Protective role of PI3-kinase-Akt-eNOS signalling pathway in intestinal injury associated with splanchnic artery occlusion shock.
OBJECTIVE
Endothelial NO synthase (eNOS) is a dynamic enzyme tightly controlled by co- and post-translational lipid modifications, phosphorylation and regulated by protein-protein interactions. Here we have pharmacologically modulated the activation of eNOS, at different post-translational levels, to assess the role of eNOS-derived NO and of these regulatory mechanisms in intestinal injury associated with splanchnic artery occlusion (SAO) shock.


METHODS
SAO shock was induced by clamping both the superior mesenteric artery and the celiac trunk for 45 min followed by 30 min of reperfusion. During ischemia, 15 min prior to reperfusion, mice were given geldanamycin, an inhibitor of hsp90 recruitment to eNOS, or LY-294002 an inhibitor of phosphatidylinositol 3-kinase (PI3K), an enzyme that initiates Akt-catalysed phosphorylation of eNOS on Ser1179. After 30 min of reperfusion, samples of ileum were taken for histological examination or for biochemical studies.


RESULTS
Either LY-294002 or geldanamycin reversed the increased activation of eNOS and Akt observed following SAO shock. These molecular effects were mirrored in vivo by an exacerbation of the intestinal damage. Histological damage also correlated with neutrophil infiltration, assessed as myeloperoxidase activity, and with an increased expression of the adhesion proteins: ICAM-I, VCAM, P-selectin and E-selectin.


CONCLUSIONS
Overall these results suggest that activation of the Akt pathway in ischemic regions of reperfused ileum is a protective event, triggered in order to protect the intestinal tissue from damage induced by ischaemia/reperfusion through a fine tuning of the endothelial NO pathway.
Roviezzo
F
F
Department of Experimental Pharmacology, University of Napoli, Federico II, Napoli, Italy.
Cuzzocrea
S
S
Di Lorenzo
A
A
Brancaleone
V
V
Mazzon
E
E
Di Paola
R
R
Bucci
M
M
Cirino
G
G
D000818:Animals; D001157:Arterial Occlusive Diseases / Q000503:physiopathology; D016227:Benzoquinones / Q000494:pharmacology; D015153:Blotting, Western; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002867:Chromones / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007082:Ileum / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D007422:Intestines / Q000098:blood supply / Q000293:injuries / Q000503:physiopathology*; D047029:Lactams, Macrocyclic / Q000494:pharmacology; D008297:Male; D051379:Mice; D009025:Morpholines / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000473:pathology; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D009195:Peroxidase / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation / Q000187:drug effects; D011499:Protein Processing, Post-Translational; D051057:Proto-Oncogene Proteins c-akt / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015427:Reperfusion Injury / Q000503:physiopathology; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D013152:Splanchnic Circulation / Q000502:physiology
D016227:Benzoquinones; D015815:Cell Adhesion Molecules; D002867:Chromones; D004791:Enzyme Inhibitors; D047029:Lactams, Macrocyclic; D009025:Morpholines; D052250:Nitric Oxide Synthase Type III; D009195:Peroxidase; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051057:Proto-Oncogene Proteins c-akt; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C001277:geldanamycin