Table of Contents for Volume 41, Issue 3 (March 2005)
Liver Biology and Pathobiology
Contribution of Toll-like receptor/myeloid differentiation factor 88 signaling to murine liver regeneration (p 443-450)
Ekihiro Seki, Hiroko Tsutsui, Yuji Iimuro, Tetsuji Naka, Gakuhei Son, Shizuo Akira, Tadamitsu Kishimoto, Kenji Nakanishi, Jiro Fujimoto
Toll-like receptors (TLRs) act as innate immune signal sensors and play central roles in host defense. Myeloid differentiation factor (MyD) 88 is a common adaptor molecule required for signaling mediated by TLRs. When the receptors are activated, cells bearing TLRs produce various proinflammatory cytokines in a MyD88-dependent manner. Liver regeneration following partial hepatectomy (PH) requires innate immune responses, particularly interleukin-6 (IL-6) and tumor necrosis factor (TNF-) production by Kupffer cells, although the recognition and activation processes are still unknown. We investigated whether TLR/MyD88 signaling is critical for induction of innate immune responses after PH. In Myd88-/- mice after PH, induction of expression of immediate early genes involved in hepatocyte replication and phosphorylation of STAT3 in the liver, and production of TNF-/IL-6 by and activation of NF-B in the Kupffer cells were grossly subnormal and were associated with impaired liver regeneration. However, TLR2, 4 and 9, which recognize gram-negative and -positive bacterial products, are not essential for NF-B activation and IL-6 production after PH, which excludes a possible contribution of TLR2/TLR4 or TLR9 to MyD88-mediated pathways. In conclusion, the TLR/MyD88 pathway is essential for incidental liver restoration, particularly its early phase. (HEPATOLOGY 2005;41:443-450.)
Lymphocyte traffic through sinusoidal endothelial cells is regulated by hepatocytes (p 451-459)
Sarah Edwards, Patricia F. Lalor, Gerard B. Nash, G. Ed Rainger, David H. Adams
Crosstalk between hepatic sinusoidal ECs and closely juxtaposed hepatocytes via vascular endothelial growth factor is essential for the maintenance of sinusoidal endothelial growth and differentiation. We propose that paracrine interactions between endothelial cells and hepatocytes also may be responsible for the unique complement of adhesion receptors expressed on sinusoidal endothelium that regulate the recruitment of lymphocytes into the liver. To address this hypothesis, we developed an in vitro model of the hepatic sinusoid in which flowing lymphocytes could interact with hepatic endothelium conditioned by the presence of hepatocytes. Human hepatic sinusoidal endothelial cells cocultured with hepatocytes were activated so that they supported the adhesion of lymphocytes at levels equivalent to those seen on endothelium stimulated with the inflammatory cytokine tumour necrosis factor-. Lymphocyte adhesion was supported by intracellular adhesion molecule 1, vascular cell adhesion molecule 1, and E-selectin, with an additional contribution from the novel adhesion receptor VAP-1. In conclusion, we show that interactions between hepatocytes and endothelial cells amplify leukocyte recruitment through the sinusoids by regulating the expression and function of endothelial adhesion molecules. These paracrine interactions may be responsible for the induction of the adhesion molecules that support constitutive lymphocyte recruitment to the liver as well as contributing significantly to the patterns of leukocyte adhesion seen during episodes of hepatic inflammation. (HEPATOLOGY 2005;41:451-459.)
Interplay among cardiotrophin-1, prostaglandins, and vascular endothelial growth factor in rat liver regeneration (p 460-469)
Naiara Beraza, Juan Martín Marqués, Eduardo Martínez-Ansó, María Iñiguez, Jesús Prieto, Matilde Bustos
Prostaglandins are hepatoprotective molecules generated in liver regeneration by the rapid induction of cyclooxygenase-2 (COX-2). Cardiotrophin-1 (CT-1) and vascular endothelial growth factor (VEGF) are other hepatoprotective mediators upregulated at 24 hours after partial hepatectomy. The interactions among these molecules during liver regeneration have not yet been defined. Here we show that rats subjected to partial hepatectomy treated with NS-398, a specific COX-2 inhibitor, exhibited cell cycle arrest, increased hepatocyte apoptosis, persistent extracellular signal-regulated kinase (ERK) 1/2 activation, and increased interleukin-6 production. These changes were associated with downregulation of CT-1 and COX-1 and altered pattern of VEGF expression. Administration of an adenovirus encoding CT-1 to NS-398-treated rats restituted normal levels of COX-1, prostaglandins, and VEGF in the liver after partial hepatectomy and restored normal liver regeneration. Furthermore, the stimulation of isolated rat hepatocytes with CT-1 increased COX-1, COX-2, and VEGF messenger RNAs and prostaglandin synthesis. Conversely, the addition of prostaglandin E1 to the culture increased CT-1 and VEGF production. In conclusion, COX-2 activation and production of prostaglandins soon after partial hepatectomy are essential requirements for hepatocyte proliferation and for the correct induction of both CT-1 and VEGF. CT-1 can restore liver regeneration after COX-2 inhibition by increasing VEGF, COX-1 expression, and prostaglandin synthesis. (HEPATOLOGY 2005;41:460-469.)
Cytokine-independent repression of rodent Ntcp in obstructive cholestasis (p 470-477)
Andreas Geier, Gernot Zollner, Christoph G. Dietrich, Martin Wagner, Peter Fickert, Helmut Denk, Nico van Rooijen, Siegfried Matern, Carsten Gartung, Michael Trauner
Cholestatic liver injury is associated not only with accumulation of bile acids but also with activation of proinflammatory cytokines. Common bile duct ligation (CBDL) induces sustained downregulation of the Na+/taurocholate cotransporter (Ntcp) in rodent liver. Although repression of Ntcp during endotoxemia is cytokine mediated, it is unclear whether inflammatory cytokines contribute to this downregulation in obstructive cholestasis. Cytokine inactivation in CBDL rats and mice was either performed directly with tumor necrosis factor alpha (etanercept) or interleukin 1 beta inactivation (anakinra/AMG 719) or indirectly Kupffer cell depletion via intraperitoneal administration of liposome-encapsulated dichloromethylene bisphosphonate. Protein and messenger RNA (mRNA) expression of Ntcp and short heterodimer partner (SHP) were analyzed via Western and Northern blotting. Key regulators of Ntcp (hepatocyte nuclear factor 1 alpha [HNF-1], HNF-4, retinoid X receptor alpha [RXR]:retinoic acid receptor alpha [RAR]) were studied via electrophoretic mobility shift analysis and nuclear Western blot analysis. Both methods of cytokine inactivation failed to maintain Ntcp protein or mRNA expression within 3 days after CBDL in either rats or mice (20%-40% of sham controls), while SHP mRNA expression increased three- to five-fold. Decreased nuclear HNF-1 and HNF-4 protein levels (45% and 60% of sham controls, respectively) and HNF-1 binding activity (32% of sham controls) were not restored during cytokine inactivation after CBDL, indicating cytokine-independent mechanisms of Ntcp regulation. RXR:RAR binding remained unchanged in all experimental conditions. In conclusion, during obstructive cholestasis accumulating bile acids per se, without major contribution of cytokines, leads to downregulation of Ntcp via repression of HNF-1 and HNF-4. (HEPATOLOGY 2005;41:470-477.)
TNF-mediated extracellular matrix remodeling is required for multiple division cycles in rat hepatocytes (p 478-486)
Anne-Laure Sérandour, Pascal Loyer, Delphine Garnier, Brice Courselaud, Nathalie Théret, Denise Glaise, Christiane Guguen-Guillouzo, Anne Corlu
During liver regeneration, hepatocytes proliferate under the control of both proinflammatory cytokines such as tumor necrosis factor (TNF) and growth factors, in parallel to extracellular matrix remodeling. This study investigated mechanisms by which mitogen and extracellular matrix signals are linked for inducing proliferation of differentiated hepatocytes. The authors used adult rat hepatocytes in coculture with liver biliary cells, because cells are stably differentiated for several weeks, capable of extracellular matrix deposition, and unable to divide in response to growth factor alone. This work demonstrated that hepatocytes could undergo several proliferation waves without loss of differentiation by using alternating periods of TNF/growth factor stimulation and deprivation. Three days after stimulation with TNF and epidermal growth factor (EGF), up to 35% of hepatocytes divided. Demonstration was also provided that EGF alone only promoted cell progression up to late G1, whereas TNF was necessary for G1/S transition and Cdk1 induction. TNF promoted an extracellular matrix (ECM) degradation that involved the matrix metalloproteinase MMP-9 induction through activation of NF-B pathway. Finally, the authors showed that ECM remodeling signal was required for initiating any new hepatocyte division wave, in presence of mitogen. In conclusion, these results highlight that hepatocyte division is dependent on ECM deposition associated with differentiation status, and that ECM degradation signal is critical in controlling G1/S transition and Cdk1 induction. These results provide new insights for understanding the unique hepatocyte proliferation control and improving regeneration in patients suffering from liver damage. (HEPATOLOGY 2005;41:478-486.)
Inducible inactivation of Notch1 causes nodular regenerative hyperplasia in mice (p 487-496)
Adrien Croquelois, Alex Blindenbacher, Luigi Terracciano, Xueya Wang, Igor Langer, Freddy Radtke, Markus H. Heim
Published Online: 18 Feb 2005
The discovery that the human Jagged1 gene (JAG1) is the Alagille syndrome disease gene indicated that Notch signaling has an important role in bile duct homeostasis. The functional study of this signaling pathway has been difficult because mice with targeted mutations in Jagged1, Notch1, or Notch2 have an embryonic lethal phenotype. We have previously generated mice with inducible Notch1 disruption using an interferon-inducible Cre-recombinase transgene in combination with the loxP flanked Notch1 gene. We used this conditional Notch1 knockout mouse model to investigate the role of Notch1 signaling in liver cell proliferation and differentiation. Deletion of Notch1 did not result in bile duct paucity, but, surprisingly, resulted in a continuous proliferation of hepatocytes. In conclusion, within weeks after Notch1 inactivation, the mice developed nodular regenerative hyperplasia without vascular changes in the liver. (HEPATOLOGY 2005;41:487-496.)
Dual role of orphan nuclear receptor pregnane X receptor in bilirubin detoxification in mice (p 497-505)
Simrat P. S. Saini, Ying Mu, Haibiao Gong, David Toma, Hirdesh Uppal, Songrong Ren, Song Li, Samuel M. Poloyac, Wen Xie
The pregnane X receptor (PXR) and the constitutive androstane receptor (CAR) are implicated in xenobiotic and endobiotic detoxification, including the clearance of toxic bilirubin. Previous studies have suggested both overlapping and preferential regulation of target genes by these receptors, but the mechanism of cross-talk remains elusive. Here we reveal a dual role of PXR in bilirubin detoxification in that both the loss and activation of PXR led to protection from hyperbilirubinemia induced by bilirubin infusion or hemolysis. The increased bilirubin clearance in PXR-null mice was associated with selective upregulation of detoxifying enzymes and transporters, and the pattern of regulation is remarkably similar to that of transgenic mice expressing the activated CAR. Interestingly, the increased bilirubin clearance and associated gene regulation were absent in the CAR-null or double-knockout mice. In cell cultures, ligand-free PXR specifically suppressed the ability of CAR to induce the multidrug resistance associated protein 2 (MRP2), a bilirubin-detoxifying transporter. This suppression was, at least in part, the result of the disruption of ligand-independent recruitment of coactivator by CAR. In conclusion, PXR plays both positive and negative roles in regulating bilirubin homeostasis, and this provides a novel mechanism that may govern receptor cross-talk and the hierarchy of xenobiotic and endobiotic regulation. PXR is a potential therapeutic target for clinical treatment of jaundice. (HEPATOLOGY 2005;41:497-505.)
Fractalkine and CX3CR1 are involved in the recruitment of intraepithelial lymphocytes of intrahepatic bile ducts (p 506-516)
Kumiko Isse, Kenichi Harada, Yoh Zen, Takashi Kamihira, Shinji Shimoda, Mine Harada, Yasuni Nakanuma
Fractalkine is a chemokine with both chemoattractant and cell-adhesive functions, and in the intestine it is involved with its receptor CX3CR1 in the chemoattraction and recruitment of intraepithelial lymphocytes. We examined the pathophysiological roles of fractalkine and CX3CR1 in normal and diseased bile ducts. Expression of fractalkine and CX3CR1 were examined in liver tissues from patients with primary biliary cirrhosis (17 cases) and controls (9 cases of primary sclerosing cholangitis, 10 cases of extrahepatic biliary obstruction, 20 cases of chronic viral hepatitis C, and 18 cases of histologically normal livers). Expression of fractalkine in biliary epithelial cells (BECs) in response to cytokine treatments was examined using a human cholangiocarcinoma cell line (HuCC-T1) and human intrahepatic BEC line. The chemotaxis of CX3CR1-expressing monocytes (THP-1) toward fractalkine was assayed using chemotaxis chambers. Fractalkine messenger RNA/protein were expressed on BECs of normal and diseased bile ducts, and their expression was upregulated in injured bile ducts of primary biliary cirrhosis. CX3CR1 was expressed on infiltrating mononuclear cells in portal tracts and on CD3+, CD4+, and CD8+ intraepithelial lymphocytes of injured bile ducts in primary biliary cirrhosis. Fractalkine messenger RNA expression was upregulated in two cultured BECs on treatment with lipopolysaccharide and Th1-cytokines (interleukin 1, interferon gamma, and tumor necrosis factor ). THP-1 cells showed chemotaxis toward fractalkine secreted by cultured cells. In conclusion, Th1-cytokine predominance and lipopolysaccharide in the microenvironment of injured bile ducts resulting from primary biliary cirrhosis induce the upregulation of fractalkine expression in BECs, followed by the chemoattraction of CX3CR1-expressing mononuclear cells, including CD4+ and CD8+ T cells, and their adhesion to BECs and the accumulation of biliary intraepithelial lymphocytes. (HEPATOLOGY 2005;41:506-516.)
Keratin mutation primes mouse liver to oxidative injury (p 517-525)
Qin Zhou, Xuhuai Ji, Lixin Chen, Harry B. Greenberg, Shelly C. Lu, M. Bishr Omary
Mutation of the cytoskeletal intermediate filament proteins keratin 8 and keratin 18 (K8/K18) is associated with cirrhosis in humans, whereas transgenic mice that overexpress K18 Arg89Cys (R89C) have significant predisposition to liver injury. To study the mechanism of keratin-associated predisposition to liver injury, we used mouse microarrays to examine genetic changes associated with hepatocyte keratin mutation and assessed the consequences of such changes. Liver gene expression was compared in R89C versus nontransgenic or wild-type K18-overexpressing mice. Microarray-defined genetic changes were confirmed by quantitative polymerase chain reaction. Nineteen genes had a more than two-fold altered expression (nine downregulated, 10 upregulated). Upregulated genes in keratin-mutant hepatocytes included the oxidative metabolism genes cytochrome P450, S-adenosylhomocysteine (SAH) hydrolase, cysteine sulfinic acid decarboxylase, and oxidation-reduction pathway genes. Downregulated genes included fatty acid binding protein 5, cyclin D1, and some signaling molecules. Several methionine metabolism-related and glutathione synthetic pathway intermediates, including S-adenosylmethionine (SAMe) and SAH, were modulated in R89C versus control mice. R89C livers had higher lipid and protein oxidation by-products as reflected by increased malondialdehyde and oxidized albumin. In conclusion, K18 point mutation in transgenic mice modulates several hepatocyte oxidative stress-related genes and leads to lipid and protein oxidative by-products. Mutation-associated decreases in SAH and SAMe could compromise needed cysteine availability to generate glutathione during oxidative stress. Hence keratin mutations may prime hepatocytes to oxidative injury, which provides a new potential mechanism for how keratin mutations may predispose patients to cirrhosis. (HEPATOLOGY 2005;41:517-525.)
Effective oral treatment of unconjugated hyperbilirubinemia in Gunn rats (p 526-534)
Anja M. Hafkamp, Rick Havinga, Maarten Sinaasappel, Henkjan J. Verkade
We sought to develop an oral treatment for unconjugated hyperbilirubinemia. In the Gunn rat model of unconjugated hyperbilirubinemia, dietary supplementation with the lipase inhibitor orlistat (Orl) or with calcium phosphate (CaP) decreases plasma unconjugated bilirubin (UCB) levels. We determined whether Orl, CaP, or their combination is superior to phototherapy, the conventional treatment, and whether the effects of Orl and CaP are influenced by dietary fat content. Gunn rats were treated with Orl (200 mg/kg chow), CaP (20 g/kg chow), Orl + CaP, or continuous phototherapy (19 W/cm2/nm) during a low-fat (LF) diet (13 energy%) or high-fat (HF) diet (35 energy%). Plasma UCB and fecal fat excretion were measured before, during, and/or at the end of treatment. Orl treatment for 2 weeks (HF diet) reduced plasma UCB concentrations similar to phototherapy (-34% and -28%, respectively); the combination of both was more effective than either treatment alone (-48%; P < .001). After 3 weeks of a HF diet, plasma UCB was 46% lower compared with the LF diet (P < .001). Plasma UCB concentrations were negatively correlated with fecal fat excretion (r = -0.96; P < .001). Irrespective of dietary fat content, 3 weeks of combined treatment (Orl + CaP) decreased plasma UCB by approximately 50% (P < .01) and was more effective than phototherapy (P < .05) at the intensity provided. In conclusion, plasma UCB concentrations in Gunn rats are negatively related to fecal fat excretion and dietary fat content. Orlistat is equally effective as phototherapy for the treatment of unconjugated hyperbilirubinemia in Gunn rats, and combined oral treatment with Orl + CaP is more effective than phototherapy. The present results support the feasibility of an efficient oral treatment of unconjugated hyperbilirubinemia. (HEPATOLOGY 2005;41:526-534.)
Transdifferentiation of rat hepatocytes into biliary cells after bile duct ligation and toxic biliary injury (p 535-544)
George K. Michalopoulos, Lindsay Barua, William C. Bowen
Rats with chimeric livers were generated by using the protocol of injecting hepatocytes from dipeptidyl peptidase IV (DPPIV)-positive donors into retrorsine-treated DPPIV-negative recipients subjected to partial hepatectomy. Rats with established chimeric livers were subjected to bile duct ligation, with or without pretreatment with the biliary toxin methylene diamiline (DAPM). Ductules bearing the donor hepatocyte marker DPPIV were seen at 30 days after bile duct ligation. The frequency of the ductules derived from the donor hepatocytes was dramatically enhanced (36-fold) by the pretreatment with DAPM. In conclusion, our results show that hepatocytes can function as facultative stem cells and rescue the biliary epithelium during repair from injury when its proliferative capacity is being compromised. (HEPATOLOGY 2005;41:535-544.)
Kupffer cells and macrophages are not required for hepatic hepcidin activation during iron overload (p 545-552)
Giuliana Montosi, Elena Corradini, Cinzia Garuti, Samuele Barelli, Stefania Recalcati, Gaetano Cairo, Linda Valli, Elisa Pignatti, Chiara Vecchi, Francesca Ferrara, Antonello Pietrangelo
Hepcidin, the iron hormone, is produced by the liver in response to iron and inflammation. Its synthesis during inflammation is triggered by cytokines, but the details of iron activation are obscure. We tested the role of Kupffer cells and macrophages by studying iron-loaded or inflamed mice with selective inactivation of Kupffer cells or the in vitro effect of conditioned human macrophages on hepcidin expression. Hepcidin messenger RNA (mRNA) expression was studied by Northern blot and reverse transcriptase polymerase chain reaction analysis in mice that were treated with 40 mg/kg gadolinium (III) chloride (GdCl3) as a Kupffer cell inactivating agent and subjected to inflammatory challenges with either lipopolysaccharide (LPS) and turpentine or iron overload by iron-dextran administration. Similar analyses were performed in human hepatoma cells (HepG2) cultured with medium from LPS- or iron-conditioned macrophages from blood donors or patients with HFE-linked hereditary hemochromatosis (HH). In vivo, LPS and particularly turpentine stimulated hepcidin mRNA expression, and this effect was prevented by the inactivation of Kupffer cells. Also, iron overload markedly upregulated hepatic hepcidin mRNA, but this activity persisted in spite of Kupffer cell blockade. In vitro, the medium of LPS-treated normal or hemocromatotic macrophages turned on hepcidin expression. On the contrary, medium of iron-manipulated macrophages, regardless of their HFE status, did not affect hepcidin mRNA steady-state levels. In conclusion, Kupffer cells are required for the activation of hepcidin synthesis during inflammation, and HH inflamed macrophages are capable of mounting a normal response, eventually leading to hepcidin stimulation. However, both Kupffer cells and human macrophages are dispensable for the regulatory activity exerted by iron on hepatic hepcidin. (HEPATOLOGY 2005;41:545-552.)
Liver Failure and Liver Disease
Evidence of normal thrombin generation in cirrhosis despite abnormal conventional coagulation tests (p 553-558)
Armando Tripodi, Francesco Salerno, Veena Chantarangkul, Marigrazia Clerici, Massimo Cazzaniga, Massimo Primignani, Pier Mannuccio Mannucci
The role played by coagulation defects in the occurrence of bleeding in cirrhosis is still unclear. This is partly due to the lack of tests that truly reflect the balance of procoagulant and anticoagulant factors in vivo. Conventional coagulation tests such as prothrombin time and activated partial thromboplastin time are inadequate to explore the physiological mechanism regulating thrombin, because they do not allow full activation of the main anticoagulant factor, protein C, whose levels are considerably reduced in cirrhosis. We used a thrombin generation test to investigate the coagulation function in patients with cirrhosis. Thrombin generation measured without thrombomodulin was impaired, which is consistent with the reduced levels of procoagulant factors typically found in cirrhosis. However, when the test was modified by adding thrombomodulin (i.e., the protein C activator operating in vivo), patients generated as much thrombin as controls. Hence, the reduction of procoagulant factors in patients with cirrhosis is compensated by the reduction of anticoagulant factors, thus leaving the coagulation balance unaltered. These findings help clarify the pathophysiology of hemostasis in cirrhosis, suggesting that bleeding is mainly due to the presence of hemodynamic alterations and that conventional coagulation tests are unlikely to reflect the coagulation status of these patients. In conclusion, generation of thrombin is normal in cirrhosis. For a clinical validation of these findings, a prospective clinical trial is warranted where the results of thrombin generation in the presence of thrombomodulin are related to the occurrence of bleeding. (HEPATOLOGY 2005;41:533-558.)
The human liver clears both asymmetric and symmetric dimethylarginine (p 559-565)
Michiel P.C. Siroen, Joost R. M. van der Sijp, Tom Teerlink, Cors van Schaik, Robert J. Nijveldt, Paul A. M. van Leeuwen
Asymmetric (ADMA) and symmetric dimethylarginine (SDMA) inhibit production of nitric oxide. The concentration of both dimethylarginines is regulated by urinary excretion, although ADMA, but not SDMA, is also subject to degradation by dimethylarginine dimethylaminohydrolase, which is highly expressed in the liver but also present in the kidney. The exact roles of the human liver and kidney in the metabolism of dimethylarginines are currently unknown. Therefore, we aimed to investigate renal and hepatic handling of ADMA and SDMA in detail in 24 patients undergoing hepatic surgery. To calculate net organ fluxes and fractional extraction (FE) rates, blood was collected from an arterial line, the portal vein, hepatic vein, and renal vein, and blood flow of the hepatic artery, portal vein, and renal vein was determined using Doppler ultrasound techniques. Results showed a significant net uptake (median [IQR]) of ADMA in both the liver (9.6 nmol/min [5.6-13.2]) and the kidney (12.1 nmol/min [1.3-17.1]). SDMA uptake was present not only in the kidney (12.7 nmol/min [3.5-25.4]), but also in the liver (7.7 nmol/min [2.8-16.4]). FE rates of ADMA for the liver and kidney were 5.0% (3.5%-7.4%) and 8.4% (1.3%-13.9%), respectively. For SDMA, hepatic and renal FE rates were 3.4% (2.1%-7.5%) and 12.5% (3.6%-16.2%), respectively. In conclusion, this study gives a detailed description of the hepatic and renal elimination of dimethylarginines and shows that the clearing of SDMA is not only confined to the kidney, but the human liver also takes up substantial amounts of SDMA from the portal and systemic circulation. (HEPATOLOGY 200541:559-565.)
Patients with cirrhosis and bare-stent TIPS may have increased risk of hepatocellular carcinoma (p 566-571)
Rafael Bañares, Oscar Núñez, María Escudero, Cristina Fernández, Javier Vaquero, Inmaculada Beceiro, Antonio Echenagusía, Gerardo Clemente, Leandro Santos
A trend toward a higher incidence of hepatocelullar carcinoma (HCC) in patients with cirrhosis treated with bare-stent transjugular intrahepatic portosystemic shunt (TIPS) has been observed in previous studies. To assess the influence of TIPS as a risk factor for developing HCC, we have compared the incidence of HCC in two retrospective cohorts of patients. The TIPS cohort (n = 138) included patients with cirrhosis who underwent TIPS placement for the treatment of portal hypertension-related complications; the non-TIPS cohort was composed of patients admitted at the hospital at the same time of TIPS insertion who were individually matched 1:1 according to age, sex, Child-Turcotte-Pugh class, and cause of cirrhosis. A stratified Cox model was used to assess risk of HCC development. The median time of follow-up was similar in TIPS and non-TIPS cohorts (30.3 [range, 7.8-119.5] and 31.4 [range, 7.8-110.8] months, respectively). The cumulative probability of developing HCC at 1, 3, and 5 years was 3%, 24%, and 34% for the TIPS cohort and 1%, 6%, and 25%, for the non-TIPS cohort, respectively (Breslow test = 5.23, P = .022). The adjusted hazard ratio was 1.52 (95% confidence interval, 1.06-2.19; P = .02). Hepatitis C virus infection and age were independent predictors of HCC development in patients without TIPS. In conclusion, patients with cirrhosis who are treated with TIPS may have a higher incidence of HCC. This observation suggests the need for a strict HCC surveillance program for these patients, especially if they are not expected to undergo a short- or medium-term liver transplantation. (HEPATOLOGY 2005;41:566-571.)
Variceal ligation plus nadolol compared with ligation for prophylaxis of variceal rebleeding: A multicenter trial (p 572-578)
Joaquin de la Peña, Enric Brullet, Eloy Sanchez-Hernández, Monserrat Rivero, Mercedes Vergara, Jose Luis Martin-Lorente, Covadonga Garcia Suárez, the EVL Study Group
Beta-Blockers and endoscopic variceal ligation (EVL) have proven to be valuable methods in the prevention of variceal rebleeding. The aim of this study was to compare the efficacy of EVL combined with nadolol versus EVL alone as secondary prophylaxis for variceal bleeding. Patients admitted for acute variceal bleeding were treated during emergency endoscopy with EVL or sclerotherapy and received somatostatin for 5 days. At that point, patients were randomized to receive EVL plus nadolol or EVL alone. EVL sessions were repeated every 10 to 12 days until the varices were eradicated. Eighty patients with cirrhosis (alcoholic origin in 66%) were included (Child-Turcotte-Pugh A, 15%; B, 56%; C, 29%). The median follow-up period was 16 months (range, 1-24 months). The variceal bleeding recurrence rate was 14% in the EVL plus nadolol group and 38% in the EVL group (P = .006). Mortality was similar in both groups: five patients (11.6%) died in the combined therapy group and four patients (10.8%) died in the EVL group. There were no significant differences in the number of EVL sessions to eradicate varices: 3.2 ± 1.3 in the combined therapy group versus 3.5 ± 1.3 in the EVL alone group. The actuarial probability of variceal recurrence at 1 year was lower in the EVL plus nadolol group (54%) than in the EVL group (77%; P = .06). Adverse effects resulting from nadolol were observed in 11% of the patients. In conclusion, nadolol plus EVL reduces the incidence of variceal rebleeding compared with EVL alone. A combined treatment could lower the probability of variceal recurrence after eradication. (HEPATOLOGY 2005;41:572-578.)
Effects of celecoxib and naproxen on renal function in nonazotemic patients with cirrhosis and ascites (p 579-587)
Joan Clària, Jeffrey D. Kent, Marta López-Parra, Ginés Escolar, Luís Ruiz-del-Arbol, Pere Ginès, Wladimiro Jiménez, Boris Vucelic, Vicente Arroyo
Nonselective inhibition of cyclooxygenase (COX) by nonsteroidal anti-inflammatory drugs frequently induces renal failure in decompensated cirrhosis. Studies in experimental cirrhosis suggest that selective inhibitors of the inducible isoform COX-2 do not adversely affect renal function. However, very limited information is available on the effects of these compounds on renal function in human cirrhosis. This investigation consists of a double-blind, randomized, placebo-controlled trial aimed at comparing the effects of the selective COX-2 inhibitor celecoxib (200 mg every 12 hours for a total of 5 doses) on platelet and renal function and the renal response to furosemide (40 mg intravenously) with those of naproxen (500 mg every 12 hours for a total of 5 doses) and placebo in 28 patients with cirrhosis and ascites. A significant reduction (P < .05) in glomerular filtration rate (113 ± 27 to 84 ± 22 mL/min), renal plasma flow (592 ± 158 to 429 ± 106 mL/min) and urinary prostaglandin E2 excretion (3430 ± 430 to 2068 ± 549 pg/min) and suppression of the diuretic (urine volume: 561 ± 128 to 414 ± 107 mL/h) and natriuretic (urine sodium: 53 ± 13 to 34 ± 10 mEq/h) responses to furosemide were observed in the group of patients treated with naproxen but not in the other two groups. Naproxen, but not celecoxib or placebo, significantly inhibited platelet aggregation (72% ± 8% to 47% ± 8%, P < .05) and thromboxane B2 production (41 ± 12 to 14 ± 5 pg/mL, P < .05). In conclusion, our results indicate that short-term administration of celecoxib does not impair platelet and renal function and the response to diuretics in decompensated cirrhosis. Further studies are needed to evaluate the long-term safety of this drug in cirrhosis. (HEPATOLOGY 2005;41:579-587.)
Pantoprazole reduces the size of postbanding ulcers after variceal band ligation: A randomized, controlled trial (p 588-594)
Nicholas J. Shaheen, Eugene Stuart, Sarah M. Schmitz, Kate L. Mitchell, Michael W. Fried, Steven Zacks, Mark W. Russo, Joseph Galanko, Roshan Shrestha
Elective esophageal variceal ligation (EVL) is performed to decrease the risk of variceal hemorrhage. Side effects of EVL include hemorrhage, chest pain, dysphagia, and odynophagia. Because gastric acid may exacerbate postbanding ulcers and delay healing, proton pump inhibition may decrease side effects associated with EVL. The aim of this study was to assess the efficacy of pantoprazole, a proton pump inhibitor, as an adjunct to elective EVL. We performed a double-blinded, randomized, placebo-controlled trial of pantoprazole after elective EVL. Subjects in the pantoprazole arm received 40 mg pantoprazole intravenously after EVL followed by 40 mg oral pantoprazole for 9 days. Control subjects received intravenous and oral placebo. Subjects underwent upper endoscopy 10 to 14 days after banding. Primary outcomes included the size and number of ulcers and the subjects' reports of dysphagia, chest pain, and heartburn. Forty-four subjects were randomized: 42 completed the protocol. At follow-up endoscopy, the mean number of ulcers was similar in the two groups. However, the ulcers in the pantoprazole group were on average half as large as in the placebo group (37 mm2 vs. 82 mm2, P < .01). Chest pain, dysphagia, and heartburn scores were not significantly different. Four subjects, all in the placebo group, had adverse outcomes, including 3 who bled from postbanding ulcers and 1 with sepsis. In conclusion, subjects receiving pantoprazole after elective EVL had significantly smaller postbanding ulcers on follow-up endoscopy than subjects receiving placebo. However, the total ulcer number and patient symptoms were not different between the groups. (HEPATOLOGY 2005;41:588-594.)
No relevant effect of ursodeoxycholic acid on cytochrome P450 3A metabolism in primary biliary cirrhosis (p 595-602)
Karin Dilger, Annette Denk, Malte H. J. Heeg, Ulrich Beuers
Induction of cytochrome P450 3A (CYP3A) has been suggested as a mechanism of action of ursodeoxycholic acid (UDCA) in cholestasis. CYP3A is of key importance in human drug metabolism, being involved in presystemic extraction of more than 50% of all drugs currently available and of various endogenous compounds. Therefore, we compared the induction potential of UDCA with that of the prototypical inducer rifampicin in a human model study with the CYP3A substrates budesonide and cortisol. Twelve patients with early-stage primary biliary cirrhosis and eight healthy volunteers were treated with UDCA (15 mg/kg daily) for 3 weeks and subsequently with rifampicin (600 mg/d) for 1 week. Extensive pharmacokinetic profiling of oral budesonide (3 mg) was performed by determination of budesonide and phase I metabolites (6-hydroxybudesonide, 16-hydroxyprednisolone) in plasma and urine at baseline and at the end of each treatment. In parallel, urinary 6-hydroxycortisol, a validated marker of CYP3A induction, was determined. UDCA did not affect biotransformation of budesonide and urinary excretion of 6-hydroxycortisol either in patients or in healthy volunteers. Ratios of areas under plasma concentration-time curves (AUC0-12 h during UDCA/AUC0-12 h before UDCA) of both metabolites were not higher than those of budesonide itself. In contrast, administration of rifampicin markedly induced CYP3A metabolism, resulting in abolished budesonide plasma levels and high urinary excretion of 6-hydroxycortisol. Metabolite formation was enhanced by rifampicin, but not by UDCA (e.g., AUC16-hydroxyprednisolone/AUCbudesonide in patients: baseline, 8.6 ± 3.9; UDCA, 10.7 ± 7.1; rifampicin, 527.0 ± 248.7). In conclusion, UDCA is not a relevant inducer of CYP3A enzymes in humans. (HEPATOLOGY 2005;41:595-602.)
Risk factors for thrombophilia in extrahepatic portal vein obstruction (p 603-608)
Massimo Primignani, Ida Martinelli, Paolo Bucciarelli, Tullia Battaglioli, Raffaella Reati, Federica Fabris, Alessandra Dell'Era, Emanuela Pappalardo, Pier Mannuccio Mannucci
Scant information exists on the role of thrombophilia in extrahepatic portal vein obstruction (EHPVO). We studied 65 patients with EHPVO, 500 with deep vein thrombosis (DVT) of the lower limbs, and 700 healthy controls referred for thrombophilia screening, including the search for gain-of-function mutations in genes encoding coagulation factor V (factor V Leiden) and prothrombin (prothrombin G20210A); antithrombin, protein C, and protein S deficiency; and hyperhomocysteinemia. At least one abnormality in the thrombophilia screening was found in 40% of patients with either EHPVO or lower limb DVT and in 13% of controls, for odds ratios of 4.0 (95% CI, 2.3-7.0) and 4.4 (95% CI, 3.3-5.9), respectively. Statistically significant associations with EHPVO were observed for the prothrombin G20210A mutation (odds ratio, 8.1; 95% CI, 3.8-17.5) and the deficiencies of antithrombin, protein C, or protein S taken together (odds ratio, 4.5; 95% CI, 1.1-18.0). The odds ratio for the prothrombin G20210A was approximately twice that for lower limb DVT. Patients with factor V Leiden had an odds ratio for EHPVO of 0.8 (95% CI, 0.1-6.4) and for lower limb DVT of 7.5 (95% CI, 4.4-13.0). The odds ratio for EHPVO in patients with hyperhomocysteinemia was 2.0 (95% CI, 0.9-4.9). At variance with lower limb DVT, oral contraceptive use was not associated with an increased risk of EHPVO. Myeloproliferative disorders were diagnosed in 35% of patients with EHPVO. In conclusion, the risk for EHPVO is increased in the presence of thrombophilia resulting from the prothrombin G20210A mutation and from the deficiencies of the naturally occurring anticoagulant proteins, but not from factor V Leiden. (HEPATOLOGY 2005;41:603-608.)
Small ubiquitin-related modifiers: A novel and independent class of autoantigens in primary biliary cirrhosis (p 609-616)
Caroline Janka, Carlo Selmi, M. Eric Gershwin, Hans Will, Thomas Sternsdorf
Published Online: 22 Feb 2005
Serum autoantibodies against components of nuclear dots (anti-NDs), namely PML and Sp100, are specifically detected in 20% to 30% of patients with primary biliary cirrhosis (PBC). Although anti-ND antibodies are nonpathogenic, the mechanisms that lead to this unique reactivity are critical to understanding the loss of immune tolerance in PBC. Importantly, Sp100 and PML are both covalently linked to small ubiquitin-related modifiers (SUMOs). Therefore, we investigated whether SUMO proteins are independent autoantigens in PBC and studied 99 PBC sera samples for reactivity against NDs, PML, and Sp100, as well as against SUMO-2 and SUMO-1 recombinant proteins. Autoantibodies against SUMO-2 and SUMO-1 were found in 42% and 15% of anti-ND-positive PBC sera, respectively. Anti-SUMO reactivity was not observed in anti-ND-negative sera. Anti-SUMO-2 autoantibodies were found in 58% of sera containing autoantibodies against both PML and Sp100 and were detected exclusively in sera containing anti-Sp100 autoantibodies. In conclusion, SUMO proteins constitute a novel and independent class of autoantigens in PBC. Furthermore, we believe our data emphasize the post-translational modification of lysine by either lipoylation in the case of AMA or SUMOylation in the case of specific anti-ND autoantibodies as the pivotal site for autoantibody generation in PBC. (HEPATOLOGY 2005 2005;41:609-616.)
Viral Hepatitis
Associations among clinical, immunological, and viral quasispecies measurements in advanced chronic hepatitis C (p 617-625)
Alan Lee Rothman, Chihiro Morishima, Herbert L. Bonkovsky, Stephen J. Polyak, Ranjit Ray, Adrian M. Di Bisceglie, Karen L. Lindsay, Peter F. Malet, Ming Chang, David R. Gretch, Daniel G. Sullivan, Atul K. Bhan, Elizabeth C. Wright, Margaret James Koziel, the HALT-C Trial Group
The relationships among host immune and viral factors and the severity of liver disease due to hepatitis C virus (HCV) are poorly understood. Previous studies have focused on individual components of the immune response to HCV, often in relatively small numbers of patients. We measured HCV-specific lymphoproliferation (LP), intrahepatic cytotoxic T lymphocyte (CTL), and neutralizing antibody (NA) responses and HCV quasispecies (QS) diversity and complexity in a large cohort of subjects with advanced liver fibrosis (Ishak stages 3-6) on entry into the HALT-C (Hepatitis C Antiviral Long-term Treatment against Cirrhosis) trial. We correlated LP, CTL, NA, and QS results with clinical characteristics, including serum alanine aminotransferase (ALT), HCV RNA level, HCV genotype, and hepatic histopathology. LP, CTL, and NA responses were detected in 37%, 22%, and 22% of subjects tested, respectively. The only association that was statistically significant was higher mean serum ALT values in patients with detectable HCV-specific CTL responses (P = .03). In conclusion, immune responses to HCV and viral diversity showed little relationship to clinical or histological features at a single time point in this selected population of patients with advanced chronic hepatitis C for whom prior interferon treatment had failed. (HEPATOLOGY 2005;41:617-625.)
G to A hypermutation of hepatitis B virus (p 626-633)
Chiemi Noguchi, Hiromi Ishino, Masataka Tsuge, Yoshifumi Fujimoto, Michio Imamura, Shoichi Takahashi, Kazuaki Chayama
G to A hypermutation of the human immunodeficiency virus type 1 (HIV-1) is induced by a deaminase APOBEC3G and is related to host antiviral defense. APOBEC3G has also been found to reduce the replication of HIV-1 by an unknown mechanism. This enzyme also reduces the production of hepatitis B virus, although the mechanism for this action has not been clearly elucidated. The hypermutated hepatitis B virus (HBV) is rarely found in usual sequencing analyses. Using peptide nucleic acid mediated by polymerase chain reaction clamping, we detected the hypermutated HBV DNA in 1 of 8 patients with acute HBV infection and 4 of 10 with chronic HBV infection. In the latter group, hypermutated genomes were found only in eAb-positive patients. As much as 72.5% of G residues were mutated in the hypermutated clones. G to A substitutions were predominant in almost all clones sequenced compared with other substitutions. G to A mutated viral genomes also were found in HepG2-derived cell lines that continuously produced HBV into the supernatant. Both alpha and gamma interferon reduced virus production in these cell lines, but they did not alter the frequency of the hypermutation. Transcripts of APOBEC3G, as well as some other deaminases, were found in these cell lines. In conclusion, our results show that part of the minus strand DNA of HBV is hypermutated both in vitro (HepG2 cell lines) and in vivo. The role and mechanism of hypermutation in reducing HBV replication should be further investigated to understand the anti-HBV defense system. (HEPATOLOGY 2005;41:626-633.)
SELDI-TOF MS profiling of serum for detection of the progression of chronic hepatitis C to hepatocellular carcinoma (p 634-642)
E. Ellen Schwegler, Lisa Cazares, Laura F. Steel, Bao-Ling Adam, David A. Johnson, O. John Semmes, Timothy M. Block, Jorge A. Marrero, Richard R. Drake
Proteomic profiling of serum is an emerging technique to identify new biomarkers indicative of disease severity and progression. The objective of our study was to assess the use of surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS) to identify multiple serum protein biomarkers for detection of liver disease progression to hepatocellular carcinoma (HCC). A cohort of 170 serum samples obtained from subjects in the United States with no liver disease (n = 39), liver diseases not associated with cirrhosis (n = 36), cirrhosis (n = 38), or HCC (n = 57) were applied to metal affinity protein chips for protein profiling by SELDI-TOF MS. Across the four test groups, 38 differentially expressed proteins were used to generate multiple decision classification trees to distinguish the known disease states. Analysis of a subset of samples with only hepatitis C virus (HCV)-related disease was emphasized. The serum protein profiles of control patients were readily distinguished from each HCV-associated disease state. Two-way comparisons of chronic hepatitis C, HCV cirrhosis, or HCV-HCC versus healthy had a sensitivity/specificity range of 74% to 95%. For distinguishing chronic HCV from HCV-HCC, a sensitivity of 61% and a specificity of 76% were obtained. However, when the values of known serum markers fetoprotein, des-gamma carboxyprothrombin, and GP73 were combined with the SELDI peak values, the sensitivity and specifity improved to 75% and 92%, respectively. In conclusion, SELDI-TOF MS serum profiling is able to distinguish HCC from liver disease before cirrhosis as well as cirrhosis, especially in patients with HCV infection compared with other etiologies. (HEPATOLOGY 2005;41:634-642.)
Plasmacytoid dendritic cells in acute and chronic hepatitis C virus infection (p 643-651)
Axel Ulsenheimer, J. Tilman Gerlach, Maria-Christina Jung, Norbert Gruener, Martin Wächtler, Markus Backmund, Teresa Santantonio, Winfried Schraut, Malte H. J. Heeg, Carl A. Schirren, Reinhart Zachoval, Gerd R. Pape, Helmut M. Diepolder
Chronic evolution of acute hepatitis C (aHC) occurs in more than 80% of patients but can frequently be prevented by early treatment with interferon (IFN)-. Plasmacytoid dendritic cells (pDCs) are the major endogenous IFN- producers, but their role in aHC is unknown. In this study, frequency, phenotype, and pDC function were analyzed in 13 patients with aHC and 32 patients with chronic hepatitis C (cHC) compared with 20 healthy controls, 33 sustained responders to antiviral treatment, 14 patients with acute hepatitis B (aHB), and 21 patients with nonviral inflammatory disease. In aHC, pDCs in the peripheral blood were significantly reduced compared with healthy controls (median, 0.1% vs. 0.36%, P < .0005) and were inversely correlated to alanine aminotransferase levels (r = -0.823; P < .005). Circulating pDCs in aHC were immature, as determined via reduced expression of HLA-DR and CCR7, and produced little amounts of IFN- (median, 3.5 pg/50,000 peripheral blood mononuclear cells [PBMCs] vs. 498.4 pg/50,000 PBMCs in healthy controls; P < .0005). Less pronounced changes were present in cHC (median, 0.17%, 28.0 pg/50,000 PBMCs IFN-, respectively). However, a significantly reduced frequency and IFN- production was also found in self-limited aHB (median 0.1%, 8.6 pg/50,000 PBMCs) and in patients with nonviral inflammatory disease (median 0.19%, 7.5 pg/50,000 PBMCs). In conclusion, in aHC frequency and IFN--producing capacity of peripheral blood pDCs are dramatically reduced and inversely correlated with the degree of liver inflammation. In cHC there is incomplete recovery of pDC function, which, however, could be solely due to the chronic inflammatory state. (HEPATOLOGY 2005;41:643-651.)
Non-Hodgkin's lymphoma and other nonhepatic malignancies in Swedish patients with hepatitis C virus infection (p 652-659)
Ann-Sofi Duberg, Marie Nordström, Anna Törner, Olle Reichard, Reinhild Strauss, Ragnhild Janzon, Erik Bäck, Karl Ekdahl
The aim of this study was to evaluate the association between hepatitis C virus (HCV) infection and non-Hodgkin's lymphoma (NHL), multiple myeloma (MM), thyroid cancer (TC), chronic lymphatic leukemia (CLL), acute lymphatic leukemia (ALL), and Hodgkin's lymphoma (HL). A Swedish cohort of 27,150 HCV-infected persons notified during 1990-2000 was included in the study. The database was linked to other national registers to calculate the observation time, expressed as person-years, and to identify all incident malignancies in the cohort. The patients were stratified according to assumed time of previous HCV infection. The relative risk of malignancy was expressed as a standardized incidence ratio (SIR) - the observed number compared to the expected number. During 1990-2000 there were 50 NHL, 15 MM, 14 ALL, 8 TC, 6 CLL, and 4 HL diagnoses in the cohort. Altogether, 20 NHL, 7 MM, 5 TC, 4 CLL, 1 ALL, and 1 HL patient fulfilled the criteria to be included in the statistical analysis. The observation time was 122,272 person-years. The risk of NHL and MM was significantly increased in the stratum with more than 15 years of infection (SIR 1.89 [95% CI, 1.10-3.03] and 2.54 [95% CI, 1.11-5.69], respectively). The association was not significant in TC or CLL. In conclusion, we report the incidence of several malignancies in a nationwide cohort of HCV-infected persons. Although the delayed diagnosis of HCV probably has resulted in an underestimation of the risk, this study showed a significantly increased risk of NHL and MM. (HEPATOLOGY 2005;41:652-659.)
Hepatitis C virus acts as a tumor accelerator by blocking apoptosis in a mouse model of hepatocarcinogenesis (p 660-667)
Yoshitaka Kamegaya, Yoichi Hiasa, Lawrence Zukerberg, Nina Fowler, Jason T. Blackard, Wenyu Lin, Won H. Choe, Emmett V. Schmidt, Raymond T. Chung
We developed hepatitis C virus (HCV) core-E1-E2 and HCV core transgenic mice on a common genetic background to assess the contribution of HCV structural proteins to hepatocarcinogenesis. Eight-week-old core-E1-E2, core, and nontransgenic mice inbred on the FVB?C57Bl/6 background were treated with diethylnitrosamine (DEN) and sacrificed at 32 weeks old. Proliferation and apoptosis were assessed by immunohistochemistry. The effect of viral proteins on apoptosis was evaluated in HepG2 cells in which apoptosis was induced by anti-Fas antibody. HCCs were identified at 32 weeks in the majority of DEN-treated mice from all three groups. The mean size of HCCs was significantly larger in core-E1-E2 transgenic (4.63 ± 1.48 mm), compared with core transgenic (0.78 ± 0.26 mm, P = .01), and nontransgenic (1.0 ± 0.19 mm, P = .002) mice. While there were no differences in proliferation, the apoptotic index in core-E1-E2 transgenic HCCs was significantly lower than those found in core and non-transgenic HCCs. Core-E1-E2 transfected HepG2 cells demonstrated a significantly lower apoptotic index (0.35 ± 0.11) compared with that of core transfected cells (0.74 ± 0.07, P = .0103). Analysis of a Fas-induced apoptosis model in HCV transgenic mice confirmed that core-E1-E2 transgenic liver underwent significantly less apoptosis than transgenic tissue expressing core only. In conclusion, HCV core-E1-E2 transgenic mice develop significantly larger tumors than transgenic mice expressing core alone or nontransgenic mice. The accelerated tumor phenotype is attributable to suppression of apoptosis rather than enhanced proliferation. These data implicate HCV E1 and/or E2 in conjunction with core as antiapoptotic, tumor accelerator proteins. (HEPATOLOGY 2005; 41:660-667.)
Concise Communication
Wilson disease in septuagenarian siblings: Raising the bar for diagnosis (p 668-670)
Aftab Ala, Jimo Borjigin, Arnold Rochwarger, Michael Schilsky
Wilson Disease (WD) usually presents in the first decades of life, although rare patients have a later presentation. We report the clincial features, diagnostic evaluation, and outcome with treatment of two septuagenarian siblings evaluated as part of a research trial for treatment of neurological WD. The index case was a 72-year-old woman who suffered progressive neurological disability, then developed sub-fulminant liver failure. Her sibling was a 70-year-old man with minimal neurological symptoms and a mild depressive disorder. His liver biopsy revealed only steatosis and minimal fibrosis and an elevated hepatic copper content (671 g/g dry weight liver). Molecular studies demonstrated compound heterozygosity for disease specific ATP7B mutations E1064A and H1069Q in both patients. Both individuals were treated with trientine and Zn followed by Zn maintenance therapy. Over the last 5 years, the clincial course stabilized and improved, although the index case recently died from bronchopneumonia. In conclusion, advanced age and different clinical presentations of these two subjects with identical ATP7B mutations raises the question of the degree of penetrance for these and other ATP7B mutations. Environmental and extragenic factors are pivotal determinants of disease phenotype. We suggest that WD must be considered at all ages in patients with hepatic disease, neurological disease, or psychiatric symptoms. (HEPATOLOGY 2005;41:668-670.)
Hepatology Elsewhere
Strong reasons make strong actions - The antiviral efficacy of NS3/4A protease inhibitors (p 671-674)
Stanley M. Lemon, MinKyung Yi, Kui Li
Background: Novel, potent and well-tolerated hepatitis C (HCV) drugs are still needed. BILN 2061 is a potent and specific inhibitor of HCV serine protease in vitro. Pre-clinical toxicology data and studies in healthy volunteers supported the administration of BILN 2061 to patients with HCV infection. Methods: The antiviral efficacy, pharmacokinetics and tolerability of 25, 200 and 500 mg bid BILN 2061 given as monotherapy for two days in 31 patients infected with chronic genotype 1 HCV infection and with minimal liver fibrosis (Ishak score 0-2) were assessed in a placebo controlled double blind pilot study. In two, subsequent, placebo controlled studies of similar design, 200 mg bid BILN 2061 was administered for two days to 10 patients with advanced liver fibrosis (Ishak score 3-4) and to 10 patients with compensated cirrhosis (Ishak 5-6). Results :Viral RNA reductions of 2-3 log10 copies/mL were achieved in most of the patients. There was a trend towards a higher number of patients receiving 500 mg BILN 2061 achieving a viral RNA reduction >3 log10 copies/mL as compared to patients receiving 25 mg BILN 2061. Advanced fibrosis or compensated cirrhosis did not affect the antiviral efficacy of BILN 2061. BILN 2061 was well tolerated in all studies. Conclusions: BILN 2061 is a well tolerated and very active compound, which reduced serum viral RNA concentrations after two days of treatment in patients infected with genotype 1 HCV independent from the degree of fibrosis. Nevertheless further clinical trials are on hold pending resolution of animal toxicity issues.
Hepatitis C virus and the threshold of natural killer cell inhibition (p 675-677)
Golo Ahlenstiel, Barbara Rehermann
Natural killer (NK) cells provide a central defense against viral infection by using inhibitory and activation receptors for major histocompatibility complex class I molecules as a means of controlling their activity. We show that genes encoding the inhibitory NK cell receptor KIR2DL3 and its human leukocyte antigen C group 1 (HLA-C1) ligand directly influence resolution of hepatitis C virus (HCV) infection. This effect was observed in Caucasians and African Americans with expected low infectious doses of HCV but not in those with high-dose exposure, in whom the innate immune response is likely overwhelmed. The data strongly suggest that inhibitory NK cell interactions are important in determining antiviral immunity and that diminished inhibitory responses confer protection against HCV.
Copyright © 2005 by the American Association for the Study of Liver Diseases. All rights reserved.
Table of Contents for March 2005 • Volume 128 • Number 3
Rapid Communication
Nonresorbable copolymer implantation for gastroesophageal reflux disease: A randomized sham-controlled multicenter trial
Jacques Devière, Guido Costamagna, Horst Neuhaus, Winfried Voderholzer, Hubert Louis, Andrea Tringali, Michele Marchese, Thomas Fiedler, Patrick Darb-Esfahani, Brigitte Schumacher
Background & Aims: This aim was to determine whether endoscopic implantation of a biocompatible nonresorbable copolymer (Enteryx; Boston Scientific Corp, Natick, MA) is a more effective therapy for gastroesophageal reflux disease (GERD) than a sham procedure. Methods: In a randomized, single-blind, prospective, multicenter clinical trial, 64 patients with GERD were enrolled whose symptoms were well controlled by proton pump inhibitor (PPI) therapy and rapidly recurred after cessation of PPI therapy. Thirty-two patients were assigned to Enteryx implantation and 32 to a sham procedure consisting of standard upper endoscopy. Patients in both groups with unsatisfactory symptom relief after 3 months were eligible for re-treatment by Enteryx implantation. The primary study end point was ≥50% reduction in PPI use. Secondary end points included ≥50% improvement in GERD score and the proportion of patients not undergoing re-treatment procedure. Follow-up evaluations were performed at 3 and 6 months. Results: The percentage of Enteryx-treated patients achieving a ≥50% reduction in PPI use (81%) was greater than that of the sham group (53%), with a rate ratio of 1.52 (confidence interval [CI], 1.06–2.28; P = .023). A higher proportion of the Enteryx (68%) than sham group (41%) ceased PPI use completely (rate ratio, 1.67; CI, 1.03–2.80; P = .033). GERD health-related quality of life heartburn score improvement ≥50% was achieved by 67% of the Enteryx group versus 22% of the sham group (rate ratio, 3.05; CI, 1.55–6.33; P < .001). More Enteryx-treated (81%) than sham-treated (19%) patients did not undergo re-treatment (rate ratio, 4.33; CI, 2.23–9.29; P < .001). Conclusions: Enteryx implantation more effectively reduces PPI dependency and alleviates GERD symptoms than a sham procedure.
Clinical-alimentary Tract
Lactobacillus and bifidobacterium in irritable bowel syndrome: Symptom responses and relationship to cytokine profiles
Liam O’Mahony, Jane McCarthy, Peter Kelly, George Hurley, Fangyi Luo, Kersang Chen, Gerald C. O’Sullivan, Barry Kiely, J. Kevin Collins, Fergus Shanahan, Eamonn M.M. Quigley
Background&Aims: The aim of this study was to compare the response of symptoms and cytokine ratios in irritable bowel syndrome (IBS) with ingestion of probiotic preparations containing a lactobacillus or bifidobacterium strain. Methods: Seventy-seven subjects with IBS were randomized to receive either Lactobacillus salivarius UCC4331 or Bifidobacterium infantis 35624, each in a dose of 1 ? 1010 live bacterial cells in a malted milk drink, or the malted milk drink alone as placebo for 8 weeks. The cardinal symptoms of IBS were recorded on a daily basis and assessed each week. Quality of life assessment, stool microbiologic studies, and blood sampling for estimation of peripheral blood mononuclear cell release of the cytokines interleukin (IL)-10 and IL-12 were performed at the beginning and at the end of the treatment phase. Results: For all symptoms, with the exception of bowel movement frequency and consistency, those randomized to B infantis 35624 experienced a greater reduction in symptom scores; composite and individual scores for abdominal pain/discomfort, bloating/distention, and bowel movement difficulty were significantly lower than for placebo for those randomized to B infantis 35624 for most weeks of the treatment phase. At baseline, patients with IBS demonstrated an abnormal IL-10/IL-12 ratio, indicative of a proinflammatory, Th-1 state. This ratio was normalized by B infantis 35624 feeding alone. Conclusions: B infantis 35624 alleviates symptoms in IBS; this symptomatic response was associated with normalization of the ratio of an anti-inflammatory to a proinflammatory cytokine, suggesting an immune-modulating role for this organism, in this disorder.
Clinical-alimentary Tract
Autologous hematopoietic stem cell transplantation in patients with refractory Crohn’s disease
Yu Oyama, Robert M. Craig, Ann E. Traynor, Kathleen Quigley, Laisvyde Statkute, Amy Halverson, Mary Brush, Larissa Verda, Barbara Kowalska, Nela Krosnjar, Morris Kletzel, Peter F. Whitington, Richard K. Burt
Background & Aims: Crohn’s disease (CD) is an immunologically mediated inflammatory disease of the gastrointestinal tract. Due to a high morbidity and/or an increase in mortality in refractory cases, a new treatment approach is needed. In theory, maximum immune ablation by autologous hematopoietic stem cell transplantation (HSCT) can induce a remission. Methods: We conducted a phase 1 HSCT study in 12 patients with refractory CD. Candidates were younger than 60 years of age with a Crohn’s Disease Activity Index (CDAI) of 250–400 despite conventional therapies including infliximab. Peripheral blood stem cells were mobilized with cyclophosphamide and granulocyte colony-stimulating factor and CD34+ enriched. The immune ablative (conditioning) regimen consisted of 200 mg/kg cyclophosphamide and 90 mg/kg equine antithymocyte globulin. Results: The procedure was well tolerated with anticipated cytopenias, neutropenic fever, and disease-related fever, diarrhea, anorexia, nausea, and vomiting. The median days for neutrophil and platelet engraftment were 9.5 (range, 8–11) and 9 (range, 9–18), respectively. The initial median CDAI was 291 (range, 250–358). Symptoms and CDAI improved before hospital discharge, whereas radiographic and colonoscopy findings improved gradually over months to years following HSCT. Eleven of 12 patients entered a sustained remission defined by a CDAI ≤150. After a median follow-up of 18.5 months (range, 7–37 months), only one patient has developed a recurrence of active CD, which occurred 15 months after HSCT. Conclusions: Autologous HSCT may be performed safely and has a marked salutary effect on CD activity. A randomized study will be needed to confirm the efficacy of this therapy.
Regional brain activation during proximal stomach distention in humans: A positron emission tomography study
Joris Vandenbergh, Patrick DuPont, Benjamin Fischler, Guy Bormans, Philippe Persoons, Jozef Janssens, Jan Tack
Background & Aims: Hypersensitivity to proximal gastric distention due to abnormal central nervous system processing of visceral stimuli has been suggested as a possible underlying pathophysiologic mechanism in functional dyspepsia. However, the cortical regions activated by distention of the proximal stomach have not been identified. The aim of this study was to investigate regional brain activation during painful and nonpainful proximal gastric distention in humans. Methods: Positron emission tomography of the brain was performed in 11 healthy volunteers during 4 conditions: (1) no distention and isobaric distention to the individual thresholds for (2) first, (3) marked, and (4) unpleasant sensation. Data were analyzed using statistical parametric mapping. Results: During maximal distention relative to baseline, significant (Pcorrected < .05) regional brain activation occurred in the left and right gyrus postcentralis (Brodmann area [BA] 43), the left gyrus temporalis superior (BA 38), the right gyrus frontalis inferior (BA 47, orbitofrontal cortex), the right midanterior cingulate gyrus (BA 24), the right anterior insula, and the left cerebellar hemisphere. These areas showed a progressive increase in activation with increasing intensity of the distending stimulus. Conclusions: We found evidence for a neuronal network processing distention stimuli of the proximal stomach that is overall consistent with the “visceral stimulation network” described in the literature. In addition, we found activation of the orbitofrontal cortex, confirming its role as a convergence zone for processing of food-related stimuli and regulation of hunger, appetite, satiety, and food intake. We found no evidence for a functional neuroanatomic divergence in the processing of noxious and innocuous gastric stimuli.
Origin of gas retention and symptoms in patients with bloating
Beatrice Salvioli, Jordi Serra, Fernando Azpiroz, Carlos Lorenzo, Santiago Aguade, Joan Castell, Juan-R. Malagelada
Background & Aims: Patients reporting abdominal bloating exhibit impaired tolerance to intestinal gas loads. The aim of this study was to identify the gut compartment responsible for gas retention. Methods: In 30 patients predominantly reporting abdominal bloating (24 with irritable bowel syndrome and 6 with functional bloating) and 22 healthy subjects, gas (nitrogen, carbon dioxide, and oxygen) was infused into the intestine for 2 hours while measuring rectal gas outflow. First, in 12 patients and 10 healthy subjects, gas transit (24 mL/min jejunal infusion labeled with 74 MBq bolus of 133Xe) was measured by scintigraphy. Second, in groups of patients and healthy subjects, the effects of gas infusion (12 mL/min) in the jejunum versus ileum, jejunum versus cecum, and jejunum versus sham infusion (n = 6 each) were compared by paired tests. Results: In patients, total gut transit of gas was delayed (50% clearance time, 33 ± 4 min vs 23 ± 4 min in healthy subjects; P < .05) owing to impaired small bowel transit (50% clearance time, 20 ± 2 min vs 12 ± 3 min in healthy subjects; P < .05), whereas colonic transit was normal (50% clearance time, 13 ± 2 min vs 11 ± 2 min in healthy subjects; not significant). Furthermore, jejunal gas infusion in patients was associated with gas retention (329 ± 81 mL vs 88 ± 79 mL in healthy subjects; P < .05), whereas direct ileal or colonic infusion was not (61 ± 103 mL and ?143 ± 87 mL retention, respectively). Conclusions:: In patients reporting bloating, the small bowel is the gut region responsible for ineffective gas propulsion.
A prospective assessment of bowel habit in irritable bowel syndrome in women: Defining an alternator
Douglas A. Drossman, Carolyn B. Morris, Yuming Hu, Brenda B. Toner, Nicholas Diamant, Jane Leserman, Michael Shetzline, Christine Dalton, Shrikant I. Bangdiwala
Background & Aims: Irritable bowel syndrome (IBS) is subtyped as IBS with diarrhea (IBS-D) or IBS with constipation (IBS-C) based on Rome II guidelines. The remaining group is considered as having mixed IBS (IBS-M). There is no standard definition of an alternator (IBS-A), in which bowel habit changes over time. Our aim was to use Rome II criteria to prospectively assess change in bowel habit for more than 1 year to understand IBS-A. Methods: Female patients (n = 317) with IBS entering a National Institutes of Health treatment trial were studied at baseline with questionnaires and 2-week daily diary cards of pain and stool frequency and consistency. Studies were repeated at the end of treatment (3 months) and at four 3-month intervals for one more year. Algorithms to classify subjects into IBS-D, IBS-C, and IBS-M groups used diary card information and modified Rome II definitions. Changes in bowel habit at 3-month intervals were then assessed using these surrogate diary card measures. Results: At baseline, 36% had IBS-D, 31% IBS-M, and 34% IBS-C. Except for stool frequency, there were no differences between groups. While the proportion of subjects in each subgroup remained the same over the year, most individuals (more than 75%) changed to either of the other 2 subtypes at least once. IBS-M was the least stable (50% changed out by 12 weeks). Patients were more likely to transition between IBS-M and IBS-C than between IBS-D and IBS-M. Notably, only 29% switched between the IBS-D and IBS-C subtypes over the year. Conclusions: While the proportion of subjects in each of the IBS subtypes stays the same, individuals commonly transition between subtypes, particularly between IBS-M and IBS-C. We recommend that IBS-A be defined as at least one change between IBS-D and IBS-C by Rome II criteria over a 1-year period.
HNPCC-associated small bowel cancer: Clinical and molecular characteristics
Karsten Schulmann, Frank E. Brasch, Erdmute Kunstmann, Christoph Engel, Constanze Pagenstecher, Holger Vogelsang, Stefan Krüger, Tilman Vogel, Hanns-Peter Knaebel, Josef Rüschoff, Stephan A. Hahn, Magnus V. Knebel-Doeberitz, Gabriela Moeslein, Stephen J. Meltzer, Hans K. Schackert, Christiane Tympner, Elisabeth Mangold, Wolff Schmiegel
Background & aims: The risk for small bowel cancer (SBC) is significantly increased in hereditary nonpolyposis colorectal cancer (HNPCC). HNPCC-associated SBCs are poorly characterized. Methods: Thirty-two SBCs were characterized according to clinical, pathologic, and germline mutation data. Histomorphologic characteristics, microsatellite instability (MSI) testing, mismatch repair (MMR) protein expression, and frameshift mutations of 7 coding mononucleotide repeats were investigated in 17 SBCs. Results: Median age at diagnosis was 39 years. Fifty percent of SBCs were located in the duodenum. The Amsterdam criteria were fulfilled in 50% of patients; 45% of patients had no personal history of previous malignancies. Two patients had a positive family history for SBC. Pathogenic germline mutations were identified in 81%; high MSI was detected in 95% and loss of MMR protein expression in 89% of cases. TGFBR2, BAX, MSH3, MSH6, ACVR2, AIM2, and SEC63 frameshift mutations were detected in 69%, 59%, 59%, 35%, 82%, 56%, and 56%, respectively. An expansive growth pattern of the tumor border and an intense intratumoral lymphocytic infiltrate were present in 75%, respectively. Conclusions: HNPCC-associated SBC often manifests at a young age and may be the first disease manifestation. Endoscopy may detect 50% of tumors. Considering recent data on gastric cancer, we propose endoscopic screening of mutation carriers starting at 30 years of age because clinical criteria cannot define a high-risk group. In addition, our study shows that histopathologic criteria, MSI, and MMR immunohistochemistry are often similar to these features in HNPCC.
Mechanisms of gastroesophageal reflux in critically ill mechanically ventilated patients
Garry Nind, Wei-Hao Chen, Richard Protheroe, Katsuhiko Iwakiri, Robert Fraser, Robert Young, Marianne Chapman, Nam Nguyen, Daniel Sifrim, Rachael Rigda, Richard H. Holloway
Background & aims: Gastroesophageal reflux is a major problem in mechanically ventilated patients and may lead to pulmonary aspiration and erosive esophagitis. Transient lower esophageal sphincter relaxations are the most common mechanism underlying reflux in nonventilated patients. The mechanisms that underlie reflux in critically ill ventilated patients have not been studied. The aim of this study was to determine the mechanisms underlying gastroesophageal reflux in mechanically ventilated patients in the intensive care unit. Methods: In 15 mechanically ventilated intensive care unit patients, esophageal motility, pH, and intraluminal impedance (11/15 patients) were recorded for 1 hour before and 5 hours during continuous nasogastric feeding. Results: Basal lower esophageal sphincter pressure was uniformly low (2.2 ± 0.4 mm Hg). The median (interquartile range) acid exposure (pH <4) was 39.4% (0%–100%) fasting and 32% (7.5%–94.2%) fed. Acid reflux occurred in 10 patients, but slow drifts in esophageal pH were also an important contributor to acid exposure. If esophageal pH decreased to pH <4, it tended to remain so for prolonged periods. A total of 46 acid reflux events were identified. Most (55%) occurred because of absent lower esophageal sphincter pressure alone; 45% occurred during straining or coughing. Conclusions: Gastroesophageal reflux in mechanically ventilated patients is predominantly due to very low or absent lower esophageal sphincter pressure, often with a superimposed cough or strain. These data suggest that measures that increase basal LES pressure may be useful to prevent reflux in ventilated patients.
Preterm birth, low birth weight, and risk for esophageal adenocarcinoma
Magnus Kaijser, Olof Akre, Sven Cnattingius, Anders Ekbom
Background & aims: Gastroesophageal reflux is common among preterm infants and those who are small for gestational age, and it is a strong risk factor for adenocarcinoma of the esophagus. Methods: In a cohort of 3364 individuals born preterm and/or small for gestational age between 1925 and 1949, we assessed the long-term risk for esophageal cancer. Results: The standardized incidence rate ratio for esophageal adenocarcinoma was increased more than 7-fold in the cohort (standardized incidence rate ratio, 7.27; 95% confidence interval, 1.98–18.62), and a birth weight <2000 g was associated with a more than 11-fold increase in risk (standardized incidence rate ratio, 11.5; 95% confidence interval, 1.39–41.5). Conclusions: The associations may be spurious, but if not, they may be explained by increased gastroesophageal reflux during infancy among infants born preterm and/or small for gestational age.
Real-time imaging of human cortical activity evoked by painful esophageal stimulation
Anthony R. Hobson, Paul L. Furlong, Sian F. Worthen, Arjan Hillebrand, Gareth R. Barnes, Krish D. Singh, Qasim Aziz
Background & aims: Current models of visceral pain processing derived from metabolic brain imaging techniques fail to differentiate between exogenous (stimulus-dependent) and endogenous (non-stimulus-specific) neural activity. The aim of this study was to determine the spatiotemporal correlates of exogenous neural activity evoked by painful esophageal stimulation. Methods: In 16 healthy subjects (8 men; mean age, 30.2 ± 2.2 years), we recorded magnetoencephalographic responses to 2 runs of 50 painful esophageal electrical stimuli originating from 8 brain subregions. Subsequently, 11 subjects (6 men; mean age, 31.2 ± 1.8 years) had esophageal cortical evoked potentials recorded on a separate occasion by using similar experimental parameters. Results: Earliest cortical activity (P1) was recorded in parallel in the primary/secondary somatosensory cortex and posterior insula (?85 ms). Significantly later activity was seen in the anterior insula (?103 ms) and cingulate cortex (?106 ms; P = .0001). There was no difference between the P1 latency for magnetoencephalography and cortical evoked potential (P = .16); however, neural activity recorded with cortical evoked potential was longer than with magnetoencephalography (P = .001). No sex differences were seen for psychophysical or neurophysiological measures. Conclusions: This study shows that exogenous cortical neural activity evoked by experimental esophageal pain is processed simultaneously in somatosensory and posterior insula regions. Activity in the anterior insula and cingulate—brain regions that process the affective aspects of esophageal pain—occurs significantly later than in the somatosensory regions, and no sex differences were observed with this experimental paradigm. Cortical evoked potential reflects the summation of cortical activity from these brain regions and has sufficient temporal resolution to separate exogenous and endogenous neural activity.
Clinical-liver, Pancreas, and Biliary Tract
Risk factors of intrahepatic cholangiocarcinoma in the United States: A case-control study
Yasser H. Shaib, Hashem B. El-Serag, Jessica A. Davila, Robert Morgan, Katherine A. McGlynn
Background & aims: The incidence of intrahepatic cholangiocarcinoma has been recently increasing in the United States. In this case-control study, we used the Surveillance, Epidemiology, and End Results-Medicare database to evaluate the prevalence of known risk factors for intrahepatic cholangiocarcinoma and explore other potential risk factors. Methods: We identified all patients with intrahepatic cholangiocarcinoma aged 65 years and older diagnosed between 1993 and 1999 in the population-based Surveillance, Epidemiology, and End Results registries (14% of the US population). Controls were randomly chosen from individuals without any cancer diagnosis in the underlying population of the Surveillance, Epidemiology, and End Results regions. We obtained information on risk factors from Medicare claims (parts A and B) for all cases and controls with at least 2 years of continuous Medicare enrollment. Unadjusted and adjusted odds ratios were calculated in logistic regression analysis. Results: A total of 625 cases and 90,834 controls satisfied the inclusion and exclusion criteria. Cases were older than controls (78.7 vs. 76.5 years; P = .02) and were more likely to be male (48.3% vs. 36.8%; P < .0001). The racial composition was similar between cases and controls. Several risk factors were significantly more prevalent among cases. These included nonspecific cirrhosis (adjusted odds ratio, 27.2; P < .0001), alcoholic liver disease (adjusted odds ratio, 7.4; P < .0001), hepatitis C virus infection (adjusted odds ratio, 6.1; P < .0001), human immunodeficiency virus infection (adjusted odds ratio, 5.9; P = .003), diabetes (adjusted odds ratio, 2.0; P < .0001), and inflammatory bowel diseases (adjusted odds ratio, 2.3; P = .002). Conclusions: This population-based study shows that in addition to previously well described risk factors, several others could be associated with intrahepatic cholangiocarcinoma. These include hepatitis C virus, human immunodeficiency virus, liver cirrhosis, and diabetes.
Contribution of metabolic factors to alanine aminotransferase activity in persons with other causes of liver disease
George N. Ioannou, Noel S. Weiss, Edward J. Boyko, Steven E. Kahn #, Sum P. Lee
Background & Aims: Nonalcoholic fatty liver disease has been defined by the presence of hepatic steatosis in the absence of other chronic liver diseases. We sought to determine whether obesity, insulin resistance, and the metabolic syndrome, which are the main risk factors for nonalcoholic fatty liver disease, are associated with similar elevations in serum alanine aminotransferase activity in persons with and those without other causes of chronic liver disease. Methods: Adult participants of the third National Health and Nutrition Examination Survey were divided into those with causes of chronic liver disease (n = 1037), defined as viral hepatitis, excessive alcohol consumption, or increased transferrin-iron saturation, and those without (n = 8004). Results: Among persons with other causes of chronic liver disease, obesity (adjusted odds ratio, 4.9; 95% confidence interval, 2.5–9.4), insulin resistance (adjusted odds ratio, 6.8; 95% confidence interval, 3.0–15.5, comparing the highest and the lowest quartile), and the metabolic syndrome (adjusted odds ratio, 3.3; 95% confidence interval, 1.4–8.0) were all strongly associated with increased alanine aminotransferase activity (>43 IU/L). Among persons without other causes of chronic liver disease, statistically similar associations were identified. Conclusions: Obesity, insulin resistance, and the metabolic syndrome are strong predictors of increased alanine aminotransferase activity in the US population, both in persons with and in persons without other causes of chronic liver disease. We hypothesize that metabolic fatty liver disease related to these conditions is the cause of the increased alanine aminotransferase activity and may be underrecognized in persons with other causes of chronic liver disease.
Insulin resistance impairs sustained response rate to peginterferon plus ribavirin in chronic hepatitis C patients
Manuel Romero-Gómez, Maria Del Mar Viloria, Raúl J. Andrade, Javier Salmerón, Moisés Diago, Conrado M. Fernández-Rodríguez, Raquel Corpas, Marina Cruz, Lourdes Grande, Luis Vázquez, Paloma Muñoz-de-Rueda, Pilar López-Serrano, Ana Gila, María L. Gutiérrez, Celia Pérez, Angela Ruiz-Extremera, Emilio Suárez, Jesús Castillo
Background & Aims: We evaluated the effect of insulin resistance and viral factors on sustained virological response in patients with chronic hepatitis C treated with peginterferon plus ribavirin. Methods: Patients (n = 159; 94 men; age, 41.7 ± 11.1 years) with chronic hepatitis C (genotype 1, n = 113; non-1 genotype, n = 46) received treatment with interferon plus ribavirin. Serum levels of leptin and insulin were measured, and the insulin resistance index (HOMA-IR: homeostasis model of assessment) and body mass index were calculated. Results: A sustained virological response was associated with lower age, insulin resistance index, body mass index, and ?-glutamyltranspeptidase and serum leptin levels. There was no association with viral load, sex, type of interferon, or cholesterol levels. A sustained virological response was achieved in 43.4% (46/113) of genotype 1 and 89% (32/36) of genotype 2 and 3 (P = .0001) patients. Necroinflammatory activity and steatosis were not associated with the sustained virological response rate. Multivariate regression analysis indicated that the independent variables related to sustained virological response were genotype (odds ratio, 3.57; 95% confidence interval, 1.49–8.3; P = .001), insulin resistance index (odds ratio, 1.82; 95% confidence interval, 1.08–3.06; P = .012), and fibrosis (odds ratio, 1.36; 95% confidence interval, 1.01–1.84; P = .029). A sustained virological response in patients with genotype 1 and insulin resistance (HOMA-IR > 2) occurred in 23 of 70 (32.8%; 95% confidence interval, 21.9%–43.9%) patients, vs. 26 of 43 (60.5%; 95% confidence interval, 45.9%–75.1%) genotype 1 patients without insulin resistance (P = .007; odds ratio, 3.12, 95% confidence interval, 1.42–6.89). Conclusions: Insulin resistance, fibrosis, and genotype are independent predictors of the response to antiviral therapy in chronic hepatitis C patients treated with peginterferon plus ribavirin.
Basic-alimentary Tract
REG I protein may function as a trophic and/or anti-apoptotic factor in the development of gastric cancer
Akira Sekikawa, Hirokazu Fukui, Shigehiko Fujii, Jun Takeda, Apichart Nanakin, Hiroshi Hisatsune, Hiroshi Seno, Shin Takasawa, Hiroshi Okamoto, Takahiro Fujimori, Tsutomu Chiba
Background & Aims: Although a significant amount of regenerating gene (REG) I? protein is present not only in normal gastric mucosa but also in gastric cancer tissues, its pathophysiologic role in gastric cancer development remains unclear. We investigated REG I? protein expression in early gastric cancers, and examined whether cytokines are responsible for REG I? gene expression and whether REG I? protein has a trophic and/or an antiapoptotic effect on gastric cancer cells. Methods: Early gastric cancer specimens were analyzed histologically using immunohistochemistry for REG I? protein and proliferating cell nuclear antigen (PCNA). The effects of cytokines on REG I? promoter activity and its messenger RNA (mRNA) expression in AGS (a kind of gastric cancer cell line) cells were examined by luciferase reporter assay and Northern blot analysis, respectively. Effects of REG I? protein on cell growth and H2O2-induced apoptosis in AGS cells were examined by 3,-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphatase nick-end labeling (TUNEL) assays, respectively. Results: REG I?-positive early gastric cancers showed a significantly higher PCNA labeling index and more severe inflammatory cell infiltration in adjacent gastric mucosa than the negative cancers. REG I? gene expression and its promoter activity were enhanced by interferon (IFN)-? and interleukin (IL)-6. REG I? protein promoted cell growth and cell resistance to H2O2-induced apoptosis in AGS cells. These effects were abolished by concomitant treatment with anti-REG I? antibody. REG I? protein enhanced Akt phosphorylation and Bcl-xL expression in AGS cells. Conclusions: REG I? gene is inducible by cytokine stimulation and its gene product may function as a mitogenic and/or an antiapoptotic factor in the development of early gastric cancer.
Proinflammatory effects of TH2 cytokines in a murine model of chronic small intestinal inflammation
Giorgos Bamias, Charles Martin, Margarita Mishina, William G. Ross, Jesus Rivera-Nieves, Marco Marini, Fabio Cominelli
Background & Aims: Strict TH1 polarization is believed to underlie the pathogenesis of intestinal inflammation in Crohn’s disease. In the present study we tested the hypothesis that TH2 cytokines also may participate in disease development in SAMP1/YitFc mice that spontaneously develop terminal ileitis with perianal manifestations. Methods: Cytokine messenger RNA (mRNA) expression was studied by real-time polymerase chain reaction (PCR). Lamina propria mononuclear cells (LPMCs) were purified and stimulated cytokine secretion was analyzed. Blockade of interferon (IFN)-? or interleukin (IL)-4 was performed by using specific neutralizing monoclonal antibodies (MAbs). CD4+/IL-4-secreting lymphocytes were purified from SAMP1/YitFc mesenteric lymph nodes (MLNs) and their ability to induce ileitis was tested after transfer to SCID recipients. Results: Initiation of ileitis in SAMP1/YitFc mice was TH1-mediated because up-regulation of IFN-? and tumor necrosis factor (TNF) preceded the histologic injury, whereas IFN-? neutralization prevented the development of chronic inflammation (P < .005) by interfering with the expansion of lymphocytes. In contrast, the establishment of chronic ileitis coincided with significant increases in IL-5 (35?) and IL-13 (29?) mRNA expression (P < .005), as well as in TH2 cytokine secretion by lamina propria lymphocytes (P < .05 vs. AKR controls). IL-4 blockade diminished IFN-? mRNA expression and significantly ameliorated the severity of established ileitis (P < .05) by decreasing the histologic indices for villous distortion and active inflammation. In addition, IL-4 augmented the in vitro IFN-? secretion by lymphocytes, whereas IL-4-secreting CD4+ lymphocytes were sufficient for adoptively transferring ileitis to SCID recipients. Conclusions: Our results indicate that both TH1 and TH2 pathways mediate Crohn’s-like ileitis and suggest that combined TH1/TH2 manipulation may offer a therapeutic advantage for the treatment of Crohn’s disease.
Role of progesterone signaling in the regulation of G-protein levels in female chronic constipation
Zuo-Liang Xiao, Victor Pricolo, Piero Biancani, Jose Behar
Background & Aims: Chronic constipation caused by slow transit is common in women with an F/M ratio of 9:1. The cause and mechanisms responsible for this syndrome are unknown. Progesterone has been suggested as a possible contributing factor. Our aim was to investigate the site and mechanisms responsible for this colonic motility disorder. Methods: Seven women with intractable constipation and slow transit time underwent colectomy and 6 women who underwent a left colectomy for adenocarcinoma (controls) were studied. Dissociated colonic circular muscle cells were obtained by enzymatic digestion. Changes in G-protein levels were measured by Western blot. The messenger RNA (mRNA) expression of G?q and progesterone receptors was determined by reverse-transcription polymerase chain reaction and Northern blot. Results: Muscle cells from patients with chronic constipation exhibited impaired contraction in response to receptor-G-protein-dependent agonists (cholecystokinin [CCK], acetylcholine) and in response to the direct G-protein activator guanosine 5?-O-(3-thiophosphate). Contraction was normal with receptor-G-protein-independent agonists (diacylglycerol and KCl). Western blot showed down-regulation of G?q/11 and up-regulation of G?s proteins in patients with chronic constipation. The mRNA expression of G?q was lower and the progesterone receptors were overexpressed in patients with chronic constipation compared with controls. These abnormalities were reproduced in vitro by pretreatment of normal colonic muscle cells with progesterone for 4 hours. Conclusions: Slow transit chronic constipation in women may be caused by down-regulation of contractile G proteins and up-regulation of inhibitory G proteins, probably caused by overexpression of progesterone receptors.
The human colonic monocarboxylate transporter Isoform 1: Its potential importance to colonic tissue homeostasis
Mark Cuff, Jane Dyer, Mark Jones, Soraya Shirazi-Beechey
Background & Aims: Butyrate serves as the major source of energy for colonic epithelial cells, and has profound effects on their proliferation, differentiation, and apoptosis. Transport of butyrate across the colonocyte luminal membrane is mediated by the monocarboxylate transporter, MCT1; the expression of which is down-regulated dramatically during colon carcinogenesis. We have proposed that the decline in MCT1 expression during colon carcinogenesis may reduce the intracellular availability of butyrate required to regulate expression of genes associated with the processes maintaining tissue homeostasis within the colonic mucosa. Methods: To test this hypothesis we used the technique of RNA interference to inhibit MCT1 expression specifically, and determined the consequences of this inhibition on the ability of butyrate to exert its recognized effects in vitro using flow cytometry, immunofluorescence, Northern analysis, and Western analysis. Results: We show that inhibition of MCT1 expression, and hence butyrate uptake, has profound inhibitory effects on the ability of butyrate to regulate expression of key target genes: p21waf1/cip1 (p21), intestinal alkaline phosphatase (IAP), and cyclin D1, and their associated processes of proliferation and differentiation. In contrast, inhibition of MCT1 expression had no effect on the ability of butyrate to modulate expression of either bcl-xL or bak, and this was reflected in a corresponding lack of effect on butyrate induction of apoptosis. Conclusions: Collectively, these results show the importance of MCT1 to the ability of butyrate to induce cell-cycle arrest and differentiation, and suggest fundamental differences in the mechanisms by which butyrate modulates specific aspects of cell function.
Interleukin-21 enhances T-helper cell type I signaling and interferon-? production in Crohn’s disease
Giovanni Monteleone, Ivan Monteleone, Daniele Fina, Piero Vavassori, Giovanna Del Vecchio Blanco, Roberta Caruso, Roberto Tersigni, Luciano Alessandroni, Livia Biancone, Gian Carlo Naccari, Thomas T. MacDonald, Francesco Pallone
Background & Aims T-helper (Th)1 cells play a central role in the pathogenesis of tissue damage in Crohn’s disease (CD). Interleukin (IL)-12/STAT4 signaling promotes Th1 cell commitment in CD, but other cytokines are needed to maintain activated Th1 cells in the mucosa. In this study, we examined the expression and role of IL-21, a T-cell–derived cytokine of the IL-2 family; in tissues and cells isolated from patients with inflammatory bowel disease. Methods IL-21 was examined by Western blotting in whole mucosa and lamina propria mononuclear cells (LPMCs) from patients with CD, ulcerative colitis (UC), and controls. We also examined the effects of exogenous IL-12 on IL-21 production, as well as the effects of blocking IL-21 with an IL-21–receptor Ig fusion protein. Interferon (IFN)-? was measured in the culture supernatants by enzyme-linked immunosorbent assay, and phosphorylated STAT4 and T-bet were examined by Western blotting. Results IL-21 was detected in all samples, but its expression was higher at the site of disease in CD in comparison with UC and controls. Enhanced IL-21 was seen in both ileal and colonic CD and in fibrostenosing and nonfibrostenosing disease. IL-12 enhanced IL-21 in normal lamina propria lymphocytes through an IFN-?–independent mechanism, and blocking IL-12 in CD LPMCs decreased anti-CD3–stimulated IL-21 expression. Neutralization of IL-21 in CD LPMC cultures decreased phosphorylated STAT4 and T-bet expression, thereby inhibiting IFN-? production. Conclusions Our data suggest that IL-21 contributes to the ongoing Th1 mucosal response in CD.
Deoxycholic acid activates protein kinase C and phospholipase C via increased Ca2+ entry at plasma membrane
Bonnie W. Lau, Matilde Colella, Warren C. Ruder, Marianna Ranieri, Silvana Curci, Aldebaran M. Hofer
Background & Aims Secondary bile acids like deoxycholic acid (DCA) are well-established tumor promoters that may exert their pathologic actions by interfering with intracellular signaling cascades. Methods We evaluated the effects of DCA on Ca2+ signaling in BHK-21 fibroblasts using fura-2 and mag-fura-2 to measure cytoplasmic and intraluminal internal stores [Ca2+], respectively. Furthermore, green fluorescent protein (GFP)-based probes were used to monitor time courses of phospholipase C (PLC) activation (pleckstrin-homology [PH]-PLC?-GFP), and translocation of protein kinase C (PKC) and a major PKC substrate, myristolated alanine-rich C-kinase substrate (MARCKS). Results DCA (50–250 ?mol/L) caused profound Ca2+ release from intracellular stores of intact or permeabilized cells. Correspondingly, DCA increased cytoplasmic Ca2+ to levels that were ?120% of those stimulated by Ca2+-mobilizing agonists in the presence of external Ca2+, and ?60% of control in Ca2+-free solutions. DCA also caused dramatic translocation of PH-PLC?-GFP, and conventional, Ca2+/diacylglycerol (DAG)-dependent isoforms of PKC (PKC-?I and PKC-?), and MARCKS-GFP, but only in Ca2+-containing solutions. DCA had no effect on localization of a novel (PKC?) or an atypical (PKC?) PKC isoform. Conclusions Data are consistent with a model in which DCA directly induces both Ca2+ release from internal stores and persistent Ca2+ entry at the plasma membrane. The resulting microdomains of high Ca2+ levels beneath the plasma membrane appear to directly activate PLC, resulting in modest InsP3 and DAG production. Furthermore, the increased Ca2+ entry stimulates vigorous recruitment of conventional PKC isoforms to the plasma membrane.
Basic-liver, Pancreas, and Biliary Tract
RNA interference-mediated control of hepatitis B virus and emergence of resistant mutant
Hui-Lin Wu, Li-Rung Huang, Chuan-Chuan Huang, Hsiao-Lei Lai, Chun-Jen Liu, Yu-Tzu Huang, Yun-Wei Hsu, Cheng-Yi Lu, Ding-Shinn Chen, Pei-Jer Chen
Background & Aims Present therapy for chronic hepatitis B attains control only in limited proportions. Small interfering RNA (siRNA) offers a new tool with potential therapeutic applications for hepatitis B virus (HBV). Given the importance of sequence identity in the effectiveness of siRNA and the heterogeneity of HBV sequences among different isolates, a short hairpin RNA (shRNA)-expressing plasmid, pSuper/HBVS1, was developed to target a region conserved among major HBV genotypes and assess its effectiveness control of HBV. Methods HBV replication-competent plasmid was cotransfected with pSuper/HBVS1 to HuH-7 cells or to mice. The levels of viral proteins, RNA, and DNA were examined in transfected cells and animals. The effects of pSuper/HBVS1 on clinical isolates with genotypes B and C were also determined. Results pSuper/HBVS1 significantly decreased levels of viral proteins, RNA, and DNA for HBV genotype A in cell culture and in mice. Comparable suppressive effects were observed on clinical isolates of genotypes B and C. A clone with a silent mutation in the target region was identified from a patient with genotype C. This mutant revealed diminished sensitivity to pSuper/HBVS1 and could be selected out in the presence of pSuper/HBVS1 in cell culture. Conclusions These findings indicated that shRNA could suppress HBV expression and replication for genotypes A, B, and C, promising an advance in treatment of HBV. However, the emergence of resistant mutants in HBV quasispecies should be considered.
Transgenic expression of pancreatic secretory trypsin inhibitor-I ameliorates secretagogue-induced pancreatitis in mice
Jaimie D. Nathan, Joelle Romac, Ruth Y. Peng, Michael Peyton, Raymond J. MacDonald, Rodger A. Liddle
Background & aims Endogenous trypsin inhibitors are believed to inhibit protease activity if trypsin becomes inadvertently activated within the acinar cell. However, this action remains unproven, and the role of endogenous pancreatic trypsin inhibitors in acute pancreatitis is unknown. In this study, we tested whether increased levels of pancreatic secretory trypsin inhibitor-I (PSTI-I) in mice could prevent secretagogue-induced pancreatitis. Methods Rat PSTI-I expression was targeted to pancreatic acinar cells in transgenic mice by creating a minigene driven by the rat elastase I enhancer/promoter. Secretagogue-induced pancreatitis was achieved by 12 hourly intraperitoneal injections of caerulein. The severity of pancreatitis was assessed by measurements of serum amylase, histologic grading, and pancreas wet weight-to-body weight ratio. Trypsinogen activation and trypsin activity were measured in pancreatic extracts. Results Targeted expression of PSTI-I to the pancreas increased endogenous trypsin inhibitor capacity by 190% (P < .01) in transgenic vs. nontransgenic mice. Caerulein administration to nontransgenic mice produced histologic evidence of acute pancreatitis, and significantly elevated serum amylase and pancreas weight ratio. In caerulein-treated transgenic mice, the histologic severity of pancreatitis was significantly reduced. There was no difference in trypsinogen activation peptide levels between caerulein-treated transgenic and nontransgenic mice. However, trypsin activity was significantly lower in transgenic mice receiving caerulein compared with nontransgenic mice. Conclusions These data demonstrate that the severity of secretagogue-induced pancreatitis is significantly ameliorated in mice with higher pancreatic levels of trypsin inhibitor. We propose that PSTI-I prevents pancreatitis by inhibiting the activity of trypsin, rather than by reducing trypsinogen activation.
Recapitulation of elements of embryonic development in adult mouse pancreatic regeneration
Jan Nygaard Jensen, Erin Cameron, Maria Veronica R. Garay, Thomas W. Starkey, Roberto Gianani, Jan Jensen
Background & Aims: The mammalian pancreas has a strong regenerative potential, but the origin of organ restoration is not clear, and it is not known to what degree such a process reflects pancreatic development. To define cell differentiation changes associated with pancreatic regeneration in adult mice, we compared regeneration following caerulein-induced pancreatitis to that of normal pancreatic development. Methods: By performing comparative histology for adult and embryonic pancreatic markers in caerulein-treated and control pancreas, we addressed cellular proliferation and differentiation (amylase, DBA-agglutinin, insulin, glucagon, ?-catenin, E-cadherin, Pdx1, Nkx6.1, Notch1, Notch2, Jagged1, Jagged2, Hes1), hereby describing the kinetics of tissue restoration. Results: We demonstrate that surviving pancreatic exocrine cells repress the terminal exocrine gene program and induce genes normally associated with undifferentiated pancreatic progenitor cells such as Pdx1, E-cadherin, ?-catenin, and Notch components, including Notch1, Notch2, and Jagged2. Expression of the Notch target gene Hes1 provides evidence that Notch signaling is reactivated in dedifferentiated pancreatic cells. Although previous studies have suggested a process of acino-to-ductal transdifferentiation in pancreatic regeneration, we find no evidence to suggest that dedifferentiated cells acquire a ductal fate during this process. Conclusions: Pancreatic regeneration following chemically induced pancreatitis in the mouse occurs predominantly through acinar cell dedifferentiation, whereby a genetic program resembling embryonic pancreatic precursors is reinstated.
Antifibrogenic role of the cannabinoid receptor CB2 in the liver
Boris Julien, Pascale Grenard, Fatima Teixeira-Clerc, Jeanne Tran van Nhieu, Liying Li, Meliha Karsak, Andreas Zimmer, Ariane Mallat, Sophie Lotersztajn
Background & Aims: Hepatic myofibroblasts are central for the development of liver fibrosis associated with chronic liver diseases, and blocking their accumulation may prevent fibrogenesis. Cannabinoids are the active components of marijuana and act via 2 G-protein-coupled receptors, CB1 and CB2. Here, we investigated whether liver fibrogenic cells are a target of cannabinoids. Methods: CB2 receptors were characterized in biopsy specimens of normal human liver and active cirrhosis by immunohistochemistry, and in cultures of hepatic stellate cells and hepatic myofibroblasts by reverse-transcription polymerase chain reaction (RT-PCR), immunocytochemistry, and GTP?S assays. Functional studies were performed in cultured hepatic myofibroblasts and activated hepatic stellate cells. Carbon tetrachloride-induced liver fibrosis was studied in mice invalidated for CB2 receptors. Results: In liver biopsy specimens from patients with active cirrhosis of various etiologies, CB2 receptors were expressed in nonparenchymal cells located within and at the edge of fibrous septa in smooth muscle ?-actin-positive cells. In contrast, CB2 receptors were not detected in normal human liver. CB2 receptors were also detected in cultured hepatic myofibroblasts and in activated hepatic stellate cells. Their activation triggered potent antifibrogenic effects, namely, growth inhibition and apoptosis. Growth inhibition involved cyclooxygenase-2, and apoptosis resulted from oxidative stress. Finally, mice invalidated for CB2 receptors developed enhanced liver fibrosis following chronic carbon tetrachloride treatment as compared with wild-type mice. Conclusions: These data constitute the first demonstration that CB2 receptors are highly up-regulated in the cirrhotic liver, predominantly in hepatic fibrogenic cells. Moreover, this study also highlights the antifibrogenic role of CB2 receptors during chronic liver injury.
Case Report
Clostridium difficile toxoid vaccine in recurrent C. difficile-associated diarrhea
Stavros Sougioultzis, Lorraine Kyne, Denise Drudy, Sarah Keates, Seema Maroo, Charalabos Pothoulakis, Paul J. Giannasca, Cynthia K. Lee, Michel Warny, Thomas P. Monath, Ciarán P. Kelly
Background & aims: Recurrent C difficile-associated diarrhea (CDAD) is associated with a lack of protective immunity to C difficile toxins. A parenteral C difficile vaccine containing toxoid A and toxoid B was reported previously to be safe and immunogenic in healthy volunteers. Our aim was to examine whether the vaccine is also well tolerated and immunogenic in patients with recurrent CDAD. Methods: Subjects received 4, 50-?g intramuscular inoculations of the C difficile vaccine over an 8-week period. Serum antitoxin antibodies were measured by ELISA, and toxin neutralizing activity was evaluated using the tissue culture cytotoxin assay. Results: Three patients with multiple episodes of recurrent CDAD were vaccinated. Two of the 3 showed an increase in serum IgG antitoxin A antibodies (3-fold and 4-fold increases, respectively) and in serum IgG antitoxin B antibodies (52-fold and 20-fold, respectively). Both also developed cytotoxin neutralizing activity against toxin A and toxin B. Prior to vaccination, the subjects had required nearly continuous treatment with oral vancomycin for 7, 9, and 22 months, respectively, to treat recurrent episodes of CDAD. After vaccination, all 3 subjects discontinued treatment with oral vancomycin without any further recurrence. Conclusions: A C difficile toxoid vaccine induced immune responses to toxins A and B in patients with CDAD and was associated with resolution of recurrent diarrhea. The results of this study support the feasibility of active vaccination against C difficile and its toxins in high-risk individuals but must be validated in larger, randomized, controlled trials.
Copyright © 2001-2005 by the American Gastroenterological Association. All rights reserved.
Table of Contents for March 2005 (Vol. 42, Issue 3)
Viral Hepatitis
Hepatitis C coinfection increases the risk of fulminant hepatic failure in patients with HIV in the HAART era, 25 November 2004
Kramer JR, Giordano TP, Souchek J, El-Serag HB
Background/Aims: It is uncertain if patients coinfected with hepatitis C and HIV are more likely to suffer fulminant hepatic failure (FHF) when compared to patients with HIV-only.
Methods: We conducted a retrospective cohort study using national administrative databases from the Department of Veterans Affairs in patients hospitalized for the first time with HIV and/or hepatitis C between 10/1991 and 9/2000. Fulminant hepatic failure was defined as occurring after the index hospitalization through 9/2001 in the absence of pre-existing liver disease. We calculated incidence rates, Kaplan Meier cumulative incidence curves, and Cox proportional hazards ratios while adjusting for demographics and other potential confounders.
Results: We identified 11,678 patients with HIV-only and 4761 patients with coinfection. There were 92 cases of fulminant hepatic failure yielding an incidence rate of 1.1/1000 person-years and 2.5/1000 person-years in the HIV-only and coinfected groups. The cumulative incidence of fulminant hepatic failure in the coinfected group was higher than in the HIV-only group (P<0.0001). The risk of fulminant hepatic failure in patients with coinfection compared to HIV-only during the HAART era was several folds higher than that during the pre-HAART era.
Conclusions: HAART and hepatitis C coinfection appeared to act synergistically in HIV-infected patients to increase the risk of fulminant hepatic failure, a rare but often fatal disease.
Etanercept as an adjuvant to interferon and ribavirin in treatment-naive patients with chronic hepatitis C virus infection: a phase 2 randomized, double-blind, placebo-controlled study, 29 November 2004
Zein NN, for the Etanercept Study Group
Background/Aims: Current therapies for patients with chronic hepatitis C virus (HCV) do not achieve sustained viral clearance in most patients, and are associated with severe toxic effects. Our aim was to investigate the efficacy and safety of etanercept as adjuvant to interferon and ribavirin in treatment-naive patients with HCV.
Methods: Double-blind, randomized, placebo controlled trial. Fifty patients with chronic HCV were randomly assigned to receive interferon alfa-2b and ribavirin with either etanercept or placebo for 24 weeks. The main outcome measure was the absence of HCV RNA at 24 weeks, the on treatment response at the end of the etanercept randomization period.
Results: At 24 weeks, HCV RNA was absent in 63% (12/19) etanercept patients compared to 32% (8/25) placebo patients (P=0.04). In addition, patients receiving etanercept had lower frequency of most adverse events categories compared to placebo.
Conclusions: Etanercept given for 24 weeks as adjuvant therapy to interferon and ribavirin significantly improved virologic response at the end of the etanercept randomization period among patients with HCV, and was associated with decreased incidence of most adverse effects associated with interferon and ribavirin.
Barriers to interferon-? therapy are higher in intravenous drug users than in other patients with acute hepatitis C, 24 November 2004
Broers B, Helbling B, François A, Schmid P, Chuard C, Hadengue A, Negro F, for the Swiss Association for the Study of the Liver (SASL 18)
Background/Aims: Treatment with interferon-? (IFN-?) may eradicate HCV in most acute hepatitis C patients, thus preventing chronic hepatitis and avoiding less efficacious combination therapy.
Methods: In a prospective study, we evaluated the impact of barriers to successful start and completion of treatment of acute and subacute (<12 months from infection) hepatitis C with pegylated IFN-?2b, 1.5?g/kg, QW, for 24 weeks.
Results :Out of 27 patients (22 were active intravenous drug users [IVDU]), 5 cleared HCV spontaneously. Antiviral treatment was indicated in 22 patients: six refused therapy for fear of side effects, whereas two others were lost to observation. Eight patients completed the treatment or received >80% of the scheduled drug: seven (88%) were sustained virological responders 24 weeks after the end of treatment. Six patients (all IVDU) stopped prematurely due to side effects: only one had a sustained virological response. Based on an intent-to-treat analysis, and considering all 14 patients in whom at least one dose of drug was administered, only 8 (57%) became sustained virological responders. Conclusions: Treatment of acute hepatitis C with pegylated IFN-? is highly beneficial, but its effectiveness is affected by a poor rate of acceptance and/or adherence to currently available regimens, especially in IVDU and women.
Efficacy of a 24-week course of PEG-interferon ?-2b monotherapy in patients with acute hepatitis C after failure of spontaneous clearance, 26 November 2004
Santantonio T, Fasano M, Sinisi E, Guastadisegni A, Casalino C, Mazzola M, Francavilla R, Pastore G
Background/Aims: Interferon (IFN) monotherapy significantly reduces the chronicity rate of acute hepatitis C (AHC) but optimal regimen and treatment timing remain undefined. The aim of this study was to assess the efficacy of a 6-month course of pegylated IFN (PEG-IFN) ?-2b monotherapy in AHC patients and to investigate if IFN treatment initiated after 12 weeks from clinical presentation, still achieved a high response rate.
Methods: Sixteen AHC patients still viremic after 12 weeks from the onset were treated with PEG-IFN ?-2b (1.5mcg/kg once weekly) for 6 months and followed for at least 12 months. Response to therapy was defined as normal ALT values and undetectable HCV RNA (<50IU/ml) at the end of therapy, after 6 (sustained response) and 12 months follow-up (long-term response).Results: At the end of treatment, HCV RNA was undetectable in 15/16 patients while ALT normalized in 14/16 patients. After 6 and 12 months follow-up, 15/16 patients (94%) showed virological and biochemical response.
Conclusions: A 6-month course of PEG-IFN ?-2b is effective in inducing resolution of AHC in 94% of patients. Our results provide a rationale for delaying treatment for 12 weeks, targeting only patients who fail to clear the virus spontaneously and truly requiring therapy without loss of efficacy.
Vascular cell adhesion molecule-1 (VCAM-1) plays a central role in the pathogenesis of severe forms of vasculitis due to hepatitis C-associated mixed cryoglobulinemia, 24 January 2005
Kaplanski G, Maisonobe T, Marin V, Grès S, Robitail S, Farnarier C, Harlé JR, Piette JC, Cacoub P
Background/Aims: To better characterize the molecules involved in leukocyte tissue infiltration during hepatitis C-mixed cryoglobulinemia (HCV-MC)-associated vasculitis.
Methods: The involvement of ELAM, ICAM-1 and VCAM-1 in 36 patients with HCV-MC vasculitis using three different approaches: concentrations of soluble forms by specific ELISA, tissue expression by immunohistochemistry on patients nerve biopsies, endothelial expression by FACS analysis, on cells activated in vitro by cryoprecipitates purified from HCV-MC patients. Results: Concentrations of sVCAM-1 were significantly elevated in the serum of HCV-MC patients compared to HCV patients without MC, the highest concentrations being found in severe vasculitis. VCAM-1 expression was detected on blood vessels from nerve biopsies performed in patients with severe vasculitis. When added to endothelial cells in vitro, HCV-MC patients cryoprecipitate induced VCAM-1 but also ELAM and ICAM-1 expression possibly through a mechanism due to the C1q complement fraction interaction with endothelial cells, since C1q was consistently present in the cryoprecipitates.
Conclusions: VCAM-1 is mainly involved in the pathogenesis of HCV-MC-associated severe vasculitis and may be a potential interesting therapeutic target.
Transplantation
Hepatitis C virus viral recurrence and liver mitochondrial damage after liver transplantation in HIV–HCV co-infected patients, 6 December 2004
Duclos-Vallée JC, Vittecoq D, Teicher E, Feray C, Roque-Afonso AM, Lombès A, Jardel C, Gigou M, Dussaix E, Sebagh M, Guettier C, Azoulay D, Adam R, Ichaï P, Saliba F, Roche B, Castaing D, Bismuth H, Samuel D
Background/Aims: As life expectancy in HIV–HCV co-infected patients improves, end stage liver disease requiring liver transplantation (LT) may become an emerging problem. We report the Paul Brousse Hospital experience of transplantation for end stage cirrhosis in HIV–HCV co-infected patients.
Methods: Seven consecutive HIV–HCV co-infected patients were transplanted between December 1999 and December 2002 for end stage liver disease due to HCV. All patients were treated by highly active antiretroviral therapy (HAART), HIV plasma viral load was <400copies/ml and median CD4 lymphocyte count was 306cells/mm3 (range, 103–510) before LT. At the time of evaluation (March 2003), the median follow-up was 21 months (range, 4–40).
Results: Two patients died, 4 and 22 months, respectively after LT. At the last biopsy, METAVIR score was staged F4 in two patients, F3 in two, and F1 in one. Microvesicular steatosis was noted in nearly all patients. The ratio of mitochondrial to nuclear DNA was low in three of four patients examined as compared with the amount of liver mtDNA found in eight HIV-negative, HCV-infected controls (P=0.01).
Conclusions: A significant defect in the activity of the respiratory chain complex IV was noted in all five patients studied. Mitochondrial hepatotoxicity and severe HCV recurrence occur in HIV–HCV co-infected patients after LT.
Liver Failure, Growth and Cancer
Amelioration of microcirculatory damage by an endothelin A receptor antagonist in a rat model of reversible acute liver failure, 29 November 2004
Palmes D, Skawran S, Stratmann U, Armann B, Minin E, Herbst H, Spiegel HU
Background/Aims: Hepatocellular damage in acute liver failure (ALF) is aggravated by proinflammatory and cytotoxic mediators released from sinusoidal-lining cells. We studied a selective endothelin A receptor (ETAR) antagonist for its potential influence on the microcirculation in the setting of ALF.
Methods: Seventy Wistar rats were divided into five groups: (I) induction of ALF by a 70% liver resection combined with injection of 400?g/kg endotoxin, (II) ALF treated with the ETAR antagonist LU 135252 (1mg/kg b.w. i.v.), (III) sham operation, (IV) injection of endotoxin, (V) 70% liver resection. Liver microcirculation was measured by intravital microscopy. Parenchymal injury, growth fractions, endothelin (ET)-1 and ETAR were studied by histology and immunohistology. Survival, liver function, and morphology were followed up to 14 days.
Results: 100% mortality, impaired liver function, widespread endothelial lesions, highest ET-1 and ETAR levels, a decreased perfusion rate, reduced sinusoidal diameter, as well as an increase in both leukocyte–endothelium interactions and sinusoidal blood flow were observed after induction of ALF. ETAR antagonist-treated rats showed decreased ET-1 and ETAR levels as well as improved microcirculatory function, morphology, liver function, and 85% survival.
Conclusions: Microcirculatory disturbances correlate with liver dysfunction in ALF. ETAR blockade represents a new therapeutic approach to ALF by reducing microcirculatory lesions and their sequelae.
Hypoxia stimulates proliferation of human hepatoma cells through the induction of hexokinase II expression, 1 December 2004
Gwak GY, Yoon JH, Kim KM, Lee HS, Chung JW, Gores GJ
Background/Aims: In a hypoxic state, a glycolytic system is operating as a salvage pathway of generating ATP, and hexokinase II, the first enzyme in this system, might be over-expressed in hepatocellular carcinomas (HCCs). This study was to evaluate if hexokinase II is participating in HCC cell survival in a hypoxic state, and to analyze the mechanism of cell death caused by hexokinase II-specific inhibition.
Methods: Human hepatoma cell lines were grown either in a normoxic or hypoxic condition. Hexokinase II and hypoxia-inducible factor-1? (HIF-1?) expression were evaluated using immunoblot techniques. Cell growth was assessed using the MTS assay. Apoptotic signaling cascades were explored by immunoblot analysis.
Results: Hypoxia stimulated HCC cellular growth through HIF-1?-dependent induction of hexokinase II expression. The hexokinase II-specific inhibitor, 3-bromopyruvate, significantly suppressed cellular growth in a hypoxic state compared to cells in a normoxic condition. This suppression was due to the induction of apoptosis through activating mitochondrial apoptotic signaling cascades.
Conclusions: This study demonstrates that hypoxia stimulates HCC cellular growth through hexokinase II induction, and its inhibition induces apoptotic cell death. Therefore, hexokinase II induction may participate in HCC progression and the blockage of this enzyme may therapeutically be efficacious in human HCCs.
Pharmacokinetics and efficacy of oral versus intravenous mixed-micellar phylloquinone (vitamin K1) in severe acute liver disease, 15 December 2004
Pereira SP, Rowbotham D, Fitt S, Shearer MJ, Wendon J, Williams R
Background/Aims: In patients with severe acute liver dysfunction, i.v. phylloquinone (vitamin K1) may be given to exclude vitamin K deficiency, rather than impaired hepatic synthesis of coagulation factors alone, as the cause of the coagulopathy. However, there have been no studies of the pharmacokinetics or efficacy of i.v. or oral K1 in such patients.
Methods: 49 adults with severe acute liver disease were randomised double-blind to a single 10mg dose of i.v. or oral mixed-micellar K1, or placebo. Serum levels of phylloquinone and undercarboxylated prothrombin (PIVKA-II) were assessed before and after treatment.
Results: At admission, 13 patients (27%) had either low serum K1 levels or elevated PIVKA-II concentrations, indicative of subclinical vitamin K deficiency. In the 16 patients who received i.v. K1, there was one (6%) treatment failure (K1 rise <10ng/ml above baseline), compared with 12 of the 15 (80%) who received oral K1 (P<0.0001). One patient in the placebo group developed overt vitamin K deficiency.
Conclusions: A minority of patients with severe acute liver dysfunction have subclinical vitamin K deficiency at the time of presentation, which is corrected by a single dose of i.v. K1. The intestinal absorption of mixed-micellar K1 is unreliable in adults with severe acute liver dysfunction.
Ethanol binging exacerbates sinusoidal endothelial and parenchymal injury elicited by acetaminophen, 22 December 2004
McCuskey RS, Bethea NW, Wong J, McCuskey MK, Abril ER, Wang X, Ito Y, DeLeve LD
Background/Aims: The pathophysiology of binge drinking of ethanol and its potentiation of acetaminophen (APAP) toxicity has received very little attention. To evaluate if ethanol binging sensitizes hepatic sinusoidal endothelial cells (SEC) and liver to APAP toxicity.
Methods: The histopathological responses to APAP were evaluated in the livers of mice gavaged with APAP alone, following a single, week-end type ethanol binge (4g/kg every 12h?5 doses) or three weekly binges.
Results: Six hours after APAP, 600mg/kg elicited severe centrilobular necrosis together with hemorrhagic congestion and infiltration of erythrocytes into the Space of Disse through large gaps that had formed in SEC. There was no evidence of parenchymal injury at 2h, but gaps already were formed through the cytoplasm of the SEC by coalescence of fenestrae. A single binge followed by 300mg/kg APAP elicited SEC and parenchymal injury equivalent to 600mg/kg APAP alone at 2 and 6h. The responses were exacerbated following three binges. Lower glutathione levels in the liver were shown in ethanol-binged animals.
Conclusions: Ethanol binging increases APAP hepatotoxicity. SEC are an early target for APAP-induced injury and ethanol binging enhances the SEC injury prior to evidence of parenchymal cell injury.
Cholestasis and Autoimmune Liver Disease
Oncosis represents the main type of cell death in mouse models of cholestasis, 8 November 2004
Fickert P, Trauner M, Fuchsbichler A, Zollner G, Wagner M, Marschall HU, Zatloukal K, Denk H
Background/Aims: Since the mechanisms leading to hepatocyte death in cholestasis are not well defined, we aimed to obtain closer insights into the related pathogenetic principles.
Methods: Cell death was assessed in common bile duct ligated (CBDL) and cholic acid (CA)-fed mice, and compared to Fas agonist Jo2-injected mice by studying H and E-stained tissue sections, DNA ladder analysis, caspase-3-like activity assay, immunohistochemistry, double immunofluorescence microscopy for activated caspase-3 and cytokeratin (CK) 18, the TUNEL method, and electron microscopy.
Results: Jo2-treated mice showed activation of caspase-3, breakdown of the CK intermediate filament network, and classical morphological features of apoptosis. In contrast, in CA-fed and CBDL mice, oncosis characterized by cell swelling and ruptured cell membranes was the predominant type of cell death, whereas in both experimental conditions significant activation of caspase-3 was absent and typical CK alterations were rare despite frequent positivity of the TUNEL assay.
Conclusions: (i) Oncosis represents the main type of hepatocyte death in acute cholestasis in mice. (ii) The importance of apoptosis in cholestasis may be overestimated if non-specific detection systems (e.g. TUNEL assay) are used.
Antibody titer to gp210-C terminal peptide as a clinical parameter for monitoring primary biliary cirrhosis, 25 November 2004
Nakamura M, Shimizu-Yoshida Y, Takii Y, Komori A, Yokoyama T, Ueki T, Daikoku M, Yano K, Matsumoto T, Migita K, Yatsuhashi H, Ito M, Masaki N, Adachi H, Watanabe Y, Nakamura Y, Saoshiro T, Sodeyama T, Koga M, Shimoda S, Ishibashi H
Background/Aims: The presence of antibodies to the 210-kDa glycoprotein of the nuclear pore complex (gp210) is highly indicative of primary biliary cirrhosis (PBC). However, the significance of anti-gp210 antibody titers for monitoring PBC remains unresolved.
Methods: We used an ELISA with a gp210 C-terminal peptide as an antigen to assess serum antibody titers in 71 patients with PBC.
Results: Patients were classified into three groups: Group A in whom anti-gp210 titers were sustained at a high level, Group B in whom anti-gp210 status changed from positive to negative under ursodeoxycholic acid (UDCA) therapy, Group C in whom anti-gp210 antibodies were negative at the time of diagnosis. The rate of progression to end-stage hepatic failure was significantly higher in group A (60%) as compared to groups B (0%) and C (4.2%). The sustained antibody response to gp210 was closely associated with the severity of interface hepatitis. The significance of anti-gp210 antibody was confirmed by National Hospital Organization Study Group for Liver Disease in Japan.
Conclusions: The serial quantitation of serum anti-gp210-C-terminal peptide antibodies is useful for monitoring the effect of UDCA and for the early identification of patients at high risk for end-stage hepatic failure.
Markers of cell activation and apoptosis in bone marrow mononuclear cells of patients with autoimmune hepatitis type 1 and primary biliary cirrhosis, 29 November 2004
Tsikrikoni A, Kyriakou DS, Rigopoulou EI, Alexandrakis MG, Zachou K, Passam F, Dalekos GN
Background/Aims: We have reported quantitative and qualitative differences in bone marrow (BM) progenitor cells in autoimmune hepatitis type-1 (AIH-1) and primary biliary cirrhosis (PBC). This study investigated the apoptotic features and cytokine suppressors of haematopoiesis in long-term cultures of BM mononuclear cells (BMMCs) from AIH-1 and PBC patients.
Methods: Apoptotic markers and CD14 expression were evaluated in 13 AIH-1 patients, 13 PBC patients, 12 cirrhotic controls and 10 healthy subjects. TNF-?, TGF-? and IFN-? were determined using ELISAs.
Results: All apoptotic markers and CD14 were increased in AIH-1 and PBC compared to controls (P<0.0001). Fas+ cells were positively correlated (P=0.0001) with apoptotic cells in AIH-1 and PBC. TNF-? and IFN-? were higher in AIH-1 (P=0.003 and P=0.001) and PBC (P=0.0001) compared to controls. No differences were found between the control groups.
Conclusions: We demonstrate for the first time that the apoptotic process, macrophage activation and the production of cytokine suppressors of haematopoiesis in BMMCs from AIH-1 and PBC patients are higher compared to controls. The Fas–FasL pathway is likely to be involved in the apoptotic process; the increased levels of selected cytokines may contribute to Fas–FasL stimulation. Cirrhosis appears unlikely to be the cause of the above findings.
Genetic and Metabolic Liver Disease
Diet induced regulation of genes involved in cholesterol metabolism in rat liver parenchymal and Kupffer cells, 24 December 2004
Hoekstra M, Out R, Kar Kruijt J, Van Eck M, Van Berkel TJC
Background/Aims: Feeding rodents atherogenic diets enriched in cholesterol or cholic acid changes hepatic cholesterol metabolism. In the present study, the effect of an atherogenic diet enriched in cholesterol and cholic acid on cellular hepatic cholesterol metabolism was studied.
Methods: Gene and protein expression analysis was performed on parenchymal, endothelial, and Kupffer cells isolated from rats fed a chow or atherogenic diet using quantitative real-time PCR and immunoblotting, respectively.
Results: The atherogenic diet raised the serum cholesterol concentration 11-fold, mostly in the VLDL fraction, and led to heavy lipid loading of rat liver parenchymal and Kupffer cells. Only moderate changes in the expression of genes involved in cholesterol metabolism were observed in parenchymal cells on the diet, while PPAR? expression was 6.8-fold decreased. Kupffer cells, however, showed a highly adaptive response with a 2- to 9-fold induction of SR-BI, ABCA1, and ABCG5/G8, and an 82-fold induction in CYP7A1 mRNA expression, respectively.
Conclusions: Heavy lipid loading of parenchymal cells leads to moderate gene expression changes, while Kupffer cells respond in a highly adaptive fashion by stimulating the expression of genes involved in cholesterol metabolism and transport.
Copyright © 2001-2005 European Association for the Study of the Liver. All rights reserved.
