Nitric oxide in early ischaemia reperfusion injury during human orthotopic liver transplantation.
BACKGROUND
Altered nitric oxide (NO) metabolism has been shown to contribute to ischemia-reperfusion (IR) injury in animal models. However, similar studies have not been performed in human liver transplantation (LT). In this study, we examined nitrate, nitrite, and nitrosothiols (NOx), NO synthases (endothelial [constitutive] nitric oxide synthase [eNOS] and inducible nitric oxide synthase [iNOS]), and nitrotyrosine in early IR injury after human LT.


METHODS
Paired biopsies were obtained from nine donor livers before cold ischemia (retrieval biopsy) and after reimplantation (reperfusion biopsy). Sections were graded for reperfusion injury using the Suzuki score. NO was detected by chemiluminescence after reduction of NOx. Expression of eNOS and iNOS was by Western blot and reverse transcriptase polymerase chain reaction and peroxynitrite by immunodetection of 3-nitrotyrosine.


RESULTS
Reperfusion biopsies showed histologic evidence of injury (median Suzuki score: retrieval 2, reperfusion 6, P=0.008) and neutrophil infiltration. NOx was reduced after reperfusion from 5.41 microM/100 mg (median, range 2.17-13.39 microM) to 3.51 microM (1.45-5.66 microM, P=0.05). eNOS protein was reduced after reperfusion from 0.6 units (median, range 0.45-1 unit) in retrieval biopsies to 0.39 units in reperfusion biopsies (range 0.2-0.79 units, P=0.007). There was no change in eNOS or iNOS mRNA expression or iNOS protein. Western blotting showed increased nitrotyrosine formation after reperfusion, median 0.42 (range 0.16-0.87) units in retrieval biopsies and 0.68 (0.29-1.06) units in reperfusion samples (P=0.007) and localized to periportal regions.


CONCLUSIONS
iNOS protein may not contribute to early reperfusion injury during human LT. However, reduced NO bioavailability caused by reduced eNOS may contribute significantly to damage at this time point.
Department of Surgery, Education and Research Centre, St. Vincent's University Hospital, Elm Park, Dublin 4, Ireland.
Raghu
R
Varadarajan
Lucy
L
Golden-Mason
Leonie
L
Young
Paul
P
McLoughlin
Niamh
N
Nolan
Gerry
G
McEntee
Oscar
O
Traynor
Justin
J
Geoghegan
John E
JE
Hegarty
Cliona
C
O'Farrelly
D000293:Adolescent; D000328:Adult; D000368:Aged; D002102:Cadaver; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D016031:Liver Transplantation / Q000009:adverse effects* / Q000473:pathology; D008297:Male; D008875:Middle Aged; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III; D012333:RNA, Messenger / Q000235:genetics; D012086:Reoperation; D015427:Reperfusion Injury / Q000209:etiology / Q000473:pathology / Q000503:physiopathology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014019:Tissue Donors / Q000706:statistics & numerical data; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000378:metabolism
D012333:RNA, Messenger; D009569:Nitric Oxide; C002744:3-nitrotyrosine; D014443:Tyrosine; C496319:NOS2 protein, human; C496314:NOS3 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D052250:Nitric Oxide Synthase Type III