Hepatocyte cytoskeleton during ischemia and reperfusion--influence of ANP-mediated p38 MAPK activation.
OBJECTIVE
To determine functional consequences of this activation, whereby we focused on a potential regulation of the hepatocyte cytoskeleton during ischemia and reperfusion.


METHODS
For in vivo experiments, animals received ANP (5 microg/kg) intravenously. In a different experimental setting, isolated rat livers were perfused with KH-buffer+/-ANP (200 nmol/L) +/-SB203580 (2 micromol/L). Livers were then kept under ischemic conditions for 24 h, and either transplanted or reperfused. Actin, Hsp27, and phosphorylated Hsp27 were determined by Western blotting, p38 MAPK activity by in vitro phosphorylation assay. F-actin distribution was determined by confocal microscopy.


RESULTS
We first confirmed that ANP preconditioning leads to an activation of p38 MAPK and observed alterations of the cytoskeleton in hepatocytes of ANP-preconditioned organs. ANP induced an increase of hepatic F-actin after ischemia, which could be prevented by the p38 MAPK inhibitor SB203580 but had no effect on bile flow. After ischemia untreated livers showed a translocation of Hsp27 towards the cytoskeleton and an increase in total Hsp27, whereas ANP preconditioning prohibited translocation but caused an augmentation of Hsp27 phosphorylation. This effect is also mediated via p38 MAPK, since it was abrogated by the p38 MAPK inhibitor SB203580.


CONCLUSIONS
This study reveals that ANP-mediated p38 MAPK activation leads to changes in hepatocyte cytoskeleton involving an elevation of phosphorylated Hsp27 and thereby for the first time shows functional consequences of ANP-induced hepatic p38 MAPK activation.
Keller
Melanie
M
Department of Pharmacy, Centre of Drug Research, University of Munich, Germany.
Gerbes
Alexander L
AL
Kulhanek-Heinze
Stefanie
S
Gerwig
Tobias
T
Grutzner
Uwe
U
van Rooijen
Nico
N
Vollmar
Angelika M
AM
Kiemer
Alexandra K
AK
D000818:Animals; D009320:Atrial Natriuretic Factor / Q000494:pharmacology*; D003599:Cytoskeleton / Q000187:drug effects; D004789:Enzyme Activation / Q000187:drug effects; D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000378:metabolism; D022781:Hepatocytes / Q000201:enzymology* / Q000473:pathology; D019194:Ischemic Preconditioning; D008297:Male; D009363:Neoplasm Proteins / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury / Q000378:metabolism* / Q000473:pathology; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins; C486114:Hspb1 protein, rat; D009363:Neoplasm Proteins; D009320:Atrial Natriuretic Factor; D048051:p38 Mitogen-Activated Protein Kinases