Hepatic apoptotic activity following transient normothermic inflow occlusion and reperfusion in the swine model

Accelerated hepatic apoptosis was first described in portal vein-ligated li vers but has since been reported in a variety of liver injuries. Because po rto-prival states can induce apoptosis we sought to determine whether trans ient ischemic periods followed by reperfusion would trigger such cell death . The cytokines TNF-alpha and TGF-beta are known to facilitate apoptosis an d are released in response to a number of stimuli including ischemia. We al so investigated alterations in plasma and tissue levels of these cytokines which might lend support to their role in increased apoptotic activity foll owing ischemia/reperfusion. Female pigs were used as the experimental model . Inflow occlusion of portal and hepatic arterial blood was performed to a portion of the swine liver directing the entire splanchnic flow to the rema ining hepatic lobes for a period of 2 h. The livers mere then reperfused an d plasma and tissue samples taken for determination of apoptotic activity u tilizing cell death immunoperoxidase staining of 3'-OH DNA ends generated b y fragmentation and ELISA assay of histone-associated DNA fragments. Plasma and tissue levels of TNF-alpha and plasma levels of TGF-beta were determin ed by ELISA assay. An increase in apoptotic activity following reperfusion was seen at Day 2 and Day 4 compared to preischemic values by the cell deat h stain. The ELISA cell death assay showed an increase in apoptotic activit y at 60 min, Day 2, and Day 4. Moreover, the ELISA cell death assay showed enhanced apoptotic activity in "hyperperfused" hepatic lobes compared to pr eischemic, or resting, liver. This was also observed when compared to sham- operated animals. Surprisingly, there was no detectable increase in plasma TNF-alpha or TGF-beta levels following ischemia/reperfusion, although homog enized liver TNF-alpha levels were increased at 60 min and Day 2 following reperfusion. We conclude that transient hepatic inflow occlusion followed b y reperfusion can induce increased apoptotic activity in the swine model. F urthermore, increased apoptotic activity also occurs in the hyperperfused l iver raising the possibility of a locally active factor or global hepatic e xpression of receptor activity in response to ischemia/reperfusion. This pe riod of ischemia/reperfusion did not produce a detectable increase in circu lating cytokine levels, and accelerated apoptosis could not be linked to he ightened TNF-alpha or TGF-beta plasma activity. Higher tissue levels of TNF -alpha could reflect enhanced binding to TNF cell surface receptors or ampl ified receptor expression. (C) 1999 Academic Press.