title: Epigallocatechin-3-Gallate Attenuates the Effects of TNF-α in Vascular Endothelial Cells by Causing Ectodomain Shedding of TNF Receptor 1.
abstract: OBJECTIVE
  We investigated the mechanism underlying anti-tumor necrosis factor-α (TNF-α) effects of epigallocatechin-3-gallate (EGCG) in human aortic endothelial cells.
  
  
  METHODS
  Tumor necrosis factor receptor 1 (TNFR1) was assessed by Western blot analysis. Cytosolic Ca2+ was measured using Fluo-4 AM. A disintegrin and metalloprotease 10 (ADAM10) was localized by immunofluorescence staining.
  
  
  RESULTS
  EGCG caused ectodomain shedding of TNFR1 within 30 min and attenuated TNF-α-induced endothelin-1 (ET-1) expression. EGCG-induced TNFR1 ectodomain shedding was prevented by BAPTA-AM (intracellular Ca2+ chelator), but not by the absence of extracellular Ca2+. In physiologic extracellular Ca2+ concentration, EGCG markedly increased cytosolic Ca2+. Even in the absence of extracellular Ca2+, EGCG raised cytosolic Ca2+, though less potently. siRNA depletion of ADAM10 prevented EGCG-induced ectodomain shedding of TNFR1 and also diminished the inhibitory effect of EGCG on TNF-α-induced ET-1 expression. EGCG caused translocation of ADAM10 to the plasma membrane, and this effect was prevented by BAPTA-AM. Besides extracellular Ca2+ influx, release of intracellular stored Ca2+ caused ADAM10-dependent ectodomain shedding of TNFR1.
  
  
  CONCLUSIONS
  EGCG decreases the responsiveness of cells to TNF-α by causing ADAM10-dependent ectodomain shedding of TNFR1. This effect was attributed to its property to increase cytosolic Ca2+ through both extracellular Ca2+ influx and release of stored Ca2+.
authors.forename: Won Seok
authors.initials: WS
authors.lastname: Yang
authors.forename: Soo Young
authors.initials: SY
authors.lastname: Moon
authors.forename: Mee Jeong
authors.initials: MJ
authors.lastname: Lee
authors.forename: Su-Kil
authors.initials: SK
authors.lastname: Park
mesh_terms: D000072197:ADAM10 Protein / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002392:Catechin / Q000031:analogs & derivatives* / Q000494:pharmacology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D019332:Endothelin-1 / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects*; D006801:Humans; D008856:Microscopy, Fluorescence; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000737:chemistry / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D019332:Endothelin-1; D034741:RNA, Small Interfering; D047888:Receptors, Tumor Necrosis Factor, Type I; D014409:Tumor Necrosis Factor-alpha; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D004533:Egtazic Acid; D002392:Catechin; C045651:epigallocatechin gallate; D000072197:ADAM10 Protein; D002118:Calcium