title: VEGF-mediated elevated intracellular calcium and angiogenesis in human microvascular endothelial cells in vitro are inhibited by dominant negative TRPC6.
abstract: OBJECTIVE
  Vascular endothelial growth factor (VEGF)-induced vascular permeability has been shown to be dependent on calcium influx, possibly through a transient receptor potential cation channel (TRPC)-mediated cation channel with properties of the TRPC3/6/7 subfamily. To investigate further the involvement of this subfamily, we determined the effects of dominant negative TRPC6 overexpression on VEGF-mediated changes of human microvascular endothelial cell (HMVEC) calcium, proliferation, migration, and sprouting.
  
  
  METHODS
  Cytoplasmic calcium concentration was estimated by fura-2 fluorescence spectrophotometry, migration by Boyden chamber assay, sprouting by immunofluorescence imaging of stimulated endothelial cells, and proliferation by flow cytometry.
  
  
  RESULTS
  Overexpression of a dominant negative TRPC6 construct in HMVECs inhibited the VEGF-mediated increases in cytosolic calcium, migration, sprouting, and proliferation. In contrast, overexpression of a wild-type TRPC6 construct increased the proliferation and migration of HMVECs.
  
  
  CONCLUSIONS
  TRPC6 is an obligatory component of cation channels required for the VEGF-mediated increase in cytosolic calcium and subsequent downstream signaling that leads to processes associated with angiogenesis.
authors.affiliation: Bristol Heart Institute, Department of Physiology and Pharmacology, School of Veterinary Sciences, University of Bristol, United Kingdom.
authors.forename: M A
authors.initials: MA
authors.lastname: Hamdollah Zadeh
authors.forename: Catherine A
authors.initials: CA
authors.lastname: Glass
authors.forename: Anette
authors.initials: A
authors.lastname: Magnussen
authors.forename: Jules C
authors.initials: JC
authors.lastname: Hancox
authors.forename: David O
authors.initials: DO
authors.lastname: Bates
mesh_terms: D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D002199:Capillary Permeability / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D049109:Cell Proliferation / Q000187:drug effects; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism*; D005799:Genes, Dominant; D006801:Humans; D009154:Mutation; D018919:Neovascularization, Physiologic / Q000187:drug effects / Q000502:physiology*; D050052:TRPC Cation Channels / Q000096:biosynthesis* / Q000235:genetics; D000074287:TRPC6 Cation Channel; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism / Q000494:pharmacology*
chemical_list: D050052:TRPC Cation Channels; D000074287:TRPC6 Cation Channel; C117032:TRPC6 protein, human; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D002118:Calcium
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