bluuebunny/crossref_metadata_2025 huggingface dataset:
Living-related liver transplantation for patients with fulminant and subfulminant hepatic failure
Authors: Miwa Shiro , Hashikura Yasuhiko , Mita Atsuyoshi , Kubota Tatsuya , Chisuwa Hisanao , Nakazawa Yuichi , Ikegami Toshihiko , Terada Masaru , Miyagawa Shinichi , Kawasaki Seiji
Published: July 2004

The prognosis for patients with fulminant (FHF) or subfulminant hepatic failure (SFHF) has improved since the introduction of liver transplantation. However, the death rate of patients awaiting liver transplantation is high, possibly because of the difficulty in obtaining grafts in a timely manner, given the relative shortage of cadaveric donors. Between June 1990 and June 1999, 106 patients underwent living-related liver transplantation (LRLT) at Shinshu University Hospital. Among them, 8 patients had FHF and 6 had SFHF; these 14 patients are the subjects of this report. The graft volumes (GV) ranged from 231 mL to 625 mL, corresponding to 35% to 105% of the recipients' standard liver volume (SLV). The postoperative courses of all donors were uneventful. Following liver transplantation, all grafts functioned favorably, with normalization of serum total bilirubin within 3 to 5 days and normalization of coagulation profiles within 4 to 7 days. Thirteen of the 14 recipients are still alive. The actuarial 6-month, 1-year, and 5-year survival rates were 100%, 90%, and 90%, respectively. In the present study, when the ratio of the GV to the recipient's SLV was more than 35%, the graft was able to support the patient's metabolic demand after liver transplantation for FHF or SFHF. Because of the urgent nature of liver transplantation in this clinical condition, concerns over informed consent may be even greater than for elective LRLT. Nevertheless, the high success rate and low donor risk may justify this option for pediatric patients, as well as for a limited population of adult patients suffering from FHF or SFHF.
Protective effect of liver ischemic preconditioning on liver and lung injury induced by hepatic ischemia-reperfusion in the rat
Authors: Peralta Carmen , Prats Neus , Xaus Carme , Gelpí Emilio , Roselló-Catafau Joan
Published: July 2004

This study evaluates whether preconditioning could modulate the injurious effects of tumor necrosis factor (TNF) on liver and lung following hepatic ischemia-reperfusion (I/R) by inhibiting hepatic postischemic TNF release. The inhibition of hepatic TNF release from Kupffer cells with gadolinium chloride (GdCl3) previous to ischemia maintained TNF at control levels, attenuating the increases in transaminases, vascular permeability, and edema associated with hepatic I/R injury. TNF addition reverted this beneficial effect, indicating the implication of the TNF released mainly from Kupffer cells in hepatic I/R injury. Preconditioning prevented hepatic TNF increases, thus attenuating the liver injury, while TNF addition abolished the benefits of preconditioning. Inhibition of nitric oxide (NO) synthesis abolished the effect of preconditioning, whereas GdCl3 addition avoided the injurious effect of NO inhibition. In addition, NO administration before I/R offered similar results to those found in preconditioning, while TNF addition abolished the benefits of NO. Thus, the effect of preconditioning on TNF release after hepatic I/R is mediated by NO. Inhibition of hepatic TNF release from Kupffer cells with GdCl3 prevented both the increase in plasma TNF and the injurious effect in lung seen after hepatic I/R, and these effects were reverted with TNF addition. Preconditioning resulting in reduced hepatic TNF levels prevented the systemic TNF release, thus reducing the lung damage following hepatic I/R. However, TNF addition abolished the protective effect of preconditioning on lung injury. These findings indicate that preconditioning attenuates hepatic postischemic TNF release from Kupffer cells, thus probably reducing the liver and lung injury following hepatic I/R, and that this effect of preconditioning is mediated by NO.
Wedged hepatic venous pressure adequately reflects portal pressure in hepatitis C virus-related cirrhosis
Authors: Escorsell Àngels , Bru Concepció , Gilabert Rosa , Moitinho Eduardo , García-Pagán Juan Carlos , Bosch Jaume
Published: July 2004

Wedged hepatic venous pressure (WHVP) is equivalent to portal venous pressure in patients with alcoholic liver diseases. However, it may underestimate portal pressure in nonalcoholics, which is important because hepatitis C virus (HCV) infection is a frequent cause of chronic liver disease. We investigated the agreement between directly measured portal pressure and WHVP in alcoholic and HCV-related liver diseases. Seventy-one patients with liver disease resulting from HCV infection (n = 32), alcohol (n = 25), or both (n = 14) underwent simultaneous measurements of WHVP (by hepatic vein catheterization) and portal pressure (by direct puncture). In 9 patients, measurements were repeated 20 minutes after acute iv propranolol administration. WHVP showed an excellent agreement with portal pressure in patients with cirrhosis resulting from either HCV, alcohol or both (intraclass correlation coefficient: 0.94, 0.93, and 0.97, respectively;P< .001). A discrepancy of ≥5 mm Hg was observed in 7 cases. WHVP underestimated portal pressure in only 1 case and exceeded portal pressure by ≥5 mm Hg in 6 patients. The WHVP response to propranolol closely and significantly correlated with changes in portal pressure (intraclass correlation coefficient: 0.87; P< .004). The simple and safe measurement of WHVP accurately reflects portal pressure in alcoholic and HCV-related liver disease. This technique also allows us to accurately assess the portal pressure response to propranolol in both alcoholic and HCV-related cirrhosis.
Intention-to-treat analysis of surgical treatment for early hepatocellular carcinoma: Resection versus transplantation
Authors: Llovet Josep M. , Fuster Josep , Bruix Jordi
Published: July 2004

Liver transplantation is proposed as the best therapy for early hepatocellular carcinoma in cirrhotic patients. However, the confrontation with the results obtained by surgical resection has never been done on an intention-to-treat basis. Between 1989 and 1997, 164 out of 1,265 patients with hepatocellular carcinoma were evaluated for surgery. Seventy-seven (48 men, mean 61 years of age, 74 Child-Pugh class A, size 33 ± 18 mm) were resected (first line option) and 87 (65 men, mean 55 years of age, 50 Child-Pugh class B/C, size 24 ± 14 mm) were selected for transplantation. The 1-, 3-, and 5-year “intention-to-treat” survival was 85%, 62%, and 51% for resection and 84%, 69%, and 69% for transplantation (8 drop-outs on waiting list). Bilirubin and clinically relevant portal hypertension were independent survival predictors after resection. Thereby, the 5-year survival of the best candidates (absence of clinically relevant portal hypertension, n = 35) was 74%, whereas it was 25% for the worst candidates (portal hypertension and bilirubin ≥1 mg/dL, n = 27) (P < .00001). The variable “drop-out on waiting list” was the sole survival predictor after transplantation. The 2-year survival rate of patients evaluated for transplantation was 84% in the 1989 to 1995 period (mean waiting time, 62 days; no drop-outs) and 54% during 1996 to 1997 (mean waiting time, 162 days; 8 drop-outs)(P < .003). This outcome was significantly lower than that of the best candidates for resection (P = .002). In conclusion, a proper selection of candidates for resection promotes better results than transplantation, in which the results are significantly hampered by the growing incidence of drop-outs because of the increasing waiting time.
Cholangiocarcinoma in patients with primary sclerosing cholangitis: A multicenter case-control study
Authors: Chalasani Naga , Baluyut Arthur , Ismail Ayaaz , Zaman Atif , Sood Gagan , Ghalib Reem , McCashland T. M. , Reddy Rajender K. , Zervos Xaralambos , Anbari Mann A. , Hoen Helena
Published: July 2004

Patients with primary sclerosing cholangitis (PSC) have a significantly increased risk of developing cholangiocarcinoma (CCA). Risk factors for developing such a complication are not well defined. We conducted a multicenter, case-control study to determine the risk factors and possible predictors for CCA in patients with PSC. The demographic, clinical, and laboratory features of 26 PSC patients with CCA diagnosed over a 7-year period at eight academic centers were compared with 87 patients with PSC but no CCA (controls). There was no statistically significant difference in demographics, smoking, signs or symptoms or complications of PSC, indices of disease severity (Mayo Risk score or Child-Pugh score), frequency or duration or complications of inflammatory bowel disease (IBD), frequency of biliary surgery, or therapeutic endoscopy between the two groups. Alcohol consumption was significantly associated with CCA in patients with PSC (odds ratio: 2.95; 95% CI: 1.04-8.3). Serum carbohydrate antigen 19-9 (CA 19-9) was significantly higher in patients with CCA than those without (177 ± 89 and 61 ± 58 U/mL, respectively;P = .002). A serum CA 19-9 level > 100 U/mL had 75% sensitivity and 80% specificity in identifying PSC patients with CCA. In conclusion, alcohol consumption was a risk factor for having CCA in PSC patients. The indices of severity of liver disease were not associated with CCA in patients with PSC. Serum CA 19-9 appeared to have good ability to discriminate PSC patients with and without CCA.
Clinicopathologic features of the intraductal growth type of peripheral cholangiocarcinoma
Authors: Suh Kyung-Suk , Roh Hye Rin , Koh Young Taeg , Lee Kuhn Uk , Park Yong-Hyun , Kim Sun-Whe
Published: July 2004

The clinicopathologic features of the intraductal growth (IG) type of peripheral cholangiocarcinoma (PCC) were examined retrospectively. Out of 112 patients who underwent surgery for PCC at Seoul National University Hospital, Seoul, Korea, between January 1980 and December 1997, 16 were classified as having the IG type. Thirteen were men and 3 were women. Their ages ranged from 38 to 73 years with a mean age of 55.9 years. Abdominal pain was the most common symptom and jaundice was found in 18.8%. Five patients had associated clonorchiasis and 6 patients experienced hepatolithiasis. Tumor sizes ranged from 0.5 cm to 14.0 cm with a mean of 4.3 cm. The tumor was located in the right lobe in 7 cases and the left lobe in 9 cases. Thirteen patients underwent major hepatic resection, and 3 underwent minor resection, involving a subsegmentectomy. Mucin was found in the bile in 4 cases. Pathology showed papillary adenocarcinoma with a background of adenomatous hyperplasia, and the absence of lymph node metastasis in all cases. The tumors were confined to the mucosa in 5 cases. Out of the 16 hepatic resections, 1 was palliative due to a positive margin. In 2 of the patients who underwent minor resections, recurrences developed, and 1 of these died 56 months after resection. Out of the 16 patients, 15 remain alive, ranging from 1 to 13 years, postoperatively. In conclusion, the IG type of PCC should be distinguished from other types of PCC because a favorable prognosis can be expected after complete surgical resection.
Apolipoprotein E genotype and the risk of gallbladder disease in pregnancy
Authors: Ko Cynthia W. , Beresford Shirley A.A. , Alderman Beth , Jarvik Gail P. , Schulte Scott J. , Calhoun Byron , Tsuchida Amy M. , Koepsell Thomas D. , Lee Sum P.
Published: July 2004

The E4 allele of apolipoprotein E (apoE4) has previously been associated with symptomatic gallstone disease. The aim of this study was to determine if apoE4 is associated with the development of gallbladder sludge and/or stones during pregnancy. We conducted a nested case-control study based on an ongoing cohort study of gallbladder disease in pregnancy. Women in this study receive gallbladder ultrasounds in each trimester of pregnancy. Cases (n = 52) were defined as women with incident gallbladder sludge or stones diagnosed at the third trimester ultrasound. Controls (n = 104) were defined as women without gallbladder sludge or stones on any of 3 study ultrasounds. ApoE genotyping was performed from stored white blood cell pellets. Data were analyzed by stratified analysis and multivariate logistic regression. Cases and controls were similar in baseline characteristics. Forty-two women had sludge, 6 had gallstones, and 4 had both sludge and stones. After adjusting for risk factors such as age, parity, and body mass index, the odds ratio (OR) for the association between heterozygosity or homozygosity for the apoE4 allele and incident gallbladder sludge or stones was 0.91 (95% confidence interval [CI], 0.41-2.02). Further adjustment for family medical history and serum lipid levels did not substantially change these results (OR, 0.73; 95% CI, 0.29-1.82). In conclusion, apoE4 appears to have little or no overall association with the development of new gallbladder sludge or stones in pregnancy. However, an effect could not be ruled out in certain subgroups, such as blacks or women who are homozygous for apoE4.
Evidence for a locally driven mucosal response and the presence of mitochondrial antigens in saliva in primary biliary cirrhosis
Authors: Reynoso-Paz Sandra , Leung Patrick S.C. , Van de Water Judy , Tanaka Atsushi , Munoz Santiago , Bass Nathan , Lindor Keith , Donald Paul J. , Coppel Ross L. , Ansari Aftab A. , Gershwin Eric M.
Published: July 2004

Primary biliary cirrhosis (PBC) is often considered to be a dry gland disease caused by frequent involvement of salivary and lacrimal glands. Although high titers of antimitochondrial autoantibodies (AMA) have long been recognized in PBC, little is known about the presence of mitochondrial autoantigens in mucosal compartments such as saliva. We investIgAted saliva and sera in PBC patients and controls for the presence of AMA and mitochondrial antigens. In PBC saliva, AMA were detected in 45 of 49 (92%), with specificity directed against pyruvate dehydrogenase complex (PDC-E2) alone in 22 of 49 (45%), against PDC-E2 and branched-chain 2-oxo-acid dehydrogenase complex E2 (BCOADC-E2) in 4 of 49 (8%), to PDC-E2 and 2-oxoglutarate dehydrogenase complex E2 (OGDC-E2) in 9 of 49 (18%), and to the 3 antigens together in 10 of 49 (20%). Isotyping of the saliva AMA showed that 80% of the patients had immunoglobulin A (IgA) against PDC-E2, 18% had IgM-specific PDC-E2, and 35% had IgG specific PDC-E2. Similar to serum and bile anti-PDC-E2 IgA antibodies, the saliva autoantibodies localized their reactivity to the inner lipoyl domain of PDC-E2. Furthermore, saliva from patients with PBC but not controls inhibited pyruvate dehydrogenase enzyme activityin vitro. In addition, and of particular interest, we detected a molecule with a molecular weight corresponding to PDC-E2 (74 kd) in PBC but not control saliva. These findings make several important points: first, there appears to be localized mucosal immunity in the secretory system of PBC; second, AMA are readily detected in PBC saliva; and third, PDC-E2 may be present in the saliva of PBC.
Partial pressure of ammonia versus ammonia in hepatic encephalopathy
Authors: Kramer Ludwig , Tribl Barbara , Gendo Alexandra , Zauner Christian , Schneider Barbara , Ferenci Peter , Madl Christian
Published: July 2004

Ammonia is considered the major pathogenetic factor of cerebral dysfunction in hepatic failure. The correlation between total plasma ammonia and the severity of hepatic encephalopathy (HE), however, is variable. Because ammonia that is present in gaseous form readily enters the brain, the correlation with the grade of HE of the pH-dependent partial pressure of gaseous ammonia (pNH3) could be better than that of total arterial ammonia levels. To test this hypothesis, 56 cirrhotic patients with acute episodes of clinical HE (median age, 54 years; range, 21-75) were studied by clinical examination and by long-latency median-nerve sensory-evoked potentials (SEPs) N70 peak, an objective and sensitive electrophysiological measure of HE. pNH3 was calculated from arterial blood according to published methods. The clinical grade of HE correlated (P < .001) with both pNH3 and total ammonia, but correlation was stronger with pNH3 (r = .79 vs. .69, P = .01). A similar correlation was found for N70 peak latency (r = .71 with pNH3 vs. .64 with total ammonia, respectively, P = .08). In summary, arterial pNH3 correlates more closely than total ammonia with the degree of clinical and electrophysiological abnormalities in HE. These findings support the ammonia hypothesis of HE and suggest that pNH3 might be superior to total ammonia in the pathophysiological evaluation of HE.
Failure of regeneration of the steatotic rat liver: disruption at two different levels in the regeneration pathway
Authors: Selzner Markus , Clavien Pierre-Alain
Published: July 2004

Hepatic resection or transplantation in patients with fatty liver is associated with increased morbidity and mortality. The regenerative capacity of fatty livers after major tissue loss is unknown. Interleukin 6 (IL-6) is a potent inducer of hepatic regeneration in normal and ischemic livers. Therefore, we studied hepatic regeneration at day 1, day 2, and day 4 in a model of 70% hepatectomy in obese and lean Zucker rats, and obese Zucker rats pretreated with recombinant interleukin 6 (rIL-6). The mitotic cycle in hepatocytes was investigated by 4 different markers of regeneration representing distinct phases of mitosis (proliferating cell nuclear antigen [PCNA] = G1 phase, bromodeoxy uridine [BrdU] = S phase, mitotic index, and regenerated liver weight = M phase). Obese Zucker rats had significantly decreased regenerative capacity compared with lean Zucker rats (PCNA, BrdU, mitotic index, regenerated liver weight) at days 1 and 2 after surgery. Four days after resection fatty animals showed an increase in the mitotic index indicating a delay of regeneration in steatotic livers. Animal survival after 70% hepatectomy was significantly decreased in obese rats compared with lean animals. Pretreatment of obese animals with rIL-6 normalized PCNA expression (G1 phase) in steatotic hepatocytes but failed to increase DNA synthesis (BrdU, S phase), mitosis (mitotic index and regenerated liver weight, M phase), and animal survival. These results indicate major impairment of hepatic regeneration in steatotic livers. Two different blockages of regeneration must be present, one rIL-6 sensitive, at the level of IL-6 or upstream, and a second, rIL-6 resistant, at the level of G1 /S-phase transition.
Centrosome abnormalities in human carcinomas of the gallbladder and intrahepatic and extrahepatic bile ducts
Authors: Kuo Kung-Kai , Sato Norihiro , Mizumoto Kazuhiro , Maehara Naoki , Yonemasu Hirotoshi , Ker Chen-Guo , Sheen Pai-Ching , Tanaka Masao
Published: July 2004

During mitosis, 2 centrosomes ensure accurate assembly of bipolar spindles and fidelity of the chromosomal segregation. The presence of more than 2 copies of centrosomes during mitosis can result in the formation of multipolar spindles, unbalanced chromosome segregation, and aneuploidy. Recent studies have provided evidence that centrosome hyperamplification plays a pivotal role in carcinogenesis. Using immunofluorescence analysis with γ-tubulin and pericentrin antibodies, paraffin-embedded sections from 40 malignant biliary diseases including gallbladder cancers (GC; n = 13), intrahepatic cholangiocellular carcinoma (CCC; n = 19), and extrahepatic bile duct cancers (BDC; n = 8) were examined. Thirty-seven benign biliary diseases including chronic cholecystitis, gallbladder adenoma, hepatolithiasis, and choledochal cyst were included as benign controls. The frequencies of the centrosome abnormalities were 70% for GC, 58% for CCC, and 50% for BDC, respectively. The frequencies of centrosome abnormalities in malignant biliary diseases were significantly higher than in their benign counterparts (GC, CCC, BDC; P = .001, .002, and .001, respectively). The results of current study also indicated that biliary malignancy in the advanced stage (III-IV) displayed a higher frequency of centrosome abnormalities than in the early stage (I-II) (P < .001). We conclude that abnormalities in size, number, and shape of the centrosome are frequently observed in biliary tract malignancy. Centrosome abnormalities started to occur in the early stage of biliary malignancy and became very frequent in the advanced stage. This implies that centrosome abnormality might relate to the transition from early to advanced malignancy in biliary malignancy.
Sublobular veins as the main site of lymphocyte adhesion/transmigration and adhesion molecule expression in the porto-sinusoidal-hepatic venous system during concanavalin A-induced hepatitis in mice
Authors: Morikawa Hiroyasu , Hachiya Kazutaka , Mizuhara Hidekazu , Fujiwara Hiromi , Nishiguchi Shuhei , Shiomi Susumu , Kuroki Tetsuo , Kaneda Kenji
Published: July 2004

Lymphocyte infiltration is a manifest feature of hepatitis. To reveal the main site and mechanism of lymphocyte adhesion/extravasation in the hepatic vasculature during inflammation, we morphometrically and histologically analyzed these events in relation to adhesion molecule expression using a murine model of T-cell mediated hepatitis induced by concanavalin A (Con A). Although lymphocyte adhesion was restricted to the sinusoids in untreated mice, it increased in all the segments of porto-sinusoidal-hepatic venous system 8 hours after Con A injection; the number of adhering lymphocytes per unit vascular circumference was the largest in the sublobular veins, relatively large in the central veins and small hepatic veins, and relatively small in the sinusoids and negligible in the portal veins. At 20 hours, extravascular lymphocytes showed similar distribution to lymphocyte adhesion at 8 hours except in the portal veins, around which they were possibly accumulated by the translocation of extrasinusoidal lymphocytes. E-selectin and vascular cell adhesion molecule-1 (VCAM-1) were transiently expressed at 4 to 6 hours, whereas P-selectin and intercellular adhesion molecule-1 were not changed between 0 and 48 hours. In particular, E-selectin expression coincided with that of lymphocyte adhesion in distribution. Lymphocyte attachment was inhibited by pretreatment with anti-E-selectin monoclonal antibody (MAb) or anti-VCAM-1 MAb, and expression of E-selectin and VCAM-1 was suppressed by pretreatment with anti-tumor necrosis factor-α (TNF-α) MAb. Electron microscopically, lymphocytes were trapped by endothelial lamellipodia and traversed the endothelium by diapedesis. These results indicate that lymphocyte adhesion/transmigration preferentially takes place in the sublobular veins in association with TNF-α-induced endothelial activation, i.e., E-selectin and VCAM-1 expression and lamellipodia formation.
The role of phosphatidic acid in platelet-derived growth factor-induced proliferation of rat hepatic stellate cells
Authors: Reeves Helen L. , Thompson Mike G. , Dack Clare L. , Burt Alastair D. , Day Christopher P.
Published: July 2004

Platelet-derived growth factor (PDGF) is the most potent mitogen for hepatic stellate cells (HSCs) in vitro. The aim of this study was to investigate the role of the lipid-derived second messenger phosphatidic acid (PA) in mediating this effect and, in particular, to determine its interaction with the extracellular signal-regulated kinase (ERK) cascade. HSCs were isolated from rat livers. PA production was determined by lipid extraction and thin-layer chromatography (TLC) after prelabeling cells with [3H]myristate. ERK activity was measured by an in vitro kinase assay after immunoprecipitation. Mitogenic concentrations of PDGF, but not those of the relatively less potent mitogen, transforming growth factor α (TGF-α), stimulated the sustained production of PA from HSCs. Exogenous PA stimulated HSC proliferation and a sustained increase in ERK activity, and proliferation was completely blocked by the inhibition of ERK activation with PD98059. The stimulation of ERK by PDGF was of a similar magnitude but more sustained than that caused by TGF-α. These results suggest that the potent mitogenic effect of PDGF in HSCs may be caused, in part, by the generation of PA and subsequently by a more sustained activation of ERK than occurs with less potent mitogens that do not induce the production of this lipid second messenger.
Hydrogen peroxide: A link between acetaldehyde-elicited α1(i) collagen gene up-regulation and oxidative stress in mouse hepatic stellate cells
Authors: Greenwel Patricia , Domínguez-Rosales José-Alfredo , Mavi Gurjeet , Rivas-Estilla A. M. , Rojkind Marcos
Published: July 2004

Ethanol induces liver fibrosis by several means that include, among others, the direct fibrogenic actions of acetaldehyde and the induction of an oxidative stress response. However, the mechanisms responsible for these activities, and the possible connections between oxidative stress and acetaldehyde-induced fibrosis are not well understood. In this communication we investigated the molecular mechanisms whereby acetaldehyde induces mouse α1(I) procollagen (col1a1) gene expression in cultured hepatic stellate cells. Transfection assays using reporter plasmids driven by different segments of the col1a1 promoter localized an acetaldehyde-responsive element (AcRE) between nucleotides −370 and −345. We also show that acetaldehyde enhances binding of a CCAAT/enhancer binding protein-β (C/EBPβ)-containing complex to this element, and that this effect is due, at least in part, to an increase in the concentration of nuclear p35C/EBPβ protein. Although this element overlaps to a previously described transforming growth factor β1 (TGF-β1)-responsive element, the stimulatory effect of acetaldehyde is not mediated through this cytokine, because addition of neutralizing anti-TGF-β1 antibodies does not prevent acetaldehyde-elicited col1a1 up-regulation. On the other hand, this effect is blocked by the addition of catalase, an H2O2 scavenger. Moreover, this ethanol metabolite stimulates production of H2O2 in stellate cells. Thus, these results suggest that acetaldehyde-induced col1a1 up-regulation is mediated, at least in part, through H2O2. Altogether, these data suggest that the −370 to −344 region of the col1a1 gene is a point of convergence of the action of numerous extracellular stimuli that ultimately leads to col1a1 up-regulation. In addition, we have established a direct connection between oxidative stress and enhanced col1a1 expression induced by acetaldehyde.
Estrogen is involved in early alcohol-induced liver injury in a rat enteral feeding model
Authors: Yin Ming , Ikejima Kenichi , Wheeler Michael D. , Bradford Blair U. , Seabra Vitor , Forman Donald T. , Sato Nobuhiro , Thurman Ronald G.
Published: July 2004

The aim of this study was to investigate whether reduction in blood estrogen by removal of the ovaries would decrease the sensitivity of female rats to early alcohol-induced liver injury using an enteral ethanol feeding model, and if so, whether estrogen replacement would compensate. Livers from ovariectomized rats with or without estrogen replacement after 4 weeks of continuous ethanol exposure were compared with nonovariectomized rats in the presence or absence of ethanol. Ethanol increased serum alanine transaminase (ALT) levels from 30 ± 6 to 64 ± 7 U/L. This effect was blocked by ovariectomy (31 ± 7) and totally reversed by estrogen replacement (110 ± 23). Ethanol increased liver weight and fat accumulation, an effect that was minimized by ovariectomy and reversed partially by estrogen replacement. Infiltrating leukocytes were increased 6.7-fold by ethanol, an effect that was blunted significantly by ovariectomy and reversed by estrogen replacement. Likewise, a similar pattern of changes was observed in the number of necrotic hepatocytes. Blood endotoxin and hepatic levels of CD14 messenger RNA (mRNA) and protein were increased by ethanol. This effect was blocked in ovariectomized rats and elevated by estrogen replacement. Moreover, Kupffer cells isolated from ethanol-treated rats with estrogen replacement produced more tumor necrosis factor α (TNF-α) than those from control and ovariectomized rats. It is concluded, therefore, that the sensitivity of rat liver to alcohol-induced injury is directly related to estrogen, which increases endotoxin in the blood and CD14 expression in the liver, leading to increased TNF-α production.
Endotoxin-stimulated macrophages decrease bile acid uptake in WIF-B cells, a rat hepatoma hybrid cell line
Authors: Sturm Ekkehard , Zimmerman Tracy L. , Crawford Aleta R. , Svetlov Stanislav I. , Sundaram Pazhani , Ferrara J. L. , Karpen Saul J. , Crawford James M.
Published: July 2004

Endotoxemia leads to cytokine-mediated alterations of the hepatocellular sodium-taurocholate-cotransporting polypeptide (ntcp). We hypothesized that stimulated macrophages are essential transducers for down-regulating hepatocellular bile salt uptake in response to endotoxin (lipopolysaccharide [LPS]) exposure. Using anin vitromodel, we exposed mouse macrophages (IC-21 cell line) to LPS for 24 hours. Concentrations of cytokines tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, and IL-6 increased 10.6-fold, 12.5-fold, and 444-fold, respectively, in LPS-conditioned IC-21 medium (CM) versus unconditioned IC-21 medium (UM). WIF-B rat hepatoma hybrid cells were incubated with either CM or UM or treated directly with medium containing recombinant TNF-α, IL-1β, and IL-6. [3H]Taurocholate ([3H]TC) uptake decreased in WIF-B cells exposed to either TNF-α (54% of control), IL-1β (78%), IL-6 (55%) as single additives, or in triple combination (TCC) (43%). A virtually identical decrease was observed after exposing WIF-B cells to CM (52%,P< .001). LPS had no direct effect on [3H]TC uptake. CM treatment did not decrease L-alanine transport in WIF-B cells. Blocking antibodies against TNF-α, IL-1β, and IL-6 restored the diminished [3H]TC uptake in cells exposed to TCC and CM to 87% and 107% of controls, respectively. Northern blotting revealed thatntcpmessenger RNA (mRNA) expression was significantly reduced in WIF-B cells after exposure to CM, and in primary rat hepatocytes exposed to CM or TNF-α (68%, 14%, and 29% of control, respectively). We conclude that macrophages and their ability to secrete the cytokines TNF-α, IL-1β, and IL-6 may be essential in mediating the endotoxin-induced cholestatic effect of decreased hepatocellular bile salt uptake.
Hypoxic stimulation of vascular endothelial growth factor expression in activated rat hepatic stellate cells
Authors: Ankoma-Sey Victor , Wang Yun , Dai Zhihao
Published: July 2004

The tissue repair response to hypoxic stimuli during wound healing includes enhanced production of angiogenic factors, such as vascular endothelial growth factor (VEGF). Hepatic stellate cells are oxygen-sensing cells, capable of producing VEGF. We hypothesized that hypoxia-stimulated signaling in activated stellate cells mediate VEGF secretion during liver injury. The specific aim was to evaluate the effect of hypoxia on the gene expression of VEGF in HSC-T6 cells, an immortalized rat hepatic stellate cell line, and in rat primary cultures of stellate cells. Hypoxic induction of VEGF mRNA was dose- and time-dependent. The hypoxic stimulation of VEGF messenger RNA (mRNA) correlated with the secretion of VEGF protein in conditioned media by hypoxic T6 cells. S-Nitroso-N-acetyl-D,L-penicillamine (SNAP), a nitric oxide (NO) donor, and desferrioxamine (DFx) and cobalt chloride, mimics of cellular hypoxia, similarly stimulated VEGF mRNA expression and secretion. Four previously described splice variants of the VEGF mRNA (VEGF-120, 144, 164, 188) were detected in both normoxic- or hypoxic-activated stellate cells. There was differential expression of the VEGF receptors, Flt-1 and Flk-1, in hypoxic T6 cells. Hypoxic conditions selectively stimulated Flt-1 mRNA expression, whereas Flk-1 mRNA remained unchanged. Hypoxic induction of VEGF was also demonstrated in primary stellate cell cultures and after in vivo injury. Hypoxia stimulates cell signaling in stellate cells, culminating in the rapid induction of VEGF and Flt-1 mRNA expression and VEGF secretion. The hypoxic induction of VEGF is mimicked by NO and may be of mechanistic importance in the pathogenesis of hepatic wound healing and hepatocarcinogenesis.
Increased toxin-induced liver injury and fibrosis in interleukin-6-deficient mice
Authors: Kovalovich Kellen , DeAngelis Robert A. , Li Wei , Furth Emma E. , Ciliberto Gennaro , Taub Rebecca
Published: July 2004

Interleukin-6 null (IL-6−/−) mice have impaired liver regeneration and increased liver necrosis following partial hepatectomy that is corrected with IL-6 treatment. Following acute carbon tetrachloride (CCl4) treatment, we found that IL-6−/− mice developed increased hepatocellular injury and defective regeneration with significant blunting of signal transducer-and-activator of transcription protein 3 (STAT3) and nuclear factor-κB (NF-κB) activation and reduced hepatocyte DNA synthetic and mitotic responses. After CCl4 treatment, unlike partial hepatectomy, increased hepatocyte apoptosis was noted in IL-6−/− livers. Pretreatment with IL-6 before CCl4 reduced acute CCl4 injury and apoptosis and accelerated regeneration in both IL-6+/+ and −/− livers. Repetitive doses of CCl4 in the presence or absence of phenobarbital resulted in increased injury and fibrosis in IL-6 −/− compared with +/+ livers. After acute and chronic injury, IL-6−/− livers showed the protracted presence of α-smooth muscle actin associated with activated stellate cells, indicating a disturbed response in wound healing that progressed to fibrosis. These data provide evidence for an important role for IL-6 in reducing CCl4-induced acute and chronic liver injury and fibrosis.
Role of oxidative stress in hypoxia-reoxygenation injury to cultured rat hepatic sinusoidal endothelial cells
Authors: Samarasinghe Dev A. , Tapner Michael , Farrell Geoffrey C.
Published: July 2004

To characterize the role of oxidative stress in cultured rat sinusoidal endothelial cells, we studied the production of superoxide after reoxygenation, the relationship of reduced glutathione (GSH) levels to cell injury, and the protective efficacy of antioxidants. Hypoxia (pO2 1-2 mm Hg) was achieved by culturing cells under 95% N2 5% CO2 for 4 hours. Reoxygenation was then reestablished, and viability was determined at 24 hours by trypan blue exclusion; putative protective agents were added at the time of reoxygenation (4 hours). As previously reported, reoxygenation after 4 hours hypoxia accentuated sinusoidal cell death fourfold compared with hypoxic or normoxic controls (P < .0001). Superoxide was not produced on reoxygenation, and superoxide dismutase provided no protection against reoxygenation injury. Cellular levels of GSH fell to 37 ± 4% of normoxic controls (P < .0001) following reoxygenation. These changes were essentially abrogated by Trolox (Aldrich Chemical Co., Milwaukee, WI) and dimethyl sulfoxide, both of which also completely protected against reoxygenation injury. When cellular GSH levels were lowered by addition of diethylmaleate (which conjugates GSH), this reduced the viability of endothelial cells cultured under normoxic conditions and greatly augmented reoxygenation injury. Conversely, addition of exogenous GSH partially protected endothelial cells against hypoxia-reoxygenation injury. Desferrioxamine also protected against reoxygenation injury, but catalase was only partly protective. It is concluded that sinusoidal endothelial cells undergo significant intracellular oxidative stress following reoxygenation, and their viability is critically dependent on GSH levels. Reactive oxygen species are likely mediators of oxidative stress in hepatic sinusoidal endothelial cells.
Ischemic preconditioning reduces Na+ accumulation and cell killing in isolated rat hepatocytes exposed to hypoxia
Authors: Carini Rita , De Cesaris Maria Grazia , Splendore Roberta , Bagnati Marco , Albano Emanuele
Published: July 2004

Short periods of ischemia followed up by reperfusion are known to protect the heart against injury caused by a subsequent sustained ischemia. This phenomenon, known as ischemic preconditioning, has also been recently shown to reduce ischemic liver damage, but the mechanisms involved are still unknown. By using isolated hepatocytes as an in vitro model of liver preconditioning, we have investigated the possible effect of preconditioning on intracellular pH and Na+ homeostasis. Freshly isolated rat hepatocytes were preconditioned by 10 minutes of incubation under hypoxic conditions followed up by 10 minutes of reoxygenation and subsequently exposed to 90 minutes of hypoxia. Although preconditioning did not ameliorate adenosine triphosphate (ATP) depletion, preconditioned hepatocytes exhibited an increased resistance to cell killing during hypoxic incubation. Intracellular acidosis and Na+ accumulation developing during hypoxia were appreciably reduced in preconditioned cells. The effects of preconditioning on intracellular pH, Na+ homeostasis, and citotoxicity were mimicked by stimulating protein kinase C (PKC) with 4β-phorbol-12-myristate-13-acetate (PMA) or 1,2 dioctanoyl-glycerol (1,2 DOG). Conversely, inhibiting PKC with chelerythrine or blocking vacuolar proton ATPase (V-ATPase) with bafilomycin A1 abolished the protection given by preconditioning or by PMA treatment on hypoxic acidosis, Na+ overload, and hepatocyte killing. Similarly, the addition of Na+ ionophore monensin also reverted the cytoprotection exerted by preconditioning. This indicated that ischemic preconditioning of isolated hepatocytes decreased cell killing during hypoxia by preventing intracellular Na+ accumulation. We propose that, after preconditioning, the stimulation of PKC might activate proton extrusion through V-ATPase, thus, limiting intracellular acidosis and Na+ overload promoted by Na+-dependent acid buffering systems.
A long-term hepatitis B viremia model generated by transplanting nontumorigenic immortalized human hepatocytes in Rag-2-deficient mice
Authors: Brown Jennifer J. , Parashar Bhupesh , Moshage Han , Tanaka Kathryn E. , Engelhardt Dean , Rabbani Elazar , Roy-Chowdhury Namita , Roy-Chowdhury Jayanta
Published: July 2004

Development of new therapies for human hepatitis B virus infection (HBV) would be greatly facilitated by the availability of a suitable small-animal model for HBV virus productionin vivo. To develop a murine model for HBV production, we established an immortalized, cloned liver cell line by transferring the Simian Virus 40 Large T-Antigen into primary human hepatocytes. These cells were stably transfected with a full-length HBV genome to generate a clone that expresses HBV genes and replicates HBV. The HBV-producing cells were transplanted into the livers of mice with combined immunodeficiency (Rag-2 deficient) by intrasplenic injection. Survival of the engrafted human hepatocytes was shown in several ways: fluorescentin situhybridization (FISH) with a human-chromosome-specific DNA probe (human alpha satellite), dot-blot hybridization of the genomic DNA extracted from liver biopsy specimens with a human-specific Alu repetitive DNA probe, Blur-8, as well as with an HBV DNA probe, and secretion of human proteins into plasma. Histological examination of mouse liver up to 8 months following human cell transplant shows completely normal architecture. Determination of plasma HBV DNA levels indicated that engrafted cells secreted 3×107to 3×108virions per mL into the blood, and HBsAg was detected in plasma. This new murine model of HBV viremia should be useful forin vivoHBV studies.
Gastric mucosa as an additional extrahepatic localization of hepatitis C virus: Viral detection in gastric low-grade lymphoma associated with autoimmune disease and in chronic gastritis
Authors: De Vita Salvatore , De Re Valli , Sansonno Domenico , Sorrentino Dario , Corte Renato La , Pivetta Barbara , Gasparotto Daniela , Racanelli Vito , Marzotto Alessandra , Labombarda Antonella , Gloghini Annunziata , Ferraccioli Gianfranco , Monteverde Angelo , Carbone Antonino , Dammacco Franco , Boiocchi Mauro
Published: July 2004

The hepatitis C virus (HCV) has been linked to B-cell lymphoproliferation and autoimmunity, and has been localized in several tissues. The clinical observation of an HCV-infected patient with Sjögren’s syndrome (SS) and Helicobacter pylori(HP) positive gastric low-grade B-cell non-Hodgkin’s lymphoma (NHL), which did not regress after HP eradication, led us to investigate the possible localization of HVC in the gastric microenvironment. HCV genome and antigens were searched in gastric biopsy specimens from the previously mentioned case, as well as from 9 additional HCV-infected patients (8 with chronic gastritis and 1 with gastric low-grade B-cell NHL). HCV-specific polymerase chain reaction (PCR) and immunohistochemistry procedures were used. The gastric B-cell NHL from the patient with SS was characterized by molecular analyses of B-cell clonality. HCV RNA was detected in both the gastric low-grade B-cell NHL and in 3 out of 6 gastric samples from the remaining cases. HCV antigens were detected in the residual glandular cells within the gastric B-cell NHL lesions, in glandular cells from 2 of the 3 additional gastric lesions that were HCV positive by PCR, and in 1 additional chronic gastritis sample in which HCV-RNA studies could not be performed. By molecular analyses, of immunoglobulin genes, the B-cell NHL from the patient with SS was confirmed to be a primary gastric lymphoma, subjected to ongoing antigenic stimulation and showing a significant similarity with rheumatoid factor (RF) and anti-HCV- antibody sequences. Our results show that HCV can localize in the gastric mucosa.
Effects of age and viral determinants on chronicity as an outcome of experimental woodchuck hepatitis virus infection
Authors: Cote Paul J. , Korba Brent E. , Miller Roger H. , Jacob James R. , Baldwin Betty H. , Hornbuckle William E. , Purcell Robert H. , Tennant Bud C. , Gerin John L.
Published: July 2004

Acute hepadnavirus infections either resolve or progress to chronicity. Factors that influence chronicity as an outcome of hepatitis B virus (HBV) infection in humans can be studied experimentally in the woodchuck model. Accordingly, several woodchuck hepatitis virus (WHV) inocula were characterized. Representative inocula had high titers of infectious virus (approximately 107.7-109.5 woodchuck 50% infectious doses per milliliter [WID50% /mL] by subcutaneous inoculation), with 1 WID50% ranging between 21 and 357 physical virion particles. WHV7P1 (standard high dose, 5 × 106WID50%) produced a 72% chronicity rate (i.e., percent chronic of total infected) in neonatal woodchucks (1-3 days old). Comparable doses of WHV8P1 resulted in a lower chronicity rate in neonates (34% chronic) indicating that it represented a strain different from WHV7P1. Neonatal woodchucks were more susceptible to chronic infection by high doses of WHV7P1 (range, 65%-75% chronic) compared with 8-week-old weanlings (33% chronic) and adult woodchucks (0% chronic; i.e., all resolved). High doses of cloned wild-type viruses also induced high rates of chronicity in neonates (70%-80% chronic). Chronicity rates in neonates were decreased for low doses of WHV7P1 (500 WID50%, 9% chronic) and for high doses of a precore WHeAg-minus mutant WHV8 clone (17% chronic). Thus, both age and viral determinants can influence chronicity as an outcome of experimental WHV infection. Standardized inocula will enable the study of mechanisms that initiate and maintain chronic hepadnavirus infection and also provide a means for developing WHV carriers for therapeutic studies.
Is an “à la carte” combination interferon alfa-2b plus ribavirin regimen possible for the first line treatment in patients with chronic hepatitis C?
Authors: Poynard Thierry , McHutchison John , Goodman Zachary , Ling Mei-Hsiu , Albrecht Janice
Published: July 2004

Randomized trials have shown the enhancement of efficacy with interferon alfa-2b and ribavirin (IFN-R) in comparison with interferon monotherapy (IFN) as first line treatment of chronic hepatitis C. Further definition of response based on disease, patient, and treatment characteristics is needed to determine the degree of benefit for the various patient subgroups. The aim of this study was to answer this question by analyzing the data from 1,744 naive patients included in trials that compared 24- or 48-week IFN-R treatment. Response factors were identified by logistic regression and receiver operating characteristics curves. Five independent characteristics were associated with a sustained loss of hepatitis C virus (HCV) RNA (<100 copies/mL) 24 weeks after the end of treatment: genotype 2 or 3, baseline viral load less than 3.5 million copies/mL, no or portal fibrosis, female gender, and age younger than 40 years. There was a significant advantage for IFN-R in comparison with IFN alone whatever the combination of factors. The most efficient strategy is to treat all patients for 24 weeks. If the 24-week polymerase chain reaction (PCR) is positive, treatment can be stopped. If the 24-week PCR is negative, patients with fewer than 4 favorable factors should be treated for an additional 24 weeks. Conclusion: The combination of IFN-R is better as first line treatment than IFN monotherapy. For patients who are PCR negative after 24 weeks of treatment, genotyping and baseline viral load, fibrosis stage, gender, and age are useful predictive factors in determining whether to continue an additional 24 weeks of treatment.
In vitro evaluation of hepatitis B virus polymerase mutations associated with famciclovir resistance
Authors: Xiong Xiaofeng , Yang Huiling , Westland Christopher E. , Zou Ruiming , Gibbs Craig S.
Published: July 2004

Several mutations (V521L, P525L, L528M, T532S, and V555I) in the gene for hepatitis B virus (HBV) polymerase have been identified in HBV isolated from patients that displayed break-through viremia during famciclovir treatment. To determine whether these mutations cause phenotypic resistance to famciclovir, we compared the inhibition constants (K
i) of penciclovir triphosphate (PCVTP, the active metabolite of famciclovir) for recombinant wild-type and mutant HBV polymerases containing these mutations. In in vitro enzymatic assays, the V555I mutation displayed the most resistance (with K
i increased by 6.2-fold) to PCVTP. The V521L and L528M mutations showed moderately decreased sensitivity to PCVTP (K
i increased by >3-fold). We also analyzed the cross-resistance profiles of these variants for adefovir and lamivudine, two other antiviral agents that also inhibit DNA replication by HBV polymerase. All 5 famciclovir-associated mutations were sensitive to adefovir diphosphate (ADVDP) in in vitro enzymatic assays (<2.3-fold decreased sensitivity). The V521L, L528M, and T532S mutations were also sensitive to lamivudine triphosphate (LAMTP); however, the P525L and V555I mutations displayed moderately decreased sensitivity to LAMTP in enzymatic assays (3.6-fold decreased sensitivity). The lamivudine-resistant mutations M552I, M552V, and L528M+M552V, which were previously shown to display 8- to 25-fold resistance to LAMTP, were less resistant (≤3.1-fold) to PCVTP.