Changes in cytosolic Ca2+ associated with von Willebrand factor release in human endothelial cells exposed to histamine. Study of microcarrier cell monolayers using the fluorescent probe indo-1

J Clin Invest. 1987 Feb;79(2):600-8. doi: 10.1172/JCI112853.

Abstract

A method for measuring fluorescence in anchored monolayers of human endothelial cells is described and used to demonstrate changes in the cytosolic free-calcium concentration ([Ca2+]c) in these cells exposed to histamine and thrombin; some endothelial responses to both agonists (e.g., mitogenesis) have been suggested to be Ca2+-mediated. Umbilical vein endothelial cells were cultured on microcarriers and loaded with the Ca2+ indicator, indo-1. Enzymatic cell detachment was avoided by monitoring the indo-1 fluorescence ratio (400/480 nm) of a stirred suspension of cell-covered microcarriers. Basal [Ca2+]c was estimated to be 70-80 nM. Thrombin induced a transient dose-dependent increase in [Ca2+]c, which was active-site dependent. Histamine stimulated a dose-dependent increase in [Ca2+]c, which was reversed by removal of histamine and inhibited competitively by the H1-receptor antagonist pyrilamine, but not by the H2-receptor antagonist cimetidine. Furthermore, histamine induced a dose-dependent secretion of von Willebrand factor, which paralleled the rise in [Ca2+]c and was similarly blocked by the H1-receptor antagonist, and which may contribute to platelet deposition at sites of inflammation.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Calcium / metabolism*
  • Cells, Cultured
  • Cytosol / metabolism
  • Endothelium / drug effects
  • Endothelium / metabolism*
  • Fluorescent Dyes
  • Histamine / pharmacology*
  • Humans
  • Indoles
  • Kinetics
  • Spectrometry, Fluorescence
  • Thrombin / physiology
  • Umbilical Veins
  • von Willebrand Factor / metabolism*

Substances

  • Fluorescent Dyes
  • Indoles
  • von Willebrand Factor
  • Histamine
  • Thrombin
  • indo-1
  • Calcium