Hepatic Iron Overload following Liver Transplantation from a C282Y/H63D Compound Heterozygous Donor.

Hereditary hemochromatosis (HH) is a genetic disease associated with progressive iron overload, eventually leading in some cases to damage of parenchymal organs, such as the liver, pancreas, and heart. Although the gene had been identified (HFE), HH pathogenesis remains to be fully elucidated. We report here, for the first time, a case of inadvertent transplantation of a liver from a donor with C282Y/H63D compound heterozygosity into a nonhemochromatotic 19-year-old Caucasian male recipient with primary sclerosing cholangitis. Progressive iron overload occurred over 1.5 years, as observed in liver biopsies and iron studies, after ruling out secondary causes of iron overload. This case strengthens the hypothesis that the liver, rather than the small intestine, plays a primary role in the maintenance of iron homeostasis.

Induction of heme oxygenase-1 protects mouse liver from apoptotic ischemia/reperfusion injury.

Ischemia/reperfusion (I/R) injury is the main cause of primary graft dysfunction of liver allografts. Cobalt-protoporphyrin (CoPP)-dependent induction of heme oxygenase (HO)-1 has been shown to protect the liver from I/R injury. This study analyzes the apoptotic mechanisms of HO-1-mediated cytoprotection in mouse liver exposed to I/R injury. HO-1 induction was achieved by the administration of CoPP (1.5 mg/kg body weight i.p.). Mice were studied in in vivo model of hepatic segmental (70 %) ischemia for 60 min and reperfusion injury. Mice were randomly allocated to four main experimental groups (n = 10 each): (1) A control group undergoing sham operation. (2) Similar to group 1 but with the administration of CoPP 72 h before the operation. (3) Mice undergoing in vivo hepatic I/R. (4) Similar to group 3 but with the administration of CoPP 72 h before ischemia induction. When compared with the I/R mice group, in the I/R+CoPP mice group, the increased hepatic expression of HO-1 was associated with a significant reduction in liver enzyme levels, fewer apoptotic hepatocytes cells were identified by morphological criteria and by immunohistochemistry for caspase-3, there was a decreased mean number of proliferating cells (positively stained for Ki67), and a reduced hepatic expression of: C/EBP homologous protein (an index of endoplasmic reticulum stress), the NF-κB's regulated genes (CIAP2, MCP-1 and IL-6), and increased hepatic expression of IκBa (the inhibitory protein of NF-κB). HO-1 over-expression plays a pivotal role in reducing the hepatic apoptotic IR injury. HO-1 may serve as a potential target for therapeutic intervention in hepatic I/R injury during liver transplantation.

Magnetic resonance cholangiopancreatography for the accurate diagnosis of biliary complications after liver transplantation: comparison with endoscopic retrograde cholangiography and percutaneous transhepatic cholangiography - long-term follow-up.

Biliary complications after liver transplantation remain a serious cause of morbidity and mortality. Direct invasive cholangiographic techniques, endoscopic retrograde cholangiography (ERCP) or percutaneous transhepatic cholangiography (PTC), have procedure-related complications. Magnetic resonance cholangiopancreatography (MRCP) is non-invasive, safe, and accurate. The aim of this study was to evaluate MRCP in detecting biliary complications following liver transplantation and comparing findings with ERCP and PTC. Twenty-seven consecutive liver transplant recipients who presented with clinical and biochemical, ultrasonographic, or histological evidence of biliary complications were evaluated with MRCP. Patients were followed up for a median period of 36 months. The presence of a biliary complication was confirmed in 18 patients (66.6%): anastomotic biliary stricture in 12 (66.6%); diffuse intrahepatic biliary stricture in 5 (27.7%): ischemic (n = 3), recurrence of primary sclerosing cholangitis (n = 2), and choledocholithiasis in one. In nine patients (33.3%), MRCP was normal. Six patients underwent ERCP, and eight PTC. There was a statistically significant correlation between the MRCP and both ERCP and PTC (p = 0.01) findings. The sensitivity and specificity of the MRCP were 94.4% and 88.9%, respectively, and the positive and negative predictive values, 94.4% and 89.9%, respectively. MRCP is an accurate imaging tool for the assessment of biliary complications after liver transplantation. We recommend that MRCP be the diagnostic imaging modality of choice in this setting, reserving direct cholangiography for therapeutic procedures.

A continuous 13C methacetin breath test for noninvasive assessment of intrahepatic inflammation and fibrosis in patients with chronic HCV infection and normal ALT.

SUMMARY: Up to 30% of patients with hepatitis C virus (HCV) infection and normal serum alanine aminotransferase (NALT) have significant liver disease. Currently, many of these patients undergo a liver biopsy to guide therapeutic decisions. The BreathID continuous online (13)C-methacetin breath test (MBT) reflects hepatic microsomal function and correlates with hepatic fibrosis. To assess its role in identifying intrahepatic inflammation and fibrosis in NALT patients, we tested 100 patients with untreated chronic HCV infection, and 100 age- and sex-matched healthy volunteers using (13)C MBT following ingestion of 75 mg methacetin. All HCV patients had undergone a liver biopsy within 12 months of performing the MBT. Patients with a necroinflammatory grade 4, based on Ishak (modified HAI) score, HAIa + HAIb + HAIc + HAId, were defined as having low or high inflammation, respectively. Patients with a histological activity fibrosis stage 2, were defined as having nonsignificant or significant fibrosis, respectively. A proprietary algorithm to differentiate intrahepatic inflammation within chronic HCV patients with NALT achieved an area under the curve (AUC) of 0.90. Setting a threshold on the point of best agreement (at 83%) results in 82% sensitivity and 84% specificity. With application of another proprietary algorithm to differentiate patients with nonsignificant or significant fibrosis, 67% of liver biopsies performed in the patient group could have been avoided. This algorithm achieved an AUC of 0.92, with a sensitivity of 91% and a specificity of 88%. There was no correlation between body mass index (BMI) and MBT scores for patients with the same histological score. The continuous BreathID(13)C MBT is an accurate tool for measuring the degree of inflammation and fibrosis in patients with chronic HCV infection and NALT. As such, it may prove to be a powerful, noninvasive alternative to liver biopsy in the management of this patient population.

Immunomodulatory effects of plasminogen activators on hepatic fibrogenesis.

Tissue-type plasminogen activators (tPA) and urokinase-type plasminogen activators (uPA) are involved in liver repair. We examined the potential immunomodulatory actions of uPA, tPA and uPA-receptor (uPAR) in carbon-tetrachloride-induced hepatic fibrosis in wild-type (WT), tPA-/-, uPA-/- and uPAR-/- mice. Carbon-tetrachloride treatment increased fibrosis in four groups but significantly less in three knock-out models. Serum cytokines and intrahepatic T cells elevated significantly following fibrosis process in WT animals but not in the knock-out groups. In culture, uPA increased lymphocyte proliferation significantly in WT and uPA-/- but not uPAR-/- animals. Following uPA exposure in vivo, there was CD8 predominance. To isolate uPA's effect on lymphocytes, WT mice were irradiated sublethally and then reconstituted with WT or uPA-/- lymphocytes. In these animals fibrosis was decreased and T cells were reduced in the uPA-/- recipients. Based on these data we postulate that plasminogen activators affect fibrosis in part by liver-specific activation of CD8 subsets that govern the fibrogenic activity of hepatic stellate cells.

The clinical spectrum of cytomegalovirus colitis in adults.

BACKGROUND: Colonic cytomegalovirus reactivation rarely occurs in adults without inflammatory bowel disease or a known immunosuppressive state. AIM: To describe our experience with such patients. METHODS: All consecutive admissions of patients with possible cytomegalovirus colitis, between 1995 and 2006, were reviewed retrospectively. RESULTS: Nineteen patients were studied. Most of the patients were elderly with multiple co-morbidities. Three main forms of disease presentation were recognized: acute diarrhoea, chronic diarrhoea and lower gastrointestinal bleeding. Colonic mucosal intranuclear inclusion bodies were found in 12 patients. Thirteen patients had cytomegalovirus viraemia (either by polymerase chain reaction and/or by white blood cell-cytomegalovirus antigenaemia test). Ganciclovir therapy was given to only eight patients; only five of these patients survived. The other subgroup of 11 patients received only supportive therapy. Most of the patients from this subgroup had a prolonged and complicated hospital course; only nine patients survived. Follow-up colonoscopies were performed only in five patients (out of the 14 patients who survived). In four of these patients, chronic mucosal inflammatory changes were noted. CONCLUSIONS: Cytomegalovirus colitis occurs rarely in adult individuals. The disease may have various and multiple acute and/or chronic clinical manifestations. Clinical awareness of this condition is needed.

Beta-glucosylceramide: a novel method for enhancement of natural killer T lymphoycte plasticity in murine models of immune-mediated disorders.

BACKGROUND: beta-Glucosylceramide, a naturally occurring glycolipid, exerts modulatory effects on natural killer T (NKT) lymphocytes. AIM: To determine whether beta-glucosylceramide can alter NKT function in opposite directions, colitis was induced by intracolonic installation of trinitrobenzenesulphonic acid, and hepatocellular carcinoma (HCC) was induced by transplantation of Hep3B cells. METHODS: The immunological effect of beta-glucosylceramide was assessed by analysis of intrahepatic and intrasplenic lymphocyte populations, serum cytokines and STAT protein expression. RESULTS: Administration of beta-glucosylceramide led to alleviation of colitis and to suppression of HCC, manifested by improved survival and decreased tumour volume. The beneficial effects were associated with an opposite immunological effect in the two models: the peripheral:intrahepatic CD4:CD8 lymphocyte ratio increased in the colitis model and decreased in the HCC group. The peripheral:intrahepatic NKT lymphocyte ratio decreased in beta-glucosylceramide-treated mice solely in the HCC model. The effect of beta-glucosylceramide was associated with decreased STAT1 and 4 expression, and with overexpression of STAT6, along with decreased interferon gamma levels in the colitis model, whereas an opposite effect was noted in the HCC model. CONCLUSIONS: beta-glucosylceramide alleviates immunologically incongruous disorders and may be associated with "fine tuning" of immune responses, by changes in plasticity of NKT lymphocytes.

CTLA-4.FasL inhibits allogeneic responses in vivo.

CTLA-4.Fas ligand (CTLA-4.FasL), a paradigmatic 'trans signal converter protein (TSCP)', can attach to APC (via CTLA-4 binding to B7) and direct intercellular inhibitory signals to responding T cells (via FasL binding to Fas receptor), converting an activating APC-to-T cell signal into an inhibitory one. Our previous studies established that CTLA-4.FasL inhibits human primary mixed lymphocyte reactions (MLR) and induces alloantigen-specific hyporesponsiveness ex vivo. The present study extends this to an in vivo context. Using splenocytes from MHC-mismatched C57BL/6 and Balb/c mice, we demonstrated that his(6)CTLA-4.FasL, effectively inhibits murine MLR. Moving in vivo, we demonstrated that subcutaneously administered his(6)CTLA-4.FasL modulates the in vivo response of infused allogeneic splenocytes. his(6)CTLA-4.FasL reduces the number of cells in each cell division, and increases the percentage of dead cells in each division. These findings are consistent with an antigen-induced cell death of the alloreactive cells, and bolsters recombinant TCSP promise as a therapeutic for transplantation diseases.

Adoptive transfer of regulatory NKT lymphocytes ameliorates non-alcoholic steatohepatitis and glucose intolerance in ob/ob mice and is associated with intrahepatic CD8 trapping.

The aim of this study was to determine the effect of adoptive transfer of regulatory natural killer T (NKT) lymphocytes on the metabolic disorder in leptin-deficient ob/ob mice, which feature depletion and defective function of NKT and CD4 lymphocytes. Leptin-deficient ob/ob mice were subjected to transplantation of 1 x 10(6) of either ob/ob or wild-type-derived NKT lymphocytes, or to transplantation of either ob/ob or wild-type-derived splenocytes. The effect on hepatic fat content was measured by magnetic resonance imaging (signal intensity index) and histology, using the steatohepatitis grading scale. The degree of glucose intolerance was measured by an oral glucose tolerance test (GTT). Adoptive transfer of wild-type or ob/ob-derived regulatory NKT cells led to a 12% decrease in hepatic fat content. A significant histological shift from macrosteatosis to microsteatosis was observed. Marked improvement in the GTT was noted in wild-type or ob/ob-derived NKT recipients. Metabolic effects were associated with a significant decrease in peripheral and intrahepatic CD4/CD8 lymphocyte ratios. Intrahepatic CD8 trapping was observed in all responders. Serum interleukin 10 levels decreased significantly. In conclusion, adoptive transfer of a relatively small number of regulatory NKT lymphocytes into ob/ob mice results in a significant reduction in hepatic fat content, a shift from macro to microsteatosis, and significant improvement in glucose intolerance. These effects were associated with decreased peripheral and intrahepatic CD4/CD8 ratios and decreased interleukin 10 levels. The results further support a role for regulatory NKT lymphocytes in the pathogenesis of non-alcoholic steatohepatitis in the leptin-deficient murine model.

Cathepsin B inactivation attenuates the apoptotic injury induced by ischemia/reperfusion of mouse liver.

BACKGROUND: A major mechanism underlying warm ischemia/reperfusion (I/R) injury during liver transplantation is the activation of the caspase chain, which leads to apoptosis. Recently, it was demonstrated that the release of cathepsin B, a cysteine protease, from the cytosol in liver injury induces mitochondrial release of cytochrome c and the activation of caspase-3 and -9, thereby leading to apoptosis. The aim of this study was to ascertain if cathepsin B inactivation attenuates the apoptotic injury due to I/R in mouse liver. METHODS: A model of segmental (70%) hepatic ischemia was used. Eighteen mice were anesthetized and randomly divided into three groups: (1) CONTROL GROUP: sham operation (laparotomy); (2) Ischemic group: midline laparotomy followed by occlusion of all structures in the portal triad to the left and median lobes for 60 min (ischemic period); (3) STUDY GROUP: like group 2, but with intraperitoneal administration of a pharmacological inhibitor of cathepsin B (4 mg/100 g) 30 min before induction of ischemia. Serum liver enzyme levels were measured by biochemical analysis, and intrahepatic caspase-3 activity was measured by fluorometric assay; apoptotic cells were identified by morphological criteria, the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) fluorometric assay, and immunohistochemistry for caspase-3. RESULTS: Showed that at 6 h of reperfusion, there was a statistically significant reduction in liver enzyme levels in the animals pretreated with cathepsin B inhibitor (p<0.05). On fluorometric assay, caspase-3 activity was significantly decreased in group 3 compared to group 2 (p<0.0001). The reduction in postischemic apoptotic hepatic injury in the cathepsin B inhibitor -treated group was confirmed morphologically, by the significantly fewer apoptotic hepatocyte cells detected (p<0.05); immunohistochemically, by the significantly weaker activation of caspase-3 compared to the ischemic group (p<0.05); and by the TUNEL assay (p<0.05). CONCLUSION: The administration of cathepsin B inhibitor before induction of ischemia can attenuate postischemic hepatocyte apoptosis and thereby minimize liver damage. Apoptotic hepatic injury seems to be mediated through caspase-3 activity. These findings have important implications for the potential use of cathepsin B inhibitors in I/R injury during liver transplantation.

Effect of adenosine A2A receptor agonist (CGS) on ischemia/reperfusion injury in isolated rat liver.

Ischemia/reperfusion injury during liver transplantation is a major cause of primary nonfunctioning graft for which there is no effective treatment other than retransplantation. Adenosine prevents ischemia-reperfusion-induced hepatic injury via its A2A receptors. The aim of this study was to investigate the role of A2A receptor agonist on apoptotic ischemia/reperfusion-induced hepatic injury in rats. Isolated rat livers within University of Wisconsin solution were randomly divided into four groups: (1) continuous perfusion of Krebs-Henseleit solution through the portal vein for 165 minutes (control); (2) 30-minute perfusion followed by 120 minutes of ischemia and 15 minutes of reperfusion; (3) like group 2, but with the administration of CGS 21680, an A2A receptor agonist, 30 microg/100 ml, for 1 minute before ischemia; (4) like group 3, but with administration of SCH 58261, an A2A receptor antagonist. Serum liver enzyme levels were measured by biochemical analysis, and intrahepatic caspase-3 activity was measured by fluorometric assay; apoptotic cells were identified by morphological criteria, the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) fluorometric assay, and immunohistochemistry for caspase-3. Results showed that at 1 minute of reperfusion, there was a statistically significant reduction in liver enzyme levels in the animals pretreated with CGS (p < 0.05). On fluorometric assay, caspase-3 activity was significantly decreased in group 3 compared to group 2 (p < 0.0002). The reduction in postischemic apoptotic hepatic injury in the CGS-treated group was confirmed morphologically, by the significantly fewer apoptotic hepatocyte cells detected (p < 0.05); immunohistochemically, by the significantly weaker activation of caspase-3 compared to the ischemic group (p < 0.05); and by the TUNEL assay (p < 0.05). In conclusion, the administration of A2A receptor agonist before induction of ischemia can attenuate postischemic apoptotic hepatic injury and thereby minimize liver injury. Apoptotic hepatic injury seems to be mediated through caspase-3 activity.

TPEN attenuates hepatic apoptotic ischemia/ reperfusion injury and remote early cardiac dysfunction.

The release of cardioactive substances during hepatic ischemia/reperfusion injury generates toxic free radicals that inflict hepatic and remote cardiac damage. The aim of the study was to determine whether TPEN, a potent iron chelator, ameliorates the apoptotic hepatic and cardiac function injuries. Three groups of isolated rat livers were studied: (1) continuously perfused with Krebs-Henseleit solution; (2) subjected to 120 min of ischemia and 15 min of reperfusion; (3) as in group 2, with TPEN administered prior to ischemia. Isolated hearts were perfused for 65 min with the effluent of the reperfused livers. Results showed that TPEN administration reduced the release of norepinephrine, epinephrine, dopamine, prostaglandin E2 and angiotensin II, decreased intrahepatic caspase-3 activity, and decreased the mean hepatocyte apoptotic index (TUNEL assay) (p = 0.001). Perfusion with post-ischemic hepatic effluent caused a transient 15-min increase in left ventricular contraction and coronary flow (p < 0.05), followed by a decrease in cardiac function at one hour. TPEN reduced the transient elevation in left ventricular contraction p < 0.05), but did not prevent the subsequent decrease in cardiac function. In conclusion, TPEN attenuates post-ischemic apoptotic hepatic injury by modulating caspase-3-like activity and reduces the cardioactive substances released from the liver.

The CCR5Delta32 allele is associated with reduced liver inflammation in hepatitis C virus infection.

CCR5Delta32 is a deletion mutation in the chemokine receptor CCR5. Liver inflammatory activity was found to be significantly reduced (P = 0.005) in Jewish Israeli patients infected with the hepatitis C virus (HCV) carrying the CCR5Delta32 allele. The CCR5Delta32 allele does not alter susceptibility to HCV infection; however, it may play a role in the progression and outcome of the disease.

Mast cells and eosinophils have a potential profibrogenic role in Crohn disease.

BACKGROUND: Mast cells and eosinophils have an important role in allergic inflammation and probably also in chronic inflammatory diseases resulting in fibrosis, such as Crohn disease where fibrosis is present as strictures. The involvement of mast cells and eosinophils in Crohn disease fibrosis was investigated. METHODS: Biopsies from diseased foci were stained for mast cells, eosinophils, anti-collagen type IV and VIII, laminin and alpha-smooth muscle actin (alpha-SMA) (IHC). Fibroblasts outgrown from the biopsies and a normal fetal intestinal fibroblast line were cultured in the presence of the human mast cell line HMC-1, or of human peripheral blood eosinophil (MACS, purity > 98%) sonicates, or of selected mediators. Fibroblast proliferation (3H-thymidine), collagen synthesis ([3H]-proline) and collagen gel contraction were evaluated. RESULTS: Mast cells were present in all the biopsies and only faintly positive for extra cellular matrix (ECM) products. Pronounced eosinophilia was detected in only two cases. Mast cell sonicates increased both Crohn disease (alpha-SMA positive) and control fibroblast proliferation, decreased collagen production and increased collagen gel contraction. Eosinophil sonicates increased fibroblast proliferation, gel contraction and collagen production. TNF-alpha decreased collagen production. Histamine, tryptase and chymase had no influence. CONCLUSIONS: These in vitro data show that mast cells and eosinophils could be involved in modulating Crohn disease fibrosis by directly influencing intestinal fibroblast properties.

Role of circulating soluble CD40 as an apoptotic marker in liver disease.

OBJECTIVES: To measure levels of soluble CD40, a laboratory marker of apoptosis in patients with liver disease, determine its origin, and correlate the findings with disease activity and histology. DESIGN: Laboratory research study with comparison group. SETTING: Liver Institute, Laboratory of HLA Typing and Histopathology Department, Rabin Medical Center, Israel. SUBJECTS: One hundred ten patients with liver disease and 20 healthy controls. METHODS: Serum samples were collected from all patients; in addition, paired hepatic and portal vein samples were collected from 23 patients, and bile samples from 5 patients. Soluble CD40 was measured with an enzyme-linked immunosorbent assay. Apoptotic cells in liver tissue were identified by morphological criteria and quantified with the TUNEL assay. RESULTS: Soluble CD40 concentration was significantly higher in patients with liver disease than controls (mean 112.9 +/- 197.2 pg/ml vs. 24.2 +/- 9.1 pg/ml, p = 0.0001), with highest levels in the chronic viral hepatitis group (mean 131.7 +/- 137.5 pg/ml, p = 0.0001). Levels of sCD40 were correlated with serum creatinine, alkaline phosphatase, alpha-feto protein, and the apoptotic index. In the 23 paired samples, CD40 level was higher in the hepatic vein (mean 74.9 +/- 114.5 pg/ml) than the portal vein (mean 51.6 +/- 67.9 pg/ml); it was highly detectable in bile (mean 115.6 +/- 119.6 pg/ml, p = 0.0123). Untreated patients with chronic viral hepatitis (B and C) had higher levels (mean 106.2 +/- 76.5 pg/ml) than treated patients (mean 59.3 +/- 68.6 pg/ml, p = 0.049). CONCLUSIONS: Levels of soluble CD40 increase in different types of liver disease. It probably derives from the liver and is secreted into the bile. Levels correlate with the apoptotic index and are affected by antiviral treatment. Soluble CD40 may serve as a serum marker of apoptosis in liver disease.

Circulating soluble cytochrome c in liver disease as a marker of apoptosis.

OBJECTIVES: To measure levels of soluble cytochrome c, a clinical marker of apoptosis in patients with liver disease; determine whether soluble cytochrome c is derived from the liver; and correlate soluble cytochrome c level with histology and disease activity. DESIGN: Laboratory research study with comparison group. SETTING: Liver Institute, at the Rabin Medical Center, Israel, and In Vitro Toxicology Laboratory, Canada. SUBJECTS: A total of 108 patients with liver disease and 30 healthy controls. INTERVENTIONS: Paired hepatic and portal vein samples were taken via the transjugular vein in patients after liver biopsy and transjugular intrahepatic portacaval shunt, and bile from patients with external biliary drainage. Soluble cytochrome c was measured with an enzyme-linked immunosorbent assay in peripheral blood. Apoptotic cells in liver tissue were identified by morphological criteria and quantitated with the dUTP nick-end-labelling (TUNEL) assay. MAIN OUTCOME MEASURES: Soluble cytochrome c level by type of liver disease by clinical and histological findings. RESULTS: Soluble cytochrome c concentration (mean 187.1 +/- 219.5 ng x mL(-1)) was significantly higher in patients with liver disease than in controls (39.8 +/- 35.1 ng x mL(-1); P = 0.0001), with highest levels in the primary sclerosing cholangitis group (mean 1041.0 +/- 2844.8 ng x mL(-1); P = 0.001). Cytochrome c levels were correlated with serum bilirubin, alkaline phosphatase, creatinine levels, necroinflammatory score and apoptotic index, but not with serum alanine aminotransferase and synthetic liver function tests. In the 16 paired samples, soluble cytochrome c level was higher in the hepatic (mean 267.9 +/- 297.0 ng x mL(-1)) than the portal vein (mean 169.2 +/- 143.3 ng x mL(-1)), and it was highly detectable in bile (mean 2288.0 +/-4596.0 ng x mL(-1)) (P = 0.001). Untreated patients with chronic viral hepatitis (B and C) had significantly higher levels (mean 282.8 +/-304.3 ng x mL(-1)) than treated patients (77.9 +/- 35.8 ng x mL(-1); P = 0.001). CONCLUSIONS: Soluble cytochrome c levels are increased in different types of liver disease. Soluble cytochrome c is probably derived from the liver and secreted into the bile. Levels correlate with the apoptotic index and are affected by antiviral treatment. Soluble cytochrome c may serve as a serum marker of apoptosis.

Mast cells involvement in the inflammation and fibrosis development of the TNBS-induced rat model of colitis.

BACKGROUND: Mast cells have been implicated in chronic inflammatory conditions resulting in fibrosis, such as Crohn disease. However, a link between inflammation, fibrosis and mast cells has not been demonstrated in human or animal intestinal diseases. This work was undertaken to analyze whether mast cells play a role in inflammation and fibrosis in the TNBS-induced rat colitis. METHODS: Rats were rectally instilled 2,4,6,-trinitrobenzene sulfonic acid in ethanol, and immediately or 4 days later injected daily i.p. with nedocromil sodium, a mast cell stabilizer, compound 48/80, a mast cell activator, or saline. Rats were sacrificed 5 days post-TNBS, or on day 21. Intestinal inflammation and fibrosis were assessed by gross and histopathological evaluation. Colonic mast cell numbers (toluidine blue) and collagen (type I mRNA expression) were evaluated. Mast cell sonicate was added to rat colon fibroblasts. Fibroblast proliferation (3H-thymidine), collagen synthesis (3H-proline) and contractile activity (tridimensional collagen lattice contraction) were then assessed. RESULTS: Nedocromil reduced inflammation and fibrosis possibly by decreasing mast cell numbers and activation and consequent collagen production. Compound 48/80 slightly enhanced the severity of the disease by activating mast cells. Mast cells increased fibroblast proliferation, collagen production and contractile activity. CONCLUSIONS: Mast cells are involved in the gastrointestinal tract inflammation and fibrosis of the TNBS-colitis rats.

Defibrotide for the treatment of veno-occlusive disease after liver transplantation.

BACKGROUND: Veno-occlusive disease (VOD) after liver transplantation is associated with acute rejection and poor outcome. The use of antithrombotic and thrombolytic agents is limited by their toxicity. Defibrotide is a polydeoxyribonucleotide with thrombolytic and antithrombotic properties and no systemic anticoagulant effect. METHODS: Defibrotide, 35-40 mg/kg/day, was administered intravenously for 21 days on a compassionate-use basis to two patients aged 66 and 49 years. VOD had developed 6 weeks and 4 months after orthotopic liver transplantation for hepatitis C and hepatitis B infection, respectively. VOD was diagnosed clinically by findings of weight gain (8.5% and 16%), ascites, jaundice (serum bilirubin 5.4 mg/dl and 21.7 mg/dl), and severe coagulopathy (in one patient), and histologically by the presence of hemorrhagic centrilobular necrosis and fibrous stenosis of the hepatic venules. One of the patients had received azathioprine as part of the immunosuppressive regimen. There was no evidence of acute cellular rejection histologically. RESULTS: After 3 weeks of defibrotide administration, the first patient showed complete clinical resolution of the VOD, and serum bilirubin level normalized. He is alive 6 months after transplantation. The second patient, treated at a later stage of disease, showed marked improvement in the coagulopathic state, but there was no resolution of the VOD. He died 2 months later of multiorgan failure due to Escherichia coli sepsis. Neither patient had side effects from the drug. CONCLUSIONS: Defibrotide is a promising drug for the treatment of VOD after liver transplantation and needs to be evaluated in large, prospective studies.

Reduced body fat and increased hepatic lipid synthesis in mice bearing interleukin-6-secreting tumor.

Chronic secretion of interleukin-6 (IL-6) in mice causes metabolic alteration in the liver, leading to increased synthesis of hepatic cholesterol and fatty acids (FA). Mice were injected with allogeneic tumor cells transduced with the murine IL-6 gene. During the 3 wk after tumor inoculation, elevated serum IL-6 levels were associated with increased spleen and liver weight. Histological examination of sections from the liver showed increased hepatocyte proliferation, resulting in liver enlargement. Body composition analysis revealed that IL-6 caused a significant loss in fat tissue without affecting lean body mass and water content. Hepatic de novo synthesis of FA and cholesterol, as measured by (3)H(2)O incorporation, was three to five times as high in mice secreting IL-6 (IL-6 mice) as in pair-fed mice bearing nonsecreting tumors. This increase in FA and cholesterol synthesis is sufficient to maintain hepatic triglyceride secretion at levels comparable with those of pair-fed mice bearing nonsecreting tumors and, presumably, is the main source of cholesterol and FA-phospholipids necessary for hepatocyte proliferation.