapoptotic signaling pathways, induces endothelial cell death, and thereby may play a role in myocardial inflammation and injury. | | Language | eng | | Pub Type(s) | Journal Article
| | PubMed ID | 14960579 |
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