Donor Pretreatment with Tetrahydrobiopterin Saves Pancreatic Isografts from Ischemia Reperfusion Injury in a Mouse Model

https://doi.org/10.1111/j.1600-6143.2010.03262.xGet rights and content
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Depletion of the nitric oxide synthase cofactor tetrahydrobiopterin (H4B) during ischemia and reperfusion is associated with severe graft pancreatitis. Since clinically feasible approaches to prevent ischemia reperfusion injury (IRI) by H4B‐substitution are missing we investigated its therapeutic potential in a murine pancreas transplantation model using different treatment regimens. Grafts were subjected to 16 h cold ischemia time (CIT) and different treatment regimens: no treatment, 160 μM H4B to perfusion solution, H4B 50 mg/kg prior to reperfusion and H4B 50 mg/kg before recovery of organs. Nontransplanted animals served as controls. Recipient survival and endocrine graft function were assessed. Graft microcirculation was analyzed 2 h after reperfusion by intravital fluorescence microscopy. Parenchymal damage was assessed by histology and nitrotyrosine immunohistochemistry, H4B tissue levels by high pressure liquid chromatography (HPLC). Compared to nontransplanted controls prolonged CIT resulted in significant microcirculatory deterioration. Different efficacy according to route and timing of administration could be observed. Only donor pretreatment with H4B resulted in almost completely abrogated IRI‐related damage showing graft microcirculation comparable to nontransplanted controls and restored intragraft H4B levels, resulting in significant reduction of parenchymal damage (p < 0.002) and improved survival and endocrine function (p = 0.0002 each). H4B donor pretreatment abrogates ischemia‐induced parenchymal damage and represents a promising strategy to prevent IRI following pancreas transplantation.

Key words:

Animal models
donor preconditioning
endothelial nitric oxide synthase
endothelial dysfunction
ischemia/reperfusion injury
mice
nitric oxide
oxidative stress
pancreas transplantation

Abbreviations:

b.w
body weight
CD
capillary diameter
CIT
cold ischemia time
DAB
diamonbenzidinetetrahydrochloride
FCD
functional capillary density
H4B
tetrahydrobiopterin
HCl
Hydrochloric Acid
HPLC
high pressure liquid chromatography
HTK
Histidine‐Tryptophane‐Ketoglutarate
i.d.
inner diameter
ICH
immunohistochemistry
i.m.
intramuscular
IRI
ischemia reperfusion injury
IVFM
intravital fluorescence microscopy
MW
molecular weight
NaOH
Sodium hydroxide
NO
nitric oxide
(e)(i)(n)NOS
(endothelial)(inducible)(neuronal) nitric oxide synthase
ONOO‐
peroxynitrite
PBS
Dulbecco's Phosphate buffered saline
p.o.
post operationem
SDS‐PAGE
sodium dodecyl sulfate poly‐acrylamide gel electrophoresis
WIT
warm ischemia time

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