Suzanne M. Wheeler

V-PYRRO/NO: an hepato-selective nitric oxide donor improves porcine liver hemodynamics and function after ischemia reperfusion.

Background. explored theThe role of nitric oxide (NO) in ischemia reperfusion (I/R) injury is controversial as both beneficial and harmful effects have been reported. We explored the potential role of a pharmacological agent recently shown to generate NO metabolically in the liver in an animal model of transplantation. Methods. The effect of a selective hepatic NO donor, O2-vinyl 1-(pyrrolidin-1-yl)diazen-1-ium-1,2-diolate (V-PYRRO/NO), on hepatic hemodynamics and biliary function was evaluated in both the in situ and I/R pig liver. Results. V-PYRRO/NO significantly reduced in situ hepatic vascular resistance (HVR) without altering systolic blood pressure. Portal vein flow was essentially unchanged during in situ infusions while hepatic artery flow nearly doubled (P =0.03). After I/R, V-PYRRO/NO infusions significantly reduced both portal vein pressure (PVP) and HVR (P= 0.04). Also, serum bile acid clearance increased from 15% when taurocholate (TC) was infused alone to 46% (P =0.007) when infused simultaneously with V-PYRRO/NO. Aqueous bile production tripled with TC and V-PYRRO/NO as compared to TC alone (P= 0.04). Analysis of bile outputs revealed a significant increase in biliary cholesterol, biliary phospholipid, and biliary bile acid (P <0.05) with V-PYRRO/NO infusion. Conclusions. The hepato-selective nitric oxide donor, V-PYRRO/NO, reduced hepatic resistance parameters of the pig liver both before and after I/R and improved the plasma clearance of bile acid and biliary outputs of bile acid-dependent compounds. The augmented function observed after I/R may be due to improvements in hepatic blood flow secondary to altered hepatic hemodynamics.

Protective effects of ischemic preconditioning on the cold-preserved liver are tyrosine kinase dependent.

Background. Little data exist regarding the use of ischemic preconditioning before sustained hepatic cold storage. We hypothesized that ischemic preconditioning protects hepatic grafts via a tyrosine kinase–dependent pathway. Methods. Six porcine livers underwent routine harvest (control). Five other livers underwent 15 min of in situ ischemia followed by 15 min of reflow before harvest (ischemic preconditioning). Another five livers were pretreated with a tyrosine kinase inhibitor (genistein) before preconditioning. Upon reperfusion and after 2 hours of cold storage, graft function, graft circulatory impairment, and markers of cellular damage were analyzed. Tissue cytoplasmic extracts were analyzed for tyrosine phosphorylation with Western blot. Significance was determined with t tests. Results. Ischemic-preconditioned grafts demonstrated enhanced bile production, augmented responses to a bile acid challenge, and elevated O2 consumption (P <0.05) compared to controls. Also, preconditioned grafts demonstrated improved hepatic tissue blood flow and decreased hepatic vascular resistance (P <0.005) compared to controls. Endothelial cell preservation (factor VIII immunostain) was improved in preconditioned graft biopsies compared to controls. With genistein pretreatment, all observed improvements returned to control levels. Analysis of cytoplasmic extracts demonstrated an increase in tyrosine phosphorylation before cold ischemia in preconditioned grafts only, but not in control or genistein-pretreated grafts. Conclusions. The data indicate that ischemic preconditioning protects the liver from sustained cold ischemia and that tyrosine kinases are involved in preconditioning responses.

Regulation of NFκB in hepatic ischemic preconditioning1

Abstract BACKGROUND: The second messengers tyrosine kinase (TK) and protein kinase C (PKC) have been implicated in mediating the cellular signaling cascade during hepatic ischemic preconditioning (IPC). We evaluated the role of TK and PKC on the modulation of the transcription factor nuclear factor kappa B (NFκB) and its inhibitor IκBα during IPC. STUDY DESIGN: Yorkshire pigs underwent routine harvest. IPC livers underwent 15 minutes of ischemia and 15 minutes of in situ perfusion before harvest, with or without pretreatment with a TK inhibitor (genistein) or a PKC inhibitor (chelerythrine). During cold storage and reperfusion, tissue extracts were analyzed for IκBα phosphorylation and NFκB levels and for TK and PKC activity by Western blot. RESULTS: Control pig livers demonstrated no change in the levels of TK, PKC, IκBα, or NFκB before cold ischemia. IPC grafts demonstrated activation of TK and PKC with increased IκBα phosphorylation and NFκB levels before cold ischemia. IPC grafts pretreated with genistein demonstrated inhibition of TK activation but not of PKC activation. Genistein-pretreated grafts also demonstrated inhibition of IκBα phosphorylation and a lack of NFκB translocation to the nucleus throughout the entire experiment. IPC grafts pretreated with chelerythrine demonstrated inhibition of PKC activation but not TK activation. Chelerythrine-pretreated grafts also demonstrated IκBα phosphorylation before cold ischemia and enhanced nuclear levels of NFκB. CONCLUSIONS: Data suggest that the role of TK in IPC might be mediated in part by NFκB, but PKC does not depend on NFκB for its effect. Two parallel signaling pathways might explain these data.

Effect of hepatic artery flow on bile secretory function after cold ischemia.

These studies evaluated the influence of hepatic arterial flow on biliary secretion after cold ischemia. Preparation of livers for transplantation or hepatic support impairs biliary secretion. The earliest indication of cold preservation injury during reperfusion is circulatory function. Arterial flow at this time may be critical for bile secretion.

Elevated intrahepatic pressures and decreased hepatic tissue blood flow prevent gas embolus during limited laparoscopic liver resections

Background: As new techniques are emerging for laparoscopic liver resections, concerns have been raised about the development of gas embolus related to the CO2 pneumoperitoneum. We hypothesized that elevated intrahepatic vascular pressures and decreased hepatic tissue blood flow (LQB) would prevent gas embolus during laparoscopic liver resections under conventional pneumoperitoneum. Methods: Intrahepatic vascular pressures and LQB were measured in nine pigs with varying CO2 pneumoperitoneum. Gas embolus was determined after hepatic incision by monitoring pulmonary arterial pressure (PAP), hepatic venous PCO2, systemic blood pressure (SBP), and suprahepatic vena cava ultrasound. Results: As the pneumoperitoneum was increased from 0 to 15 mmHg, intrahepatic vascular pressures increased significantly (p < 0.05), while LQB decreased significantly (p < 0.05). A 2.0-cm hepatic incision at 4, 8, 15, and 20mmHg produced no ultrasound evidence of gas embolus and no changes in PAP, SBP, or hepatic venous PCO2 (p = NS). Conclusion: These data suggest that the risk of significant embolus under conventional pneumoperitoneum is minimal during laparoscopic liver resections.

Bosentan, an endothelin antagonist, augments hepatic graft function by reducing graft circulatory impairment following ischemia/reperfusion injury.

Endothelin is a potent hepatic vasoconstrictor. We evaluated the role of an endothelin antagonist in hepatic ischemia/reperfusion injury. Bosentan, a novel endothelin receptor antagonist, was infused directly into the portal vein prior to cold ischemia and immediately on reperfusion, in five porcine livers. Five other pigs underwent routine liver harvest and reperfusion without bosentan treatment. Hepatic vascular resistance and liver tissue blood flow, as measured by thermistor flow probes, were determined following reperfusion. Hepatocellular damage was assessed through hepatic venous levels of sorbitol dehydrogenase and lactate dehydrogenase. Endothelial cell damage was determined in sections immunostained for factor VIII. Graft function was determined through oxygen consumption, bile production, and response to bile acid challenge. Organs treated with bosentan demonstrated lower vascular resistance and enhanced tissue blood flow (P <0.05) as compared to untreated organs. Portal vein inflow to hepatic tissue was significantly enhanced (4.4-fold) in the bosentan-treated organs (P <0.05). No difference was observed in hepatocellular damage. Pathology scores for factor VIII immunohistochemical staining were 2.3-fold higher in the bosentan-treated livers as compared to untreated livers (P <0.05). The bosentantreated livers also demonstrated enhanced oxygen consumption, increased bile production, and augmented biliary response to a bile acid challenge (P <0.05). These results indicate that administration of bosentan before and after ischemia/reperfusion reduces hepatic circulatory disturbances, diminishes endothelial cell damage, and augments hepatic graft function.

Donor Hepatic Function ☆: A Factor in Postreperfusion Syndrome

Reperfusion of support livers after cold preservation produces hemodynamic instability (i.e., postreperfusion syndrome) in the recipient during both orthotopic liver transplantation and extracorporeal liver perfusion. We evaluated the effect of the normal porcine cold-preserved support liver on healthy recipient hemodynamics and in situ liver function during extracorporeal liver perfusion. Support livers were harvested from Yorkshire pigs and reperfused in an extracorporeal circuit with a healthy, anesthetized recipient pig. Correlation analyses were performed between support liver variables of function (oxygen consumption, bile flow, and biliary phospholipid and cholesterol output) and both recipient hemodynamic stability (heart rate, blood pressure, urine output, and vasopressor use) and hepatic function (bile flow and biliary phospholipid secretion). The data indicate that optimally functioning support livers are associated with improved recipient hemodynamic stability manifested by decreased recipient heart rate and vasopressor use and increased recipient urine output. Support livers exhibiting poor biliary secretory function (i.e., bile flow and phospholipid output) were associated with similarly diminished recipient liver biliary secretory function. These data indicate that the functional condition of the support liver after harvest and cold preservation may influence both recipient hemodynamic parameters and the endogenous function of the recipient liver. (J Gastrointest Surg 2002;6:248-254.)

Alterations in intrahepatic hemodynamics of the harvested porcine liver

Hemodynamic properties of a donor liver, during initial reperfusion, are associated with the degree of graft preservation injury and have been proposed to correlate with subsequent markers of liver function. In the present study, hepatic hemodynamics, that is, portal venous pressure, hepatic vascular resistance, and compliance (vascular distensibility), were characterized (1) in situ before porcine livers were manipulated, (2) after these same livers were isolated and perfused within a bypass circuit, and (3) on reperfusion after 2 hours of cold ischemia. Hepatic vascular resistance was determined in each of these three states from the portal vein pressure response to differing hepatic blood flows. In addition, the response of the same livers to norepinephrine and nitroprusside was evaluated in each condition. In the in situ and isolated perfused liver, portal venous pressure increased only modestly despite doubling of hepatic flows. After cold ischemia, the pressure response to higher flows was significantly greater and much less of a reduction in hepatic vascular resistance was noted than in studies prior to cold ischemia. Unlike livers prior to cold ischemia, the pressure response to norepinephrine was attenuated following cold ischemia. The response to nitroprusside, however, remained intact reducing the portal pressure to that of in situ livers. Therefore the portal hypertension that follows cold ischemia appears to be largely provoked by the preservation injury and not by surgical manipulation or the bypass circuit. This increment in portal pressure is responsive to a nitric oxide donor.