Breakdown of hepatic tight junctions during reoxygenation injury.
We investigated whether reoxygenation following anoxia increased biliary permeability and whether or not allopurinol had a protective effect. Isolated rat livers were perfused for 30 min in a one-pass system with buffer equilibrated with 100% nitrogen after stabilization, and then for 60 min with the oxygenated buffer. Hepatic tight junction permeability was assessed by quantifying the early appearance in the bile of horseradish peroxidase (HRP) injected with the perfusate. This early peak represents paracellular passage of HRP, whereas a later second peak results from transcellular passage. In the control livers, 7% of the total HRP passage (93 +/- 50 pg/g liver) was paracellular and 93% was transcellular. After 30 min of reoxygenation following anoxia, however, 516 +/- 20 pg/g liver of HRP passed paracellularly. Addition of allopurinol (5 micrograms/ml) to the perfusate from the start of perfusion reduced paracellular passage of HRP to 219 +/- 49 pg/g liver after anoxia and reperfusion (P less than 0.01). Allopurinol also reduced the cumulative lactate dehydrogenase (LDH) release during the first 30 min of reoxygenation from 2.1 +/- 0.3 x 10(4) to 1.4 +/- 0.4 x 10(4) units/g liver (P less than 0.01). Reduction of the anoxic period from 30 min to 25 min significantly reduced the change in tight junction permeability and the extent of cellular injury: Paracellular passage of HRP was 336 +/- 20 pg/g and LDH release was 0.7 +/- 0.1 x 10(4) units/g liver, both significantly lower than those at 30 min (P less than 0.01). No significant difference in hepatic ATP levels after 60 min of reoxygenation was noted among the experimental groups, but all had lower levels than the control group. The protective effect of allopurinol suggests that the mechanism of biliary reoxygenation injury involves free radical generation. Susceptibility of tight junctions suggests a pattern of injury similar to that involved in anoxic damage of the vascular endothelium.
Konno
H
H
Second Department of Surgery, Hamamatsu University School of Medicine, Japan.
Lowe
P J
PJ
Hardison
W G
WG
Miyai
K
K
Nakamura
S
S
Baba
S
S
D000255:Adenosine Triphosphate / Q000032:analysis; D000493:Allopurinol / Q000494:pharmacology; D000818:Animals; D002463:Cell Membrane Permeability / Q000187:drug effects; D006735:Horseradish Peroxidase / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008099:Liver / Q000098:blood supply* / Q000737:chemistry / Q000201:enzymology; D008297:Male; D010101:Oxygen Consumption; D051381:Rats; D011919:Rats, Inbred Strains; D015427:Reperfusion Injury / Q000503:physiopathology*
D000493:Allopurinol; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase; D006735:Horseradish Peroxidase