Rho and rho kinase modulation of barrier properties: Cultured endothelial cells and intact microvessels of rats and mice

R. H. Adamson, F. E. Curry, G. Adamson, B. Liu, Y. Jiang, K. Aktories, H. Barth, A. Daigeler, N. Golenhofen, W. Ness, D. Drenckhahn

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Abstract

Previus experiments using cultured endothelial monolayers indicate that Rho-family small GTPases are involved in modulation of endothelial monolayer permeability by regulating assembly of the cellular actin filament scaffold, activity of myosin-based contractility and junctional distribution of the Ca2+-dependent endothelial cell adhesion molecule, VE-cadherin. We investigated these mechanisms using both cultured endothelial cells (from porcine pulmonary artery and mouse heart) and vascular endothelium in situ (mouse aorta, and individually perfused venular microvessels of mouse and rat mesentery). Exposure to Clostridium difficile toxin B (100 ng ml-1) inactivated 50-90% of all endothelial Rho proteins within 60-90 min. This was accompanied by considerable reduction of actin filament stress fibres and junctional F-actin in cultured endothelial monolayers and in mouse aortic endothelium in situ. Also, VE-cadherin became discontinuous along endothelial junctions. Inhibition of Rho kinase with Y-27632 (30 μM) for 90-120 min induced F-actin reduction both in vitro and in situ but did not cause redistribution or reduction of VE-cadherin staining. Perfusion of microvessels with toxin B increased basal hydraulic permeability (LP) but did not attenuate the transient increase in LP of microvessels exposed to bradykinin. Perfusion of microvessels with Y-27632 (30 μM) for up to 100 min reduced basal LP but did not attenuate the permeability increase induced by platelet activating factor (PAF) or bradykinin. These results show that toxin B-mediated reduction of endothelial barrier properties is due to inactivation of small GTPases other than RhoA. Rho proteins as well as RhoA-mediated contractile mechnisms are not involved in bradykinin- or PAF-induced hyperpermeability of intact microvessels.

Original languageEnglish (US)
Pages (from-to)295-308
Number of pages14
JournalJournal of Physiology
Volume539
Issue number1
DOIs
StatePublished - Feb 15 2002

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rho-Associated Kinases
Microvessels
Cultured Cells
Endothelial Cells
Bradykinin
Permeability
Monomeric GTP-Binding Proteins
Platelet Activating Factor
Actin Cytoskeleton
Actins
Perfusion
Stress Fibers
Mesentery
Vascular Endothelium
Cell Adhesion Molecules
Myosins
Pulmonary Artery
Endothelium
Aorta
Proteins

ASJC Scopus subject areas

  • Physiology

Cite this

Adamson, R. H., Curry, F. E., Adamson, G., Liu, B., Jiang, Y., Aktories, K., ... Drenckhahn, D. (2002). Rho and rho kinase modulation of barrier properties: Cultured endothelial cells and intact microvessels of rats and mice. Journal of Physiology, 539(1), 295-308. https://doi.org/10.1113/jphysiol.2001.013117

Rho and rho kinase modulation of barrier properties : Cultured endothelial cells and intact microvessels of rats and mice. / Adamson, R. H.; Curry, F. E.; Adamson, G.; Liu, B.; Jiang, Y.; Aktories, K.; Barth, H.; Daigeler, A.; Golenhofen, N.; Ness, W.; Drenckhahn, D.

In: Journal of Physiology, Vol. 539, No. 1, 15.02.2002, p. 295-308.

Research output: Contribution to journalArticle

Adamson, RH, Curry, FE, Adamson, G, Liu, B, Jiang, Y, Aktories, K, Barth, H, Daigeler, A, Golenhofen, N, Ness, W & Drenckhahn, D 2002, 'Rho and rho kinase modulation of barrier properties: Cultured endothelial cells and intact microvessels of rats and mice', Journal of Physiology, vol. 539, no. 1, pp. 295-308. https://doi.org/10.1113/jphysiol.2001.013117
Adamson, R. H. ; Curry, F. E. ; Adamson, G. ; Liu, B. ; Jiang, Y. ; Aktories, K. ; Barth, H. ; Daigeler, A. ; Golenhofen, N. ; Ness, W. ; Drenckhahn, D. / Rho and rho kinase modulation of barrier properties : Cultured endothelial cells and intact microvessels of rats and mice. In: Journal of Physiology. 2002 ; Vol. 539, No. 1. pp. 295-308.
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AU - Adamson, R. H.

AU - Curry, F. E.

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AU - Liu, B.

AU - Jiang, Y.

AU - Aktories, K.

AU - Barth, H.

AU - Daigeler, A.

AU - Golenhofen, N.

AU - Ness, W.

AU - Drenckhahn, D.

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N2 - Previus experiments using cultured endothelial monolayers indicate that Rho-family small GTPases are involved in modulation of endothelial monolayer permeability by regulating assembly of the cellular actin filament scaffold, activity of myosin-based contractility and junctional distribution of the Ca2+-dependent endothelial cell adhesion molecule, VE-cadherin. We investigated these mechanisms using both cultured endothelial cells (from porcine pulmonary artery and mouse heart) and vascular endothelium in situ (mouse aorta, and individually perfused venular microvessels of mouse and rat mesentery). Exposure to Clostridium difficile toxin B (100 ng ml-1) inactivated 50-90% of all endothelial Rho proteins within 60-90 min. This was accompanied by considerable reduction of actin filament stress fibres and junctional F-actin in cultured endothelial monolayers and in mouse aortic endothelium in situ. Also, VE-cadherin became discontinuous along endothelial junctions. Inhibition of Rho kinase with Y-27632 (30 μM) for 90-120 min induced F-actin reduction both in vitro and in situ but did not cause redistribution or reduction of VE-cadherin staining. Perfusion of microvessels with toxin B increased basal hydraulic permeability (LP) but did not attenuate the transient increase in LP of microvessels exposed to bradykinin. Perfusion of microvessels with Y-27632 (30 μM) for up to 100 min reduced basal LP but did not attenuate the permeability increase induced by platelet activating factor (PAF) or bradykinin. These results show that toxin B-mediated reduction of endothelial barrier properties is due to inactivation of small GTPases other than RhoA. Rho proteins as well as RhoA-mediated contractile mechnisms are not involved in bradykinin- or PAF-induced hyperpermeability of intact microvessels.

AB - Previus experiments using cultured endothelial monolayers indicate that Rho-family small GTPases are involved in modulation of endothelial monolayer permeability by regulating assembly of the cellular actin filament scaffold, activity of myosin-based contractility and junctional distribution of the Ca2+-dependent endothelial cell adhesion molecule, VE-cadherin. We investigated these mechanisms using both cultured endothelial cells (from porcine pulmonary artery and mouse heart) and vascular endothelium in situ (mouse aorta, and individually perfused venular microvessels of mouse and rat mesentery). Exposure to Clostridium difficile toxin B (100 ng ml-1) inactivated 50-90% of all endothelial Rho proteins within 60-90 min. This was accompanied by considerable reduction of actin filament stress fibres and junctional F-actin in cultured endothelial monolayers and in mouse aortic endothelium in situ. Also, VE-cadherin became discontinuous along endothelial junctions. Inhibition of Rho kinase with Y-27632 (30 μM) for 90-120 min induced F-actin reduction both in vitro and in situ but did not cause redistribution or reduction of VE-cadherin staining. Perfusion of microvessels with toxin B increased basal hydraulic permeability (LP) but did not attenuate the transient increase in LP of microvessels exposed to bradykinin. Perfusion of microvessels with Y-27632 (30 μM) for up to 100 min reduced basal LP but did not attenuate the permeability increase induced by platelet activating factor (PAF) or bradykinin. These results show that toxin B-mediated reduction of endothelial barrier properties is due to inactivation of small GTPases other than RhoA. Rho proteins as well as RhoA-mediated contractile mechnisms are not involved in bradykinin- or PAF-induced hyperpermeability of intact microvessels.

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