Rho-kinase signalling regulates CXC chemokine formation and leukocyte recruitment in colonic ischemia-reperfusion.
OBJECTIVE
Leukocyte recruitment is a key feature in ischemia-reperfusion (I/R)-induced tissue injury. The aim of the present study was to investigate the effect of Rho-kinase inhibition on I/R-provoked leukocyte recruitment in the colon.


METHODS
C57BL/6 mice were subjected to 30 min of ischemia by clamping of the superior mesenteric artery followed by 120 min of reperfusion. Intraperitoneal pretreatment with the selective Rho-kinase inhibitors fasudil (4-40 mg/kg) and Y-27632 (1-10 mg/kg) was administered prior to induction of colonic I/R. Leukocyte-endothelium interactions were analyzed by intravital fluorescence microscopy. Colonic content of tumour necrosis factor-alpha (TNF-alpha) and the CXC chemokines macrophage inflammatory protein-2 (MIP-2) and cytokine-induced neutrophil chemoattractant (KC) were determined by ELISA. Additionally, colonic activity of myeloperoxidase (MPO), a marker of leukocyte infiltration, and malondialdehyde (MDA), were quantified.


RESULTS
Fasudil and Y-27632 pretreatment decreased I/R-induced leukocyte rolling and adhesion by 76% and 96%, respectively. Moreover, Rho-kinase interference reduced formation of TNF-alpha, MIP-2 and KC by more than 68% in the reperfused colon. Additionally, the reperfusion-provoked increase in the levels of MPO and MDA in the colon decreased after Rho-kinase inhibition by 69% and 42%, respectively.


CONCLUSIONS
Our data demonstrate that inhibition of Rho-kinase activity decrease I/R-induced leukocyte rolling, adhesion and recruitment in the colon. Moreover, these findings show that Rho-kinase signalling regulates TNF-alpha and CXC chemokine formation as well as lipid peroxidation in the reperfused colon. Thus, targeting Rho-kinase signalling may be a useful strategy in order to protect against pathological inflammation in the colon.
Santen
Stefan
S
Department of Surgery, Malmö University Hospital, Lund University, 205 02 Malmö, Sweden. stefan.santen@skane.se
Wang
Yusheng
Y
Laschke
Matthias W
MW
Menger
Michael D
MD
Jeppsson
Bengt
B
Thorlacius
Henrik
H
D000818:Animals; D002448:Cell Adhesion / Q000187:drug effects; D002465:Cell Movement / Q000187:drug effects; D054360:Chemokine CXCL1 / Q000096:biosynthesis / Q000378:metabolism; D054426:Chemokine CXCL2 / Q000096:biosynthesis; D019743:Chemokines, CXC / Q000096:biosynthesis*; D003106:Colon / Q000187:drug effects / Q000201:enzymology / Q000473:pathology*; D019040:E-Selectin / Q000378:metabolism; D006439:Hemodynamics / Q000187:drug effects; D007958:Leukocyte Count; D036904:Leukocyte Rolling / Q000187:drug effects; D007962:Leukocytes / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D008315:Malondialdehyde / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D019007:P-Selectin / Q000378:metabolism; D009195:Peroxidase / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D015427:Reperfusion Injury / Q000097:blood / Q000201:enzymology* / Q000473:pathology / Q000503:physiopathology; D015398:Signal Transduction / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D054360:Chemokine CXCL1; D054426:Chemokine CXCL2; D019743:Chemokines, CXC; C475895:Cxcl1 protein, mouse; D019040:E-Selectin; D019007:P-Selectin; D047428:Protein Kinase Inhibitors; D014409:Tumor Necrosis Factor-alpha; D008315:Malondialdehyde; D009195:Peroxidase; D054460:rho-Associated Kinases