The role of HMGB-1 on the development of necrosis during hepatic ischemia and hepatic ischemia/reperfusion injury in mice

BACKGROUND: High-mobility group 1 (HMGB-1) is a late mediator of endotoxin lethality in mice. The release of HMGB-1 is delayed compared to other proinflammatory cytokines that mediate shock and tissue injury. The purpose of this study was to investigate the role of HMGB-1 levels in response to hepatic ischemia, hepatic I/R injury, and the relationship between changes in HMGB-1 and other cytokines. MATERIALS AND METHODS: Three murine models were employed: our robust model of segmental hepatic warm ischemia (SHWI), a model of partial hepatic ischemia/reperfusion injury (PHIRI), and a model of total hepatic ischemia/reperfusion injury (THIRI). Over a 48-h period following ischemic insult and reperfusion using these models, serum HMGB-1 concentrations, concentrations of HMGB-1 in ischemic and nonischemic lobes, and serum concentrations of TNF-alpha and IL-6 levels were determined in mice. An anti-HMGB-1 antibody treatment was used in SHWI and THIRI to evaluate what aspects of response to ischemia and reperfusion were potentially mediated by HMGB-1. RESULTS: Hepatic HMGB-1 tissue concentrations exhibited biphasic changes in SHWI mice, which were increased in the ischemic lobes relative to nonischemic lobes at 12 h but decreased relative to nonischemic lobes at 24 h after ischemic insult. These results suggested that HMGB-1 was released into the systemic circulation by necrotic cells over the first 12 h but this process may be complete by 24 h postischemia. By 6 to 12 h after SHWI, serum TNF-alpha began to increase significantly and continued to increase for 18 h, followed by a sudden decline. Similarly, serum IL-6 increased over 1-3 h after SHWI and then decreased over the next 6 h. Treatment with an anti-HMGB-1 antibody significantly prolonged survival time in SHWI and THIRI. CONCLUSIONS: HMGB-1 plays a significant role in the response to hepatic ischemia and hepatic ischemia/reperfusion injury. The present study demonstrated a time-dependent production of HMGB-1 following hepatic warm ischemia in mice. The inherent HMGB-1 in ischemic areas was exhausted and HMGB-1 may be released by necrotic cells. HMGB-1 activation is involved in immediate proinflammatory stress response to I/R and anti-HMGB-1 antibody treatment remarkably improved survival. We demonstrated that systemic HMGB-1 accumulation was measured at an earlier phase of the hepatic ischemia and ischemia/reperfusion injury model than LPS-induced endotoxemia.     

Hepatorenal interaction in glutathione redox state during partial hepatic ischemia-reperfusion in rats.

The hepatic glutathione metabolism during partial hepatic ischemia-reperfusion was studied in a rat model with special reference to the hepatorenal glutathione metabolism system. Bile samples, and in- and outflow blood and tissue samples of the rat liver, kidney and small intestine were collected during 60 min of 70% partial liver ischemia and reperfusion. Both reduced and oxidized glutathione levels were determined by HPLC. The tissue ratio of reduced to oxidized glutathione (GSH/GSSG ratio) decreased significantly in the ischemic hepatic lobe at 60 min after reperfusion. The GSH/GSSG ratio in bile from the ischemic hepatic lobes decreased significantly after 60 min of ischemia and gradually recovered after reperfusion. The net release of GSH from the nonischemic hepatic lobe increased at 60 min after reperfusion, since the calculated net release of GSH from the whole liver increased significantly whereas there was no change in the net release from the ischemic hepatic lobe. The tissue GSH level in the kidney increased significantly at 180 min after reperfusion. The calculated net uptake of GSH into the kidney, and the net release of total cysteine from the kidney, tended to decrease at the end of 60 min of ischemia, to increase at 60 min after reperfusion and then decrease at 180 min after reperfusion. We found that the hepatorenal glutathione metabolism was changed by partial hepatic ischemia-reperfusion. These changes might reflect a hepatorenal interaction to maintain the glutathione redox state of the vital organs.     

Tumor Necrosis Factor-α, Interleukin-1β and Nitric Oxide: Induction of Liver Megamitochondria in Prehepatic Portal Hypertensive Rats

Abstact It has been shown that portal hypertension in the rat causes microvesicular hepatocytic fatty infiltration. Formation of megamitochondria (MG) is one of the most prominent alterations in steatosis. Because nitric oxide (NO), tumor necrosis factor-α (TNFα), and interleukin-1β (IL-1β) impair mitochondrial function, these mediators have been studied in prehepatic portal hypertensive rats to verify their coexistence with MG and therefore with steatosis. Male Wistar rats were divided into two groups: a control group (n = 7) and a group with partial portal vein hgation (n = 19) at 6 weeks of evolution. TNFα and IL-1β were quantified in liver by enzyme-linked immunosorbent assay, and NO was measured in the portal vein, suprahepatic inferior vena cava, and infrahepatic inferior vena cava by the Griess reaction. In portal hypertensive rats, the-serum concentration of NO of hepatic origin increases (132.10 ± 34.72 vs. 52.44 ± 11.32 nmol/ml; p < 0.001), as do TNF-α (2.02 ± 0.20 vs. 1.12 ± 0.43 μmol/mg protein) and IL-1β (18.95 ± 2.59 vs. 5.48 ± 1.70 μmol/mg protein) (p = 0.005) in the liver. The most frequent hepatic histologic findings are the presence of MG (p < 0.001), steatosis, and hyperplasia. An increase in hepatic release of NO, TNFα and IL-Iβ with MG formation is produced in rats with portal hypertension. Therefore these proinflammatory mediators and this morphologic mitochondrial alteration could both be involved in the etiopathogenesis of steatosis.     

Induction of different types of cell death after normothermic liver ischemia-reperfusion

Normothermic liver ischemia-reperfusion (I-R) may induce hepatocellular autophagy, apoptosis, and necrosis. The aim of this study was to investigate these three types of cell death in normothermic liver I-R in rats. A segmental normothermic ischemia of the liver was induced for 120 minutes. Liver autophagy was evaluated by transmission electron microscopy and LC3 (Light Chain 3) immunohistochemical studies. Liver apoptosis was assessed by FLIVO (FLuorescence in vIVO) and TUNEL (TdT-mediated dUTP nick end labeling) assays. Liver necrosis was determined by optical microscopic examination. Autophagy was increased in ischemic liver lobes at 6 hours after reperfusion, compared with nonischemic lobes. Fluorescence microscopy showed in situ caspase-3 and -7 specific activity to be increased in ischemic liver lobes after 6 hours of reperfusion, compared with nonischemic lobes. Quantitative analysis of apoptotic cells evaluated by the TUNEL method showed a clearly significant increase in ischemic liver lobes at 6 hours after reperfusion, compared with nonischemic lobes. Necrotic cell death was significantly increased in ischemic liver lobes at 6 hours after reperfusion, compared with nonischemic lobes (P < .005). In conclusion, 120 minutes normothermic liver I-R resulted in increased autophagic, apoptotic and necrotic cell death.     

The Effect of <Emphasis Type="SmallCaps">l</Emphasis>-arginine and Aprotinin on Intestinal Ischemia–reperfusion Injury

Background Intestinal ischemia/reperfusion (I/R) results in local mucosal injury, systemic injuries, and organ dysfunction. These injuries are characterized by altered microvascular and epithelial permeability and villous damage. Activation of neutrophils, platelets, and endothelial factors are known to be involved in this process. Cytokines such as TNF-α, IL-1, IL-6, and oxygen-derived free radicals are believed to be important pathogenic mediators. Capillary no-reflow is also known to play a role in I/R. The aim of our study was to examine the role of l-arginine, a known nitric oxide (NO) donor, and aprotinin, a protease inhibitor with multiple effects, on intestinal I/R. Methods Pigs weighing 20–25 kg were used. Ischemia was established by clamping the superior mesenteric artery (SMA) at its origin and was sustained for 2 hours. Duration of reperfusion was 2 hours. The animals were divided into four groups: group A, the control group, which was submitted to I/R injury only; group B, in which l-arginine was administered at a rate of 5 mg/kg/min during ischemia and continuing throughout reperfusion; group C, in which aprotinin was administered with an initial bolus dose of 20,000 U/kg during ischemia followed by a continuous dose at 50 U/hour throughout reperfusion; and group D in which both substances were administered. In all groups TNF-α, IL-1, and IL-6 levels were measured using ELISA at baseline, 2 hours of ischemia, and 1 hour and 2 hours of reperfusion. SMA blood flow was measured with a Doppler probe at baseline, 10 min, 1 hour, and 2 hours of reperfusion. Histological changes of the intestinal mucosa were examined and graded on a five-point scale in all groups. Results In the control group, levels of TNF-α, IL-1, and IL-6 were significantly increased during reperfusion (p < 0.05) compared to baseline. Administration of l-arginine and aprotinin led to suppression of the release of TNF-α, IL-1, and IL-6 during reperfusion in a statistically significant manner (all p < 0.05). A synergistic or additive effect of l-arginine and aprotinin was not observed. SMA blood flow in the control group was decreased (p > 0.05) during reperfusion compared to baseline. In animals treated with l-arginine and aprotinin, SMA blood flow during reperfusion was significantly increased (p < 0.05) compared to the control group. Histologic examination of the intestinal mucosa was characterized by flattening of the villi and necrosis in the control group. In the treated animals, less severe histological changes were noted. Conclusions Administration of l-arginine and aprotinin may lead to amelioration of intestinal I/R injury. We did not note a synergistic or additive effect of these two substances. These findings warrant further studies in clinical settings for future treatment efforts. This paper was presented as a poster at the 47th Annual Meeting of the Society for Surgery of the Alimentary Tract, Los Angeles, California, May 20–24, 2006.     

Effect of prostaglandin E1 derivative on labilization of liver lysosomal membrane in partial liver ischemia]

The change of liver lysosomal enzymes in tissue and serum during a reperfusion period was studied in partial liver ischemic model in rats and effect of Prostaglandin E1 (PGE1) derivative on partial liver ischemia was investigated. Partial liver ischemia was induced by clamping the branches of the vessels to the right and caudate lobes of rat liver. The clamp was released after 30 minutes of ischemia. Ischemic and nonischemic lobes of the liver were separately removed and the serum was also collected immediately and two hours after the release of the clamp. Lysosomal enzyme activities from free and bound lysosomal fraction were measured separately and the fragility index (F.I.) was calculated. PGE1 derivative was administered intraperitoneally 24, 6, 0.5 hours prior to the induction of ischemia at each dose of 0.05 microgram/kg. Pretreatment with PGE1 derivative prevented lysosomal labilization in ischemic lobe, since there was a significant decrease in F.I. of cathepsin D in the PGE1-pretreated group (preischemia; 28.3 +/- 2.4%, immediately after reperfusion; 30.3 +/- 2.5%, two hours after reperfusion; 30.3 +/- 2.5%) compared to the placebo group (immediately after reperfusion; 40.9 +/- 3.4%, two hours after reperfusion; 41.7 +/- 3.4%, p less than 0.05, p less than 0.05, p less than 0.01, respectively). Pretreatment with PGE1 derivative also significantly suppressed the increase of serum lysosomal enzyme activity. These results showed that PGE1 derivative improved liver lysosomal labilization in partial liver ischemia.     

The protective effects of 17beta-estradiol on hepatic ischemia-reperfusion injury in rat model, associated with regulation of heat-shock protein expression

BACKGROUND: Ischemia-reperfusion (I/R) injury, which was commonly seen in the field of hepatic surgical intervention, impaired liver regeneration and predisposed to liver failure. Previous studies have shown gender dimorphic response of the liver for various hepatic stresses including I/R injury, hemorrhagic shock-resuscitation, liver cirrhosis, endotoxemia, and chronic alcoholic consumption, and demonstrated gender dimorphism in hepatocellular dysfunction after experimental trauma and hemorrhage. The objective of this study was to examine the hypothesis that the protective effects of 17beta-estradiol (E2) in hepatic I/R injury were associated with increasing heat-shock protein 70 expression. MATERIALS AND METHODS: Sprague-Dawley male and female rats were randomly divided into male and female sham, I/R, and E2 + I/R groups. The model of reduced-size liver ischemia and reperfusion was used. Except for the sham-operated groups, all rats were subjected to 70% liver ischemia for 45 min followed by resection of the remaining 30% nonischemic lobes and reperfusion of ischemic tissue. For each group, five rats were used to investigate the survival during a week after operation; blood samples and liver tissues were obtained in the remaining animals after 3, 12, and 24 h of reperfusion to assess serum alanine aminotransferase, aspartate aminotransferase, liver tissue NO(2)(-) + NO(3)(-), malondialdehyde content, superoxide dismutase, nitric oxide synthase, and myeloperoxidase activity, Hsp70 expression, and apoptosis ratio. RESULTS: Compared with I/R groups, male and female E2 + I/R groups showed less I/R-induced injury, and SOD and eNOS activity and Hsp70 expression were increased significantly (P < 0.01). A higher rate of apoptosis was observed in the I/R group versus the E2 + I/R group, a significant increase of MDA, NO(2)(-) + NO(3)(-), and MPO of liver tissues and serum transaminase were also observed in the I/R group versus the E2 + I/R group. The survival rate was significantly higher in the male E2 + I/R group than in the male I/R group. CONCLUSION: E2 pretreatment had protective effects on liver in hepatic I/R injury. The mechanism of this protection might be related to overexpression of Hsp70.     

Alkaline phosphatase reduces hepatic and pulmonary injury in liver ischaemia -- reperfusion combined with partial resection

BACKGROUND: Lipopolysaccharides mediate inflammation in liver ischaemia-reperfusion (I/R) and partial liver resection (PHX). Bovine intestinal alkaline phosphatase (BIAP) detoxifies lipopolysaccharides by dephosphorylation and reduces inflammation in models of sepsis. This study examined the protective effects of BIAP administration in models of partial (70 per cent) liver I/R with or without partial resection of all non-ischaemic lobes during ischaemia (30 per cent). METHODS: Male Wistar rats were divided into six groups: I/R + BIAP, I/R + saline, I/R + PHX + BIAP and I/R + PHX + saline, PHX only or sham laparotomy only. A single dose of BIAP (0.5 units/g) or vehicle (saline) was administered 5 min before reperfusion. Inflammatory response, and hepatic and pulmonary injury were assessed during 24 h of reperfusion. RESULTS: I/R, with or without PHX, increased all markers of inflammation, and hepatic and pulmonary damage (P < 0.050 versus sham operation). I/R + PHX significantly increased release of aspartate aminotransferase (AST) and alanine aminotransferase (ALT), and hepatic neutrophil influx compared with I/R only (P < 0.050). BIAP treatment decreased hepatic wet/dry ratios, neutrophil influx and histopathological damage after I/R with or without PHX (P < 0.050), and also AST, ALT and interleukin (IL)-6 production after I/R + PHX (P < 0.050). BIAP treatment reduced the neutrophil influx after I/R, and pulmonary histopathological injury was decreased after I/R with or without PHX. CONCLUSION: BIAP attenuates hepatic and pulmonary injury after partial liver I/R and PHX.     

Hepatic ischemia-reperfusion promotes liver metastasis of colon cancer

BACKGROUND: The effects of hepatic ischemia/reperfusion (I/R) on liver metastasis have not been fully examined. We examined hepatic I/R and liver metastasis of colorectal cancer in a rat model; we also quantitated expression of E-selectin (ELAM-1) mRNA after I/R. MATERIALS AND METHODS: Rats underwent 30 or 60 min of 70% partial hepatic ischemia. After 60 min of reperfusion, rat colon adenocarcinoma cells (RCN-H4) were inoculated intrasplenically. The number of tumor nodules on the liver surface was determined 3 weeks later. Expression of E-selectin mRNA was determined at 1, 3, and 6 h after ischemia by quantitative RT-PCR. RESULTS: Hepatic I/R promoted liver metastasis of RCN-H4 and induced the expression of E-selectin mRNA in both clamped and unclamped liver lobes. The number of tumor nodules and the expression of E-selectin mRNA after 60 min of ischemia was greater than that after 30 min. CONCLUSIONS: Hepatic I/R, especially with a long duration of ischemia, induces expression of E-selectin and promotes liver metastasis of colon cancer in rats.     

Liver apoptosis following normothermic ischemia-reperfusion: in vivo evaluation of caspase activity by FLIVO assay in rats

Normothermic liver ischemia-reperfusion (I-R) may induce hepatocellular apoptosis. Caspase activation is involved in the initiation and execution of apoptosis. The aim of this study was to determine in vivo caspase activity in normothermic liver I-R in rats. Segmental normothermic ischemia of the liver was induced for 120 minutes in rats. After intravenous injection of the green probe FLIVO, in vivo caspase-3- and -7-specific activity was determined using fluorescence microscopy, in either nonischemic or ischemic liver lobes at 3 and 6 hours after reperfusion. Liver apoptosis was assessed by the deoxynucleotide transferase-mediated deoxyuridine triphosphate nick end labeling (TUNEL) assay. Fluorescence microscopy showed that in vivo caspase-3- and -7-specific activities were significantly increased (P< .005) in ischemic lobes at 3 and 6 hours of reperfusion, compared with nonischemic liver lobes. Quantitative analysis of apoptotic cells measured by the TUNEL method showed a significant increase among apoptotic cells in ischemic lobes at 3 and 6 hours after reperfusion (P< .005), compared with nonischemic liver lobes. In conclusion, 120-minute normothermic liver I-R resulted in increased caspase-3- and -7-specific activities and in liver cell apoptosis.     

Effects of Ozone Oxidative Preconditioning on Liver Regeneration after Partial Hepatectomy in Rats

ABSTRACT

Aim: Similar protective effect of ischemic and ozone oxidative preconditioning (OzoneOP) in hepatic ischemia–reperfusion (I/R) injury was demonstrated, providing evidences that both preconditioning settings shared similar biochemical mechanisms of protection. We investigated the effects of OzoneOP on liver regeneration after 70% partial hepatectomy (PHx) in rats. Methods: Rats were divided into three groups: PHx, I/R + PHx, and OzoneOP + I/R + PHx groups. Ozone (intraperitoneal, 1.2 mg/kg) was given to rats subjected to I/R and 70% hepatectomy daily five times before operation. At 24 hr and 48 hr after resection, samples were collected for the measurement of serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), tumor necrosis factor alpha (TNF-α), and interleukin-6 (IL-6). Moreover, liver regeneration rate, proliferating cell nuclear antigen (PCNA) labeling index, mitotic index, and histopathological examination were evaluated. Results: OzoneOP reduced liver injury determined by liver histology and serum transaminases. There was a rise in serum TNF-α and IL-6 levels in the I/R + PHx group whereas OzoneOP significantly decreased the rise in the level of TNF-α but not IL-6 on the 24 hr and 48 hr of reperfusion. Moreover, liver regeneration in OzoneOP + PHx group, as assessed by the regenerated liver weight, mitotic, and PCNA-labeling index, was significantly improved when compared to I/R + PHx group. Conclusion: These results suggest that OzoneOP ameliorates the hepatic injury associated with I/R and has a stimulatory effect on liver cell regeneration that may make it valuable as a hepatoprotective modality.     

Protective effects of Huang Qi Huai granules on adriamycin nephrosis in rats

Huang Qi Huai (HQH) granules, a mixture of Chinese herbs, contains trametes robiniophila murr, wolfberry fruit, and Polygonatum. We investigated the mechanism of the protective effects of HQH on adriamycin nephrosis (ADR) in rats. Adriamycin nephrotic rats were induced by a single dose of 5 mg/kg adriamycin. For the HQH-treated adriamycin nephrosis group, 1 day after treatment with 5 mg/kg adriamycin, the rats were administered once-daily oral gavage of 2 mg/kg HQH for 15 days. All the rats were killed at day 15. Histological changes were observed by light microscopy and transmission electron microscope. Nephrin and podocin expression levels were measured by real-time RT-PCR and Western blot. Proteinuria was measured by the Bradford protein assay. Serum TNF-α and IL-1β levels were evaluated by ELISA. Macrophage infiltration was detected by immunohistochemistry and immunoblotting, respectively. ADR rats showed heavy proteinuria, podocyte and tubulointerstitial injury, macrophage infiltration, and increased levels of serum cytokines TNF-α and IL-1β. HQH significantly ameliorated the adriamycin-induced renal injury. These data were validated in the cultured podocytes. The podocytes were treated by adriamycin in the presence or absence of HQH and nephrin and podocin expression and TNF-α and IL-1β synthesis and secretion were determined by real-time RT-PCR, immunoblotting, and ELISA, respectively. Adriamycin significantly reduced nephrin and podocin expression, which was significantly restored by the treatment of HQH. HQH treatment inhibited adriamycin-induced TNF-α and IL-1β expression. Our findings suggest that HQH significantly reduces proteinuria, prevents podocyte injury, and ameliorates tubulointerstitial damage. Inhibition of inflammatory cytokine expression and macrophage infiltration may be the protective mechanism of HQH.     

Bone-Resorbing Cytokines from Peripheral Blood Mononuclear Cells after Hormone Replacement Therapy: A Longitudinal Study

Conflicting results have been reported in several cross-sectional studies measuring cytokine production from adherent monocytes in pre- and postmenopausal women. Furthermore, the target cells for the action of estrogen are still debated. We therefore assessed in a longitudinal manner the cytokine production from different fractions of peripheral blood mononuclear cells (PBMC) cultured for 48 h. PBMC were obtained from 30 postmenopausal women before and after 6 months of hormone replacement therapy (HRT). Women were randomly allocated to two groups: an adherent PBMC group (n= 20) and a total PBMC group (n= 9). After 6 months of treatment, urinary pyridinoline levels were markedly decreased in both groups (353 ± 24 vs 114 ± 13 μg/mmol creatinine and 325 ± 35 vs 164 ± 31 μg/mmol creatinine respectively, p<0.01). Culture supernatants were assayed for interleukin 1β (IL-1β), interleukin 6 (IL-6), soluble IL-6 receptor (IL-6rs) and tumor necrosis factor alpha (TNF-α). In the adherent PBMC group, HRT induced a nonsignificant trend toward decreased levels of IL-1β (35 ± 10 vs 13 ± 5 pg/ml), TNF-α (333 ± 58 vs 222 ± 30 pg/ml) and IL-6 (115 ± 70 vs 17 ± 10 pg/ml). In contrast, in the total PBMC group, HRT induced a consistent and dramatic decrease in levels of IL-1β (104 ± 22 vs 25 ± 8 pg/ml), IL-6 (5950 ± 1041 vs 1011 ± 361 pg/ml), IL-6rs (148 ± 33 vs 35 ± 12 pg/ml) (p<0.01) and TNF-α (1468 ± 315 vs 585 ± 207 pg/ml, p= 0.05). We then evaluated whether HRT had the same effect in vitro. Adherent or total PBMC of 8 postmenopausal women were cultured with or without 10⁻⁸M 17β-estradiol or tibolone for 48 h. Production of IL-1β, TNF-α, IL-6 and IL-6rs was not affected by the presence of 17β-estradiol or tibolone in cultures of these cell fractions. In conclusion, our data indicate that non-adherent PBMC could mediate the response to HRT. HRT may exert its action indirectly via noncirculating cells, as suggested by the absence of an in vitro effect. Received: 11 July 2000 / Accepted: 15 January 2001     

Role of Purines on Hepatic Ischemia-Reperfusion Lesions in Rabbit

In this work, we evaluate the effects of adenosine 5′ triphosphate (ATP) on hepatic lesions caused by ischemia/reperfusion (I/R) in liver rabbit. Rabbits were pretreated with ATP (15 mg/kg IV) or saline solution 0.9% (SS), before the hepatic I/R procedure. We evaluated the effects of ATP on hepatic injury before and after I/R. The warm hepatic I/R procedure caused profound acute liver injury, as indicated by elevated serum aspartate aminotransferase, alanine aminotransferase, and lactic dehydrogenase levels, as well as a high apoptotic cell count. All these changes were attenuate by ATP treatment before the hepatic I/R procedure. These results suggested that ATP exerted protective effects on hepatic I/R lesions in the rabbit. This ATP effect may be related to improved energy metabolism during reperfusion in ischemic livers protecting against functional damage of cellular and subcellular membranes during lipid peroxidation.     

Interleukin-6 and Tumor Necrosis Factor-α Levels following Hepatic Cryotherapy: Association with Volume and Duration of Freezing

Although morbidity following cryotherapy is usually minor, a syndrome of multiorgan failure and disseminated intravascular coagulation (DIC) has been described and referred to as the cryoshock phenomenon. We hypothesized that mediators similar to those in septic shock may be involved in this syndrome. In this study we aimed to assess the plasma concentrations of the cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) following hepatic cryotherapy and to relate them to the duration and volume of freezing and to hepatocellular injury. Between April and December 1997 blood samples were taken preoperatively and at different times postoperatively from patients undergoing hepatic artery catheter-insertion (HAC) (n= 15), cryotherapy (n= 5), liver resection (n= 9), liver resection and edge cryotherapy (n= 7), or liver resection and cryotherapy of additional lesions (n= 9). They were analyzed for serum aspartate transaminase (AST) and plasma TNF-α and IL-6 levels. There was a significant association (Pearson correlation) of serum AST levels 1 hour postoperatively with plasma TNF-α and IL-6 levels at the end of the procedure. In patients undergoing cryotherapy or resection with cryotherapy of additional lesions (n= 14), the volume and duration of hepatic freezing were significantly associated with postoperative serum AST and plasma TNF-α and IL-6 levels at various postoperative times. Hepatic cryotherapy is followed by cytokine release, with postoperative plasma TNF-α and IL-6 levels associated with the degree of hepatic cryotrauma. These mediators may be involved in the occurrence of cryoshock following large-volume hepatic freezing.     

Local secretion of TNF-α from the liver does not correlate with endotoxin, IL-6, or organ function in the early phase after orthotopic liver transplantation

Hepatic ischemia/reperfusion leads to an excessive release of proinflammatory cytokines, which promotes local and remote cell damage. The value of cytokine measurement in humans for predicting graft function after orthotopic liver transplantation (OLT) remains unclear. Therefore, in this study, tumor-necrosis-factor-α (TNF-α), interleukin-6 (IL-6), and endotoxin (ET) levels were determined in the blood taken from the hepatic veins of 31 patients who underwent OLT. Peak levels of TNF-α in hepatic venous blood were measured shortly after reperfusion and were significantly higher than concentrations in the systemic circulation. IL-6 concentrations, peaking 90 min after reperfusion, only correlated with postoperative pulmonary dysfunction. ET was detectable in 21 patients, but levels did not correlate with either IL-6 or TNF-α concentrations. Additionally, serum cytokine levels did not correlate with the duration of ischemia or with histological changes seen in liver biopsies. In general, our study suggests that local secretion of cytokines does not predict liver function in the early posttransplant phase. Received: 25 October 1999 Revised: 7 July 2000 Accepted: 22 November 2000     

Effect of Portal Vein Occlusion on the Pancreas: An Experimental Model

Background The effects of portal vein occlusion on the pancreas are not clearly understood. Therefore, we studied histomorphological changes induced in the rat pancreas by various periods of portal vein occlusion. Materials and methods Sixty female Wistar albino rats were randomly allocated into four groups of 15 each. In Group I (control), rats underwent sham laparotomy to expose the portal vein proximal to its bifurcation. In Groups II–IV, rats underwent laparotomy followed by portal vein occlusion by clamping for 15, 30, and 60 minutes respectively. The pancreas was removed immediately after sham laparotomy in Group I and immediately after clamp release in Groups II–IV. Pancreatic tissue specimens were subjected to histochemical analysis for cell typing and diagnosis, immunohistochemical analysis for identification of the inflammatory markers tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), endothelial nitric oxide synthase (eNOS), and inducible NOS (iNOS), and TUNEL analysis was carried out for identification of apoptotic cells. Results Histochemistry revealed signs of inflammation in pancreatic tissue from rats subjected to portal vein occlusion. Immunohistochemistry revealed that the expression of proinflammatory cytokines TNF-α and IL-1β and the oxidative damage indicator iNOS in rat pancreatic tissue increased progressively with the duration of portal vein occlusion. TUNEL assay revealed no signs of apoptosis in any of the groups. Conclusion We conclude that portal vein occlusion triggers an inflammatory response in the pancreas that worsens the longer the occlusion lasts.     

Role of P-selectin expression in hepatic ischemia and reperfusion injury

Background. Researchers have shown that reperfusion of ischemic tissues initiates a complex series of reactions that paradoxically injure tissues. Although several mechanisms have been proposed to explain the pathobiology of ischemic/reperfusion (I/R) injury, much attention has focused on adhesion molecules. Our research is intended to show the kinetics of P-selectin in the liver in response to I/R injury.Methods. Left-lobar hepatic ischemia was induced for 30 min in 35 C57BL-6 mice and 20 P-selectin-deficient (K-O) mice. P-selectin expression was measured in these mice at 20 min, 2, 5, 12 and 24 h reperfusion times, as well as in control and sham animals. The animals were injected with radio-labeled P-selectin monoclonal antibody and the organs were harvested for counts/g tissue, expressed as the percentage injected dose. Serum liver enzymes were measured and pathological sections of ischemic and control livers were performed. The unpaired t -test was used for statistical analysis.Results. P-selectin expression showed two peaks in this animal model. The first peak was at 20 min and the second peak at 5 h of reperfusion (p<0.001). We documented an 8-fold increase in aspartate aminotransferase (AST), alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) levels 10 h following I/R injury. Pathological specimens showed periportal necrosis consistent with an ischemic event. P-selectin K-O mice showed no up-regulation as a separate control group, and the liver enzymes were significantly lower than the wild-type mice at 10 h (p<0.001).Conclusion. P-selectin has a bimodal expression following hepatic I/R injury. The first peak is attributed to the Weibel–Palade bodies and the second to new translational P-selectin. We noted no difference in the up-regulation of P-selectin in the ischemic and non-ischemic liver lobes in the same animal.