Christian Eipel

Ischemic Preconditioning Impairs Liver Regeneration in Extended Reduced-Size Livers

Objective:To evaluate the effect of ischemic preconditioning (IPC) in an experimental setting of extended liver resection with 30 minutes of inflow occlusion in rats. Summary Background Data:IPC has been proven an effective strategy against hepatic ischemia-reperfusion injury in both animal and human studies. However, decreased protective effects in terms of transaminase levels were found in patients with larger resection volume, questioning the benefit of IPC in case of small liver remnants. Methods:Rats undergoing 90% hepatectomy under strict inflow occlusion for 30 minutes were subjected to either receive or not receive an IPC period (5 minutes of ischemia followed by 30 minutes of reperfusion). In addition to 10-day survival rate, laser Doppler flowmetry of hepatic blood flow and fluorescence microscopic analysis of the hepatic microcirculation were performed to assess the effect of IPC on initial microvascular reperfusion of liver remnants after 90% resection. Moreover, regeneration capacity of livers undergoing IPC and 70% resection was studied over 7 days by means of histology and immunohistochemistry. Results:Ten-day survival of rats which underwent IPC and 90% hepatectomy was 0 out of 10 animals versus 1 out of 10 animals without IPC. Hemodynamic and microcirculatory analysis revealed signs of hyperperfusion during initial reperfusion of preconditioned liver remnants in 90% hepatectomized animals. In addition to increased transaminase levels, IPC impaired hepatic proliferative response after 70% organ resection, as indicated by both a significant reduction in mitotic figures and Ki-67 nuclear staining of hepatocytes, as well as a decrease in restitution of liver mass. Conclusions:Though portal hypertension reflecting shear stress has been reported to trigger liver regeneration, remnant liver tissue after major hepatectomy may not benefit from hyperperfusion-induced trigger for cell cycle entry but is rather dominated from hyperperfusion-induced local organ injury. Further studies are required to finally judge on the harmfulness of IPC in extended liver resection.

Hyperperfusion Syndrome in Small-for-Size Livers

Background: Portal hyperperfusion in small-for-size livers might seriously impair postoperative liver regeneration. Using an experimental model, we investigated splenectomy as a measure to reduce portal blood flow and its impact on postoperative recovery following extended liver resection. Method: Wistar rats underwent partial (90%) hepatectomy with or without splenectomy under temporary inflow occlusion (30 min). In addition to 10-day survival rate, laser Doppler flowmetry of hepatic blood flow and fluorescence microscopic analysis of hepatic microcirculation were performed to assess the effect of splenectomy on initial microvascular reperfusion of liver remnants. Results: While postischemic perfusion failure was comparable between both groups, portal blood flow was significantly reduced after simultaneous splenectomy (3.5 ± 0.4 vs. 5.4 ± 0.4 ml/min). Moreover, red blood cell velocity and volumetric blood flow were reduced in splenectomized animals. These animals experienced lower AST levels (421 ± 36 vs. 574 ± 73 U/l) and a significantly increased survival rate, reaching 6.6 ± 1.3 vs 2.6 ± 0.8 days. Conclusion: Simultaneous splenectomy significantly reduced the risk for postoperative hyperperfusion syndrome in small-for-size livers. Shear-stress-induced liver injury was diminished due to a significant reduction of portal venous blood flow, which positively influenced postoperative regeneration resulting in significantly higher survival.

Hepatocellular apoptosis is mediated by TNFα-dependent Fas/FasLigand cytotoxicity in a murine model of acute liver failure

There is increasing evidence that the active contribution of hepatocytes to liver disease is strongly dependent on local cytokine environment. It has been shown in vitro that TNFα can enhance hepatocyte FasLigand (FasL)-mediated cytotoxicity. Here, we demonstrate that TNFα-induced apoptosis was associated with Fas and FasL upregulation and that a FasL-neutralizing antibody prevented TNFα-induced apoptosis. We further examined in vivo the relevance of the Fas/FasL pathway to hepatocellular apoptosis in a TNFα-driven model of acute liver failure. Livers of galactosamine/lipopolysaccharide (Gal/LPS)-exposed Fas wild-type mice highly expressed both Fas and FasL and revealed marked hepatocellular apoptosis that was almost completely blocked by soluble TNFα-receptor; this was also almost absent in Gal/LPS-exposed Fas lymphoproliferation mutant mice. Our data provide evidence for a direct link between TNFα and Fas/FasL in mediating hepatocyte apoptosis. Fratricidal death by Fas–FasL interactions of neighbouring hepatocytes may actively contribute to acute liver failure.

Splenectomy improves survival by increasing arterial blood supply in a rat model of reduced‐size liver

Prevention of acute portal hyperperfusion in small‐for‐size livers by inflow modulation results in beneficial postoperative outcome. The objective of this study was to unravel the underlying mechanism, emphasizing the intimate relationship between portal venous (PV) and hepatic arterial (HA) blood flow (BF). Rats underwent partial hepatectomy (pHx), splenectomy before pHx or splenectomy and ligation of the A. hepatica before pHx. Portal venous blood flow (PVBF), hepatic arterial blood flow (HABF), and tissue pO2 were assessed during stepwise resection from 30% to 90%. Hepatic regeneration and hypoxia‐responsive gene expression were analyzed in each group after nonlethal 85% pHx. 90% pHx caused a fourfold rise in PVBF, a slight decrease in HABF with a 50% reduction in pO2, and high mortality. Splenectomy before pHx reduced the PVBF and caused a rise in HABF with doubling in tissue pO2. An attenuation of hypoxia‐responsive gene expression turned into enhanced hepatocellular regeneration and improved survival. A. hepatica ligation abolished the beneficial effect of splenectomy on tissue oxygenation, proliferation, and outcome. In conclusion, the beneficial effect of splenectomy in small‐for‐size livers can be attributed to a rise in HABF with sufficient oxygen supply rather than to a reduced portal venous hyperperfusion to the remnant liver.

A critical appraisal of the hemodynamic signal driving liver regeneration

BackgroundMany aspects of the signaling mechanisms involved in the initiation of hepatic regeneration are under current investigation. Nevertheless, the actual mechanisms switching liver regeneration on and off are still unknown. Hemodynamic changes in the liver following partial hepatectomy have been suggested to be a primary stimulus in triggering liver regeneration. Most of the new knowledge about the impact of hemodynamic changes on liver regeneration is both conceptually important and directly relevant to clinical problems.PurposeThe purpose of this review is therefore to exclusively address the hemodynamic signal driving the liver regeneration process.

Role of the perforin/granzyme cell death pathway in d-Gal/LPS-induced inflammatory liver injury

Cytotoxic T lymphocytes and their granule components, such as perforin and granzyme, play an important role in the defense of hepatic infections caused by different pathogens. Moreover, it has been shown in vitro that hepatocytes can initiate cell death via a perforin-dependent mechanism. Although it is well known that hepatocellular apoptosis in D-galactosamine/lipopolysaccharide (D-Gal/LPS)-associated liver failure is mediated by TNF-alpha-dependent Fas/FasL cytotoxicity, there is no information on the role of perforin-mediated mechanisms in vivo. Therefore, we studied whether the cytolytic perforin/granzyme pathway contributes to the D-Gal/LPS-associated hepatotoxicity. Perforin knockout (Pko) mice showed significantly higher hepatic TNF-alpha and IL-6 mRNA expression as well as plasma TNF-alpha and IL-6 concentrations within the first hour upon D-Gal/LPS challenge compared with perforin wild-type (Pwt) mice. At 6 h upon D-Gal/LPS challenge, Pko mice further presented with higher transaminase release and onconecrotic tissue damage, whereas hepatocellular apoptosis and caspase-3 cleavage remained unaffected by the perforin deficiency. Pretreatment with a recombinant human TNF-alpha receptor fusion protein attenuated necrotic and apoptotic tissue damage and reduced plasma transaminase activities as well as cytokine release, thereby preventing acute liver failure in Pko mice as effectively as in Pwt mice. These data do not only confirm the significance of TNF-alpha as distal mediator of hepatic injury in this model but simultaneously reveal a contribution of a perforin-dependent immunoregulation, limiting the D-Gal/LPS-induced overwhelming cytokine release and onconecrotic tissue injury.

Kupffer cells are mandatory for adequate liver regeneration by mediating hyperperfusion via modulation of vasoactive proteins.

Objective: Physiological liver regeneration requires adequate microvascular perfusion after partial hepatectomy. Although Kupffer cells (KCs) are known to play a key role in modulating hepatocyte proliferation, their impact on regulating hepatic microcirculation during liver regeneration has so far been disregarded. With respect to their expression and modulation of vasoactive mediators, KCs may provide important signals that regulate hepatic perfusion during liver regeneration.

Nitric oxide reduces organ injury and enhances regeneration of reduced-size livers by increasing hepatic arterial flow

Reduced‐size livers suffer from portal hyperperfusion, diminished arterial blood flow and the risk of postoperative liver injury. The aim of this experimental study was to unravel the role of nitric oxide in this setting.

Kupffer cell depletion reduces hepatic inflammation and apoptosis but decreases survival in abdominal sepsis.

Objective During abdominal sepsis, the activation of hepatic Kupffer cells (KC) and its consequences are of central interest. This study evaluates the impact of selective KC depletion on hepatic microcirculation, cytokine release, and systemic alterations in the colon ascendens stent peritonitis (CASP), a model of polymicrobial abdominal sepsis. Methods For KC depletion clodronate liposomes were injected 24 h before CASP surgery in female C57BL/6N mice. Three and 12 h after CASP, in-vivo fluorescence microscopy of the liver was performed. Analysis of hepatocellular apoptosis was conducted by immunohistochemistry. In addition, levels of tumor necrosis factor (TNF), IL-6, and IL-10 in the liver, lungs, spleen, and plasma were determined, and bacteriology and survival analysis were performed. Results CASP led to significant sinusoidal perfusion failure, increased leukocyte recruitment, hepatocellular apoptosis and increased levels of TNF, IL-6, and IL-10 in the liver and plasma. KC depletion before CASP significantly reduced leukocyte recruitment to the liver and hepatocellular apoptosis. IL-10 secretion decreased dramatically in the liver and plasma of KC-depleted septic mice. In contrast, TNF levels were clearly elevated after clodronate treatment. In the lung and spleen, a compensatory upregulation of IL-10 could be detected after KC depletion. Clodronate treatment resulted in a significant reduction in survival. Conclusion The results indicate that KC depletion is locally protective in polymicrobial abdominal sepsis, as it reduces hepatic inflammation and apoptosis. These effects could be observed in the presence of clearly elevated TNF levels. However, the lack of IL-10 in KC-depleted mice resulted in a detrimental systemic proinflammation.

Antileukoproteinase protects against hepatic inflammation, but not apoptosis in the response of D-galactosamine-sensitized mice to lipopolysaccharide

There is major evidence for the strong bi‐directional interrelation of parenchymal cell apoptosis and leukocyte accumulation and inflammation in acute liver injury. Therefore, the aim of this in vivo study was to investigate the anti‐apoptotic and anti‐inflammatory potential of antileukoproteinase (ALP) in a murine model of acute liver failure.