Regulation of endothelin-1 by angiopoietin-1: implications for inflammation.
Endothelin-1 (ET-1) is increasingly recognized as a proinflammatory mediator in various diseases, such as atherosclerosis and acute respiratory distress syndrome (ARDS). Angiopoietin-1 (Ang-1), a ligand of the endothelial receptor Tie2, inhibits endothelial apoptosis, reduces vascular leakage, and suppresses the induction of inflammatory markers, indicating that it has diverse vasoprotective, anti-inflammatory actions. Thus, we examined the effects of Ang-1 on ET-1 production in vitro and in vivo and investigated cell-based gene transfer of Ang-1 in a rat model of lipopolysaccharide (LPS)-induced ARDS. Cultured human endothelial cells were treated with recombinant Ang-1 with or without tumor necrosis factor-alpha (TNF-alpha) (100 U/ml). ET-1 release into the culture medium after 24 hrs was determined by enzyme-linked immunosorbent assay. Levels of preproendothelin-1 (ppET-1) mRNA were measured by quantitative reverse transcription-polymerase chain reaction. Fisher344 rats were subjected to cell-based gene transfer to the lung circulation by injecting syngeneic fibroblasts transfected with Ang-1 cDNA or a null plasmid vector. After 24 hrs, LPS (100 microg/kg body wt) was instilled intratracheally to induce pulmonary inflammation. Bronchoalveolar lavage was performed 6 hrs later, and lungs were harvested for histologic and molecular analyses. ET-1 release from cultured endothelial cells was dose-dependently reduced by Ang-1, which also prevented induction of ET-1 release by TNF-alpha (P < 0.05). RNA expression of ppET-1 was similarly reduced. In LPS-challenged lungs, ppET-1 RNA was induced 3.4-fold, and ET-1 protein in lavage fluid was increased 5.6-fold (P < 0.05). Ang-1 gene transfer attenuated the LPS-induced increases in ppET-1 RNA and lavage ET-1 protein by 34% and 33%, respectively (P < 0.05). The downregulation of ET-1 correlated with the amelioration of pulmonary inflammation, as indicated by reductions in leukocyte infiltration (by 43%) and intra-alveolar septal thickening (by 40%). These results show that ET-1 transcript and protein levels are downregulated by Ang-1 in both in vitro and in vivo systems and that cell-based Ang-1 gene transfer markedly ameliorated inflammation in vivo in an experimental model of ARDS. Thus, cell-based gene transfer of Ang-1 may provide a novel treatment strategy for ARDS by attenuating vascular inflammation via suppression of ET-1.
McCarter
Sarah D
SD
Terrence Donnelly Research Laboratories, Division of Cardiology, St. Michae's Hospital, Toronto, Ontario, Canada.
Lai
Patrick F H
PF
Suen
Renée S
RS
Stewart
Duncan J
DJ
D042683:Angiopoietin-1 / Q000235:genetics / Q000494:pharmacology* / Q000627:therapeutic use; D000818:Animals; D002478:Cells, Cultured; D003470:Culture Media / Q000032:analysis; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation / Q000187:drug effects; D019332:Endothelin-1 / Q000032:analysis / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects*; D018014:Gene Transfer Techniques; D007249:Inflammation / Q000235:genetics; D008171:Lung Diseases / Q000188:drug therapy / Q000473:pathology; D008297:Male; D011897:Random Allocation; D051381:Rats; D011916:Rats, Inbred F344; D012867:Skin / Q000166:cytology
D042683:Angiopoietin-1; D003470:Culture Media; D019332:Endothelin-1