EVALUATION OF PURINE NUCLEOSIDE PHOSPHORYLASE RELEASE AS A MEASURE OFHEPATIC ENDOTHELIAL-CELL INJURY

With emerging data that endothelial cell (EC) injury is the limiting f actor in Liver preservation and hepatic function, a simple and reliabl e biochemical technique for monitoring EC injury is needed. Measuremen t of purine nucleoside phosphorylase (PNP) release into the circulatio n 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. Therefo re, we re-examined the suitability of using PNP as a measure of nonpar enchymal injury by measuring its distribution in purified populations of hepatocytes, ECs, and Kupffer cells (KCs) and correlating cell inju ry and enzyme release in short-term cultures at 37 degrees C of each c ell 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 th e lowest specific activity (27 +/- 9 U/mg protein; mean +/- standard e rror of the mean [SEM]) compared with both hepatocytes (115 +/- 15) an d KCs (66 +/- 18). Despite a decrement in EC and KC viability over tim e in each incubation solution, there was poor correlation between time of incubation and PNP release (r = .01 to .22), and between cell viab ility and PNP release (r = .01 to .16). In contrast, PNP release from incubated hepatocytes correlated with the length of incubation (r = .5 7 to .78) as well as cell injury (r = .63 to .77) in all four test sol utions. This data suggest that PNP release is unlikely to specifically reflect EC injury in the intact liver.