Evaluation of purine nucleoside phosphorylase release as a measure of hepatic endothelial cell injury.
With emerging data that endothelial cell (EC) injury is the limiting factor in liver preservation and hepatic function, a simple and reliable biochemical technique for monitoring EC injury is needed. Measurement of purine nucleoside phosphorylase (PNP) release into the circulation from perfused liver has been proposed as such a method. However, our experiments with perfused rat liver did not display a clear or direct relationship between PNP release and endothelial cell injury. Therefore, we re-examined the suitability of using PNP as a measure of nonparenchymal injury by measuring its distribution in purified populations of hepatocytes, ECs, and Kupffer cells (KCs) and correlating cell injury and enzyme release in short-term cultures at 37 degrees C of each cell type. Purified cells were incubated (4 x 10(6) cells/mL) in oxygen or nitrogen saturated. Wisconsin solution or Krebs buffer for 6 hours, with cell viability and PNP release assayed every 2 hours. ECs had the lowest specific activity (27 +/- 9 U/mg protein; mean +/- standard error of the mean [SEM]) compared with both hepatocytes (115 +/- 15) and KCs (66 +/- 18). Despite a decrement in EC and KC viability over time in each incubation solution, there was poor correlation between time of incubation and PNP release (r = .01 to .22), and between cell viability and PNP release (r = .01 to .16). In contrast, PNP release from incubated hepatocytes correlated with the length of incubation (r = .57 to .78) as well as cell injury (r = .63 to .77) in all four test solutions.(ABSTRACT TRUNCATED AT 250 WORDS)
Brass
C A
CA
Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia.
Mody
M G
MG
D000241:Adenosine / Q000494:pharmacology; D000493:Allopurinol / Q000494:pharmacology; D000818:Animals; D002021:Buffers; D002469:Cell Separation; D004727:Endothelium / Q000378:metabolism / Q000473:pathology; D005978:Glutathione / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology; D007552:Isotonic Solutions / Q000494:pharmacology; D007728:Kupffer Cells / Q000378:metabolism; D008099:Liver / Q000187:drug effects / Q000473:pathology*; D008297:Male; D019188:Organ Preservation Solutions; D011683:Purine-Nucleoside Phosphorylase / Q000378:metabolism*; D011887:Raffinose / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D014018:Tissue Distribution
D002021:Buffers; D007328:Insulin; D007552:Isotonic Solutions; C051865:Krebs-Ringer solution; D019188:Organ Preservation Solutions; C055206:University of Wisconsin-lactobionate solution; D000493:Allopurinol; D011683:Purine-Nucleoside Phosphorylase; D005978:Glutathione; D000241:Adenosine; D011887:Raffinose