Bradykinin regulates calpain and proinflammatory signaling through TRPM7-sensitive pathways in vascular smooth muscle cells.
Transient receptor potential melastatin-7 (TRPM7) channels have recently been identified to be regulated by vasoactive agents acting through G protein-coupled receptors in vascular smooth muscle cells (VSMC). However, downstream targets and functional responses remain unclear. We investigated the subcellular localization of TRPM7 in VSMCs and questioned the role of TRPM7 in proinflammatory signaling by bradykinin. VSMCs from Wistar-Kyoto rats were studied. Cell fractionation by sucrose gradient and differential centrifugation demonstrated that in bradykinin-stimulated cells, TRPM7 localized in fractions corresponding to caveolae. Immunofluorescence confocal microscopy revealed that TRPM7 distributes along the cell membrane, that it has a reticular-type intracellular distribution, and that it colocalizes with flotillin-2, a marker of lipid rafts. Bradykinin increased expression of calpain, a TRPM7 target, and stimulated its cytosol/membrane translocation, an effect blocked by 2-APB (TRPM7 inhibitor) and U-73122 (phospholipase C inhibitor), but not by chelerythrine (PKC inhibitor). Expression of proinflammatory mediators VCAM-1 and cyclooxygenase-2 (COX-2) was time-dependently increased by bradykinin. This effect was blocked by Hoe-140 (B2 receptor blocker) and 2-APB. Our data demonstrate that in bradykinin-stimulated VSMCs: 1) TRPM7 is upregulated, 2) TRPM7 associates with cholesterol-rich microdomains, and 3) calpain and proinflammatory mediators VCAM-1 and COX2 are regulated, in part, via TRPM7- and phospholipase C-dependent pathways through B2 receptors. These findings identify a novel signaling pathway for bradykinin, which involves TRPM7. Such phenomena may play a role in bradykinin/B2 receptor-mediated inflammatory responses in vascular cells.
Yogi
Alvaro
A
Kidney Research Centre, Ottawa Health Research Institute, University of Ottawa, 451 Smyth Rd. Ottawa, ON, Canada KIH 8M5.
Callera
Glaucia E
GE
Tostes
Rita
R
Touyz
Rhian M
RM
D000818:Animals; D053119:Benzophenanthridines / Q000494:pharmacology; D001896:Boron Compounds / Q000494:pharmacology; D001920:Bradykinin / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D065094:Bradykinin B2 Receptor Antagonists; D002154:Calpain / Q000378:metabolism*; D021941:Caveolae / Q000378:metabolism; D002478:Cells, Cultured; D051546:Cyclooxygenase 2 / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D018836:Inflammation Mediators / Q000378:metabolism*; D008274:Magnesium / Q000378:metabolism; D008638:Mesenteric Arteries / Q000276:immunology / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology / Q000276:immunology / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology / Q000276:immunology / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D021381:Protein Transport; D011760:Pyrrolidinones / Q000494:pharmacology; D051381:Rats; D011921:Rats, Inbred WKY; D043782:Receptor, Bradykinin B2 / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D050053:TRPM Cation Channels / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000378:metabolism; D015854:Up-Regulation; D019010:Vascular Cell Adhesion Molecule-1 / Q000378:metabolism
D053119:Benzophenanthridines; D001896:Boron Compounds; D065094:Bradykinin B2 Receptor Antagonists; D004791:Enzyme Inhibitors; D004963:Estrenes; D018836:Inflammation Mediators; D011760:Pyrrolidinones; D043782:Receptor, Bradykinin B2; D050053:TRPM Cation Channels; D019010:Vascular Cell Adhesion Molecule-1; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; C065679:icatibant; C016299:chelerythrine; C109986:2-aminoethoxydiphenyl borate; D051546:Cyclooxygenase 2; C489691:Ptgs2 protein, rat; C486584:Trpm7 protein, rat; D011493:Protein Kinase C; D010738:Type C Phospholipases; D002154:Calpain; D008274:Magnesium; D001920:Bradykinin