Mechanisms of ionomycin-induced endothelial cell barrier dysfunction

Joe G.N. Garcia, Kane L. Schaphorst, Shu Shi, Alexander D. Verin, C. Michael Hart, Karleen S. Callahan, Carolyn E. Patterson

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86 Scopus citations

Abstract

Myosin light chain (MLC) phosphorylation catalyzed by the Ca2+- calmodulin-dependent MLC kinase (MLCK) is critical to thrombin-mediated endothelial cell gap formation and barrier dysfunction. We have tested the hypothesis that the Ca2+ ionophore ionomycin stimulates MLCK-dependent endothelial cell contraction and permeability. Ionomycin significantly increased albumin clearance and decreased electrical resistance across confluent bovine pulmonary microvascular and macrovascular endothelial cell monolayers in a concentration-dependent manner that was temporally similar to that produced by thrombin. In contrast, however, ionomycin produced a significant Ca2+-dependent reduction in the levels of phosphorylated MLC with evidence of serine/threonine phosphatase activation. Potential MLCK- independent mechanisms of endothelial cell permeability were examined with little evidence to support a role for stimulated nitric oxide synthase or phospholipase A2 activities. Importantly, ionomycin produced 1) reductions in the activities of the barrier protective adenylate cyclase and the adenosine 3',5'-cyclic monophosphate-dependent protein kinase A, 2) dramatic dose- and time-dependent inhibition of endothelial cell tyrosine kinase activities, and 3) marked decreases in the phosphotyrosine content of the p125 focal adhesion kinase. These data indicate that ionomycin produces endothelial cell barrier dysfunction by mechanisms that are independent of MLCK activation and may involve reductions in endothelial cell tethering forces via inhibition of protein kinase A and tyrosine kinase activities, especially the p125 focal adhesion kinase.

Original languageEnglish (US)
Pages (from-to)L172-L184
JournalAmerican Journal of Physiology - Lung Cellular and Molecular Physiology
Volume273
Issue number1 17-1
Publication statusPublished - Jul 1 1997
Externally publishedYes

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Keywords

  • Adenosine 3',5'
  • Cyclic monophosphate
  • Focal adhesion kinase
  • Myosin light chain kinase
  • Protein kinase A
  • Tyrosine phosphorylation

ASJC Scopus subject areas

  • Physiology
  • Pulmonary and Respiratory Medicine
  • Physiology (medical)
  • Cell Biology

Cite this

Garcia, J. G. N., Schaphorst, K. L., Shi, S., Verin, A. D., Hart, C. M., Callahan, K. S., & Patterson, C. E. (1997). Mechanisms of ionomycin-induced endothelial cell barrier dysfunction. American Journal of Physiology - Lung Cellular and Molecular Physiology, 273(1 17-1), L172-L184.