Role of calmodulin and myosin light-chain kinase in lung ischemia/
reperfusion injury.
Khimenko, Pavel L., Timothy M. Moore, Peter S. Wilson, and Aubrey E.
Taylor.
Department of Physiology, College of Medicine, University of South
Alabama, Mobile, Alabama 36688
APStracts 3:0042L, 1996.
It is generally accepted that microvascular permeability is controlled
by intercellular endothelial cell gap size. This process is
controlled in endothelial cell monolayers and peripheral blood
vessels by calmodulin (CaM)-dependent myosin light-chain kinase
(MLCK) which phosphorylates myosin light chain (MLC20) with
subsequent actin-myosin interaction. In the present study both CaM
and MLCK blockers were studied during ischemia and reperfusion (I/R)
-induced injury in isolated buffer perfused rat lungs. The effects of
a calcium ionophore (CaI) were tested in isolated intact rat lungs in
order compare the effects of increasing intracellular Ca2+ to I/R
-induced damage. Because protein kinase C (PKC) could also be a
mediator of I/R injury, a PKC inhibitor was studied in lungs
subjected to either I/R or CaI. In lungs subjected to I/R alone, a
five-fold increase in microvascular permeability occurred after 30
minutes of reperfusion (p<0.001) and a ten-fold increase was
present after an additional 60 minutes of reperfusion (p<0.01).
Pretreatment of the I/R lungs with a CaM inhibitor (trifluoperazine,
100 [mu]M) or with a MLCK inhibitor (ML-7, 500nM) blocked the
microvascular damage at both 30 minutes and 90 minutes of
reperfusion. When the CaM inhibitor was introduced into the venous
reservoir after 46 minutes of reperfusion, after the microvascular
damage was present, no further increase in microvascular permeability
occurred. Pretreatment of the lungs with a PKC inhibitor
(staurosporine, 100 nM) did not alter the magnitude of the increased
microvascular permeability produced by I/R or the time course of the
damage. The calcium ionophore (A23187, 7.5 [mu]M) caused increases in
Kfcs similar to those produced by I/R. Pretreatment of A23187 treated
lungs with a CaM inhibitor produced no protective effect on the
microvascular injury at 30 minutes after administration. Pretreatment
of the CaI-challenged lungs with staurosporine significantly
increased the microvascular barrier injury at 30 minutes as compared
to that occurring with I/R. When a [beta]-adrenergic receptor agonist
(isoproterenol, 10 [mu]M) was introduced to the lung after CaI
-induced damage had occurred, no further increase in microvascular
permeability was observed and a trend towards reversal of injury
occurred. We conclude from these studies that CaM/MLCK/MLC20 system
is involved in our model of I/R-induced rat lung injury but is not
involved in lung injury associated with Ca2+ entering the cell.
Received 3 November 1995; accepted in final form 5 March 1996.
APS Manuscript Number L316-5.
Article publication pending Am. J. Physiol. (Lung Cell. Mol.
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 27 March 96