Mechanotransduction by integrin is essential for IL-6 secretion from endothelial cells in response to uniaxial continuous stretch.
We previously reported that uniaxial continuous stretch in human umbilical vein endothelial cells (HUVECs) induced interleukin-6 (IL-6) secretion via IkappaB kinase (IKK)/nuclear factor-kappaB (NF-kappaB) activation. The aim of the present study was to clarify the upstream signaling mechanism responsible for this phenomenon. Stretch-induced IKK activation and IL-6 secretion were inhibited by application of alpha(5)beta(1) integrin-inhibitory peptide (GRGDNP), phosphatidylinositol 3-kinase inhibitor (LY-294002), phospholipase C-gamma inhibitor (U-73122), or protein kinase C inhibitor (H7). Although depletion of intra- or extracellular Ca(2+) pool using thapsigargin (TG) or EGTA, respectively, showed little effect, a TG-EGTA mixture significantly inhibited stretch-induced IKK activation and IL-6 secretion. An increase in the intracellular Ca(2+) concentration ([Ca(2+)](i)) upon continuous stretch was observed even in the presence of TG, EGTA, or GRGDNP, but not in a solution containing the TG-EGTA mixture, indicating that both integrin activation and [Ca(2+)](i) rise are crucial factors for stretch-induced IKK activation and after IL-6 secretion in HUVECs. Furthermore, while PKC activity was inhibited by the TG-EGTA mixture, GRGDNP, LY-294002, or U-73122, PLC-gamma activity was retarded by GRGDNP or LY-294002. These results indicate that continuous stretch-induced IL-6 secretion in HUVECs depends on outside-in signaling via integrins followed by a PI3-K-PLC-gamma-PKC-IKK-NF-kappaB signaling cascade. Another crucial factor, [Ca(2+)](i) increase, may at least be required to activate PKC needed for NF-kappaB activation.
Sasamoto
Akitoshi
A
Department of Physiology, Nagoya Univ. Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan. msokabe@med.nagoya-u.ac.jp
Nagino
Masato
M
Kobayashi
Satoshi
S
Naruse
Keiji
K
Nimura
Yuji
Y
Sokabe
Masahiro
M
D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects / Q000502:physiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D006801:Humans; D051550:I-kappa B Kinase; D039081:Integrin alpha5beta1 / Q000378:metabolism*; D015850:Interleukin-6 / Q000378:metabolism*; D040542:Mechanotransduction, Cellular / Q000187:drug effects / Q000502:physiology*; D009025:Morpholines / Q000494:pharmacology; D009842:Oligopeptides / Q000494:pharmacology; D051966:Phospholipase C gamma; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011760:Pyrrolidinones / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000494:pharmacology; D014471:Umbilical Veins / Q000166:cytology / Q000187:drug effects / Q000378:metabolism
D002867:Chromones; D004791:Enzyme Inhibitors; D004963:Estrenes; D039081:Integrin alpha5beta1; D015850:Interleukin-6; D009025:Morpholines; D009842:Oligopeptides; D011760:Pyrrolidinones; C103774:glycyl-arginyl-glycyl-aspartyl-asparaginyl-proline; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D019284:Thapsigargin; D017346:Protein Serine-Threonine Kinases; C496559:CHUK protein, human; D051550:I-kappa B Kinase; C496560:IKBKB protein, human; C496565:IKBKE protein, human; D011493:Protein Kinase C; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D002118:Calcium