bluuebunny/crossref_metadata_2025 huggingface dataset:
Iron overload facilitates hepatic fibrosis in the rat alcohol/low-dose carbon tetrachloride model
Authors: Mackinnon Malcolm , Clayton Cindy , Plummer John , Ahern Michael , Cmielewski Patricia , Ilsley Anthony , Hall Pauline
Published: March 2007

The role of iron deposition in initiating hepatic fibrosis in iron overload disorders is not clearly established, and it is becoming increasingly recognized that iron may be interacting with other potential liver-damaging agents. The authors therefore examined the interplay of iron and alcohol in rats administered subtoxic doses of carbon tetrachloride (CCl4) vapor at 20 ppm in customized chambers. At birth, the offspring of seven pregnant Porton rats were divided into two groups: one group was fed a normal rat chow diet and the other a diet supplemented with 3% (w/w) carbonyl iron for 10 weeks after weaning. In this latter group, the mothers were fed an iron supplement while breastfeeding. At 10 weeks, the animals from the first group (normal chow) were divided into two groups of six animals and fed a Lieber-DeCarli liquid diet with daily exposure to CCl4 vapor: group 1, liquid diet + CCl;group 2, liquid diet + alcohol 150 kcal/l + CCl4. The animals from the second iron-supplemented group were divided into two groups of six animals and fed a liquid diet with 3% (w/v) carbonyl iron and exposed to CCl4 vapor for 10 weeks: group 3, liquid diet + iron + CCl4;group 4, liquid diet + iron + alcohol supplement + CCl4. Two animals from each group of six had a liver biopsy at 4, 6, and 8 weeks, and all animals were killed after 10 weeks of CCl4 exposure. After the first 10-week iron loading period, the rats fed the carbonyl iron-supplemented diet had a 10-fold elevation in hepatic iron concentration. In the second 10-week (CCl4 exposure) period, fibrosis was scored on a four-point scale in each liver biopsy and in all animals at 10 weeks. At 10 weeks, the animals exposed to iron and alcohol in addition to CCl4 all had an established or developing cirrhosis with the development of fibrosis apparent at 4 weeks. Animals in the other groups had markedly less fibrosis, with none seen in the control group up to 10 weeks. Thus, the addition of iron to alcohol facilitates the development of fibrosis in animals exposed to subtoxic doses of CCl4 vapor. This model should allow a more detailed analysis of the mechanism(s) underlying this process. (Hepatology 1995; 21:1083-1088.)
Ethane exhalation and vitamin E/ubiquinol status as markers of lipid peroxidation in ferrocene iron—loaded rats
Authors: Dresow Bernd , Albert Claudia , Zimmermann Inge , Nielsen Peter
Published: March 2007

Organ damage caused by iron overload has been mostly attributed to iron-induced peroxidation of membrane lipids. Using the ferrocene iron—loaded rat model, we studied ethane exhalation as a direct marker of in vivo lipid peroxidation, as well as concentrations of α-tocopherol and ubiquinol 9/10 in liver and plasma as indirect markers of this process. The feeding of a diet enriched with 0.5% TMH-ferrocene up to 31 weeks resulted in a large increase in liver iron concentration to about 25 mg/g wet weight (w wt). At lower, predominantly hepatocellular liver siderosis, the breath ethane exhalation was dependent on dietary vitamin E (VitE) supplements (onset of ethane exhalation at liver-Fe >2 mg/g w wt on VitE—restricted diet; >5 mg Fe per gram on VitE—replete diet). At severe liver siderosis, breath ethane exhalation reached a maximum of approximately 8 nmol/kg/hr independent of VitE supplementation. Plasma as well as hepatic α-tocopherol decreased with progressive iron loading. In addition, a significant depletion in hepatic ubiquinol 9 and 10 was noted.
The structure and organization of the bile canalicular cytoskeleton with special reference to actin and actin-binding proteins
Authors: Tsukada Nobuhiro , Ackerley Cameron A. , Phillips James M.
Published: March 2007

The distribution of actin filaments and actin-binding proteins in the bile canaliculus (BC) of normal human hepatocytes was determined as a means of establishing the structure and organization of the BC cytoskeleton. Immunoblots demonstrated that actin, and the actin-binding proteins, myosin II, tropomyosin, vinculin, α-actinin, villin, were present, as were the non-actin-related proteins β-tubulin, and cytokeratins. Three actin filament regions were identified: microvillus core filaments, a membrane-associated microfilamentous network, and a circumferential pericanalicular actin filament band. Actin-binding proteins were nonrandomly associated with actin in these regions. In the case of the pericanalicular band, there was also association with the zonula adherens junction. Intermediate filaments inserted into desmosomes. The ultrastructural localization of the actin-binding proteins was fundamentally linked to the arrangement and organization of the major canaliculus-associated microfilament structures. Structural organization of the cytoskeleton was also linked to distinct components of the intercellular junctions. It is notable that tropomyosin and a-actinin, which in muscle cells are regulatory proteins of contractile activity, and myosin II are associated with the pericanalicular actin microfilament band; it is the BC counterpart of the contractile actin filament band found in the apical region of other secretory cells. The outer sheath of noncontractile intermediate filaments likely stabilizes the canalicular compartment. (Hepatology 1995; 21:1106-1113.)
In vitro tumor necrosis factor cytotoxicity in HEP G2 liver cells
Authors: Hill Daniell B. , Schmidt Jack , Shedlofsky Steven I. , Cohen Donald A. , McClain Craig J.
Published: March 2007

Tumor necrosis factor-α(TNF-α) is a mediator of liver injury. The objective of this study was to develop an in vitro model of TNF-mediated liver cell injury using the Hep G2 cell line. Hep G2 cells normally are insensitive to TNF cytotoxicity, but they were rendered susceptible, or sensitized, to TNF cytotoxicity by inhibitors of RNA and protein synthesis. The concentration of TNF required to kill 50% of Hep G2 cells sensitized with 0.8 μmol/L actinomycin D (Act D) was 35 pmol/L compared with 5 pmol/L for LM fibroblasts, a classic target cell used in TNF cytotoxicity bioassays. Similarly, TNF cytotoxicity occurred in Hep G2 cells sensitized with cyclohexmide (CHX), and cytotoxicity to both inhibitors was dose dependent. Both protein and RNA synthesis were inhibited in Hep G2 cells by the concentrations of CHX and Act D associated with TNF cytotoxicity. Hep G2 cells pretreated with TNF alone and later exposed to normally toxic concentrations of TNF with DACT did not develop cytotoxicity. Thus, in vitro tolerance to TNF was induced. Cytotoxicity also was more severe at modestly increased temperatures (39° C versus 37° C), which may have clinical relevance to hepatic decompensation during febrile episodes. We suggest that the Hep G2 cell line sensitized by inhibiting RNA and protein synthesis is a useful in vitro model for evaluating mechanism(s) of TNF-mediated liver cell injury. (Hepatology 1995; 21:1114-1119.)
Functional and ultrastructural features of ethanol/bile salts interaction in the isolated perfused rat liver
Authors: Alvaro Domenico , Benedetti Antonio , Gigliozzi Alessandro , Bini Adriano , Guardia Paola Della , la Rosa Tiziana , Jezequel Anne Marie , Capocaccia Livio
Published: March 2007

We investigated whether bile salts (BS) with different hydrophobic-hydrophilic properties interact with ethanol on bile secretion, enzyme (aspartate transaminase [AST], lactate dehydrogenase [LDH]) release in the perfusate, liver ultrastructure, and vesicular exocytosis in the isolated perfused rat liver. Ethanol (0.1 or 1%) promoted a rapid decrease of bile flow and BS secretion in livers perfused with taurocholate (TCA), the physiologic BS in the rat (−28% decrease of baseline values with 0.1% and −34% with 1% ethanol). The inhibitory effect of ethanol on bile flow and BS secretion was significantly (P < .02) attenuated by perfusing liver with the hydrophilic BS, tauroursodeoxycholate (TUDCA), and it was exacerbated (P < .02) by perfusion with the hydrophobic BS, taurodeoxycholate (TDCA). The release of AST and LDH in the perfusate was unaffected by 0.1% ethanol, but increased threefold to fivefold by 1% ethanol in TCA-perfused livers. This cytolitic effect of ethanol was not observed in TUDCA-perfused livers, but it was enhanced (P < .03) by perfusion with TDCA. No ultrastructural abnormalities were found in either TCA- or TUDCA-perfused livers, with or without 1% ethanol. Only minimal changes were found in livers perfused with TDCA alone, but, in the presence of TDCA, 1% ethanol induces marked mitochondrial damage. The biliary excretion of the fluid phase marker horseradish peroxidase was inhibited by ethanol, an effect reversed by TUDCA (P < .02) and exacerbated by TDCA (P < .04). In conclusion, this study demonstrates that hydrophilic BS such as TUDCA counteract the inhibitory effect of ethanol on bile secretion and vesicular exocytosis as well as the ethanol-induced cytolitic effect in the isolated perfused rat liver. In the presence of hydrophobic BS such as TDCA, the exposure to ethanol promotes a marked inhibition of bile secretion and vesicular exocytosis as well as prominent mitochondrial damage. (Hepatology 1995; 21:1120-1129.)
Evidence of preservation injury to bile ducts by bile salts in the pig and its prevention by infusions of hydrophilic bile salts
Authors: Hertl Martin , Harvey Robert P. C. , Swanson Paul E. , West Delin D. , Howard Todd K. , Shenoy Surendra , Strasberg Steven M.
Published: March 2007

Preservation injury to bile ducts is a serious problem in liver transplantation, especially when preservation exceeds 12 hours. The authors hypothesized that the injury was caused by contact of bile ducts with bile salts during cold preservation and might be preventable by infusion of more hydrophilic bile salts. Swine livers were harvested after intraportal infusions of saline (control), of the hydrophobic bile salt taurodeoxycholate, or of the hydrophilic bile salts tauroursodeoxycholate or dehydrocholate. The effect of infusing a combination of hydrophilic and hydrophobic bile acids was also studied. Bile samples were taken before and during the infusions. Then livers were perfused with UW solution, ducts were flushed retrograde with UW, and livers were stored at 0 to 1°C for 20 hours. Bile ducts were harvested after preservation, and coded microscopic slides of the specimens were examined by light microscopy. There was large variability in baseline bile salt concentration. Injury after preservation consisted of sloughing and pyknosis of surface and glandular epithelium. The histologic injury score determined after preservation was directly related to bile salt concentration in bile ducts at the time of flushing. During bile salt infusions, the infused bile salt replaced most or all of the other bile salts present in bile. Severe postpreservation injury of intrahepatic ducts occurred after taurodeoxycholate infusions, but injury was minimal when either of the two hydrophilic bile salts was infused. The mixture of bile acids produced intermediate results. Retrograde flushing with UW does not prevent injury to intrahepatic ducts. The authors conclude that the injury is caused by contact with bile salts, is dependent on bile salt concentration and composition, and is preventable. (Hepatology 1995; 21:1130-1137.)
Response of plasma and tissue endothelin-1 to liver ischemia and its implication in ischemia-reperfusion injury
Authors: Kawamura Eisuke , Yamanaka Naoki , Okamoto Eizo , Tomoda Fumihito , Furukawa Kazutaka
Published: March 2007

This study was designed to investigate the changes in plasma and tissue endothelin-1/endothelin-2 (ET) after liver ischemia and to assess the protective effect of anti-ET 1/ET 2 monoclonal antibody (ET antibody) against ischemia-reperfusion injury. The ET levels in the liver tissue, hepatic venous blood of the ischemic and non-ischemic sides, and in the portal venous blood were measured before and after partial liver ischemia for 1 hour in the adult dog. The ET levels in the liver tissue and hepatic venous blood on the ischemic side increased slightly during ischemia and markedly after reperfusion, whereas those on the nonischemic side showed no significant increases. The ET levels in the portal venous blood peaked at 1 to 3 hours after ischemia, which was significantly higher than the levels in the hepatic venous blood on the ischemic side and which correlated with the portal venous pressure elevated because of the partial liver clamping. The administration of antibody (2 mg/kg, intravenous) before reperfusion resulted in a significant inhibition of the postreperfusion elevations of serum-glutamic-oxaloacetic transaminase (GOT), S-glutamic pyruvic transaminase (GPT), and the indocyanine green (ICG) dye retention rate. In conclusion, ET was produced both in the liver tissue exposed to ischemia and in the vascular endothelium of the portal bed exposed to portal congestion. The ET released from the vascular endothelium, including the liver and the portal bed, was found to be a possible factor of ischemia-reperfusion injury.
Proliferation and hypertrophy of liver cells surrounding islet grafts in diabetic recipient rats
Authors: De Paepe Monique E. , Keymeulen Bart , Pipeleers Daniel , Klöppel Günter
Published: March 2007

The liver offers an adequate site for the metabolic function of pancreatic islet implants. Little is known about the effects of the islet grafts on the host organ. This study examines liver tissue of normal or streptozotocin (STZ)-diabetic rats at different intervals following intraportal injection of syngeneic islets. Implantation of 800-islet—grafts, containing 0.9 million beta cells, normalized overt diabetes within 14 days. This period of metabolic normalization was characterized by a specific sequence of alterations in the implant area. During the first days after transplantation, islet cells migrated into the liver lobules, whereby tight hepatocyte-islet cell contacts were established. Hepatocytes surrounding grafts showed massive lipid accumulation and hypertrophy (cellular profile area 603 ± 72 μm2 in diabetic islet recipients vs. 382 ± 42 μm2 in diabetic controls; P < .005). The implant area also contained significantly more liver cells in proliferative activity than hepatic tissue in normal controls (bromodeoxyuridine labeling index of peri-islet hepatocytes 6.2%, 4.6%, and 0.9% on posttransplantation days 2, 4, and 14, respectively, compared with 0.02% in normal controls). The cellular hypertrophy and hyperplasia explain the sudden increase in liver weight of diabetic recipients (from 8.0 ± 1.1 g to 13.8 ± 2.2 g on post-transplantation day 2; P < .005). Both alterations can be attributed to the massive local discharge of insulin in an insulin-deficient organ containing an excess of extracellular nutrients. Progressive revascularization of the implant sites and overall metabolic normalization are thought to explain the return of a normal liver histology by the third week after transplantation. In conclusion, intraportal islet grafts exert profound effects on the liver of diabetic rat recipients. The morphological features of the implant sites may serve as markers for the function of the islet grafts as well as for the adaptive capacity of the recipient liver. (Hepatology 1995; 21:1144-1153.)
Chronic ethanol feeding increases the quantity of Gαs-protein in rat liver plasma membranes
Authors: Iles Karen E. , Nagy Laura E.
Published: March 2007

The liver is a primary target for both acute and chronic effects of ethanol. Because ethanol is known to alter the function of guanine nucleotide regulatory proteins (G-proteins), changes in hepatic G-proteins could contribute to the adverse effects of ethanol on liver function. Male Wistar rats were fed a liquid diet containing 36% of calories as ethanol for 4 weeks. Control rats were pair-fed or allowed free access to a diet that isocalorically substituted maltose dextrins for ethanol. Liver plasma membranes were isolated and separated into basolateral and canalicular fractions by sucrose-density gradients. Enrichment of marker enzymes (5′-nucleoti-dase for canalicular membranes and forskolin-stimulated adenylyl cyclase activity for basolateral membranes) was not affected by ethanol feeding. Quantity of Gαs, and Gαi proteins in membrane fractions was determined by immunoblot. After ethanol feeding, immunoreactive Gαs protein was increased in liver plasma membranes compared with pair-fed controls. Gαi and Gαs were present in both the basolateral and canalicular fractions of the plasma membrane in control and ethanol-fed rats. Gαs quantity in the basolateral membrane was greater in ethanol-fed rats compared with controls, with no differences in Gαs observed in canalicular membranes. The quantity of Gαi did not change in response to ethanol feeding in any of the membrane fractions. Treatment of isolated plasma and basolateral membranes with 10 μmol/L 5′-guanylimidophosphate, a non-hydrolyzable guanosine triphosphate analogue that activates G-proteins, increased cAMP production to a greater extent in ethanol-fed rats compared with controls. These data indicate that ethanol increases the quantity and function of Gαs protein in rat liver plasma membranes. (Hepatology 1995; 21:1154-1160.)
Cyclosporin A and FK-506 in inhibition of rat ito cell activation in vitro
Authors: Ikeda Hitoshi , Fujiwara Kenji
Published: March 2007

Ito cells are the primary matrix-producing cells in the liver. In hepatic fibrosis in vivo or culture on plastic, these cells undergo activation, a process characterized by cell proliferation, fibrogenesis, and smooth muscle α-actin expression. The cytosolic-binding proteins of cyclosporin A (CsA) and FK506 accelerate folding of various proteins including collagen and become inactivated by binding to those agents. CsA is shown to inhibit collagen synthesis in cultured fibroblasts. These findings prompted us to examine the effect of cyclosporin A and FK506 on Ito cell activation. CsA and FK506 reduced DNA synthesis in a dose-related manner, to 26% and 45% of controls at 5 μmol/L, respectively, without affecting total protein synthesis. CsA reduced collagen synthesis in a dose-related manner, to 70% of controls at 5 μmol/L without affecting noncollagenous protein synthesis, whereas FK506 changed neither collagen synthesis nor noncollagenous protein synthesis. Moreover, smooth muscle α-actin expression was reduced by 0.5 μmol/L CsA, but not by FK506. CsA merits consideration for the therapy of hepatic fibrosis. FK506 may also be a candidate for such therapy through inhibitory action on Ito cell proliferation. (Hepatology 1995; 21:1161-1166.)
Small intestinal absorption of polyethylene glycol 400 to 1,000 in the portacaval shunted rat
Authors: Pantzar Niclas , Bergqvist Peter B. F. , Bugge Mogens , Olaison Gunnar , Lundin Stefan , Jeppsson Bengt , Weström Björn , Bengtsson Finn
Published: March 2007

Functional changes of the intestinal barrier that may occur after the creation of a portacaval shunt (PCS) were investigated. After chronic PCS in the rat, the intestinal absorption of and the jejunal permeability to the inert polymer marker polyethylene glycol (PEG) with molecular weight (Mw) ranging from 400 to 1,000 g/mol were investigated. The PEG mixture was orally fed to PCS and sham-operated rats, and urine was collected for 24 hours to obtain the urinary recovery of the different PEG polymers as a measure of intestinal absorption. To study the intestinal permeability, segments from the proximal small intestine were incubated in diffusion chambers with the PEG mixture on the mucosal side, and samples were withdrawn from the serosal side for analysis. The urinary recovery for the PEGs increased (P < .01) while the tissue permeability decreased (P < .001) in the PCS group rats in comparison with Sham-operated rats. The increased absorption in vivo was caused neither by altered renal clearance, nor by changed portal blood pressure. The decreased jejunal permeability in the PCS rats could be explained by a reduction of the mucosal area by shortening of the microvilli. This discrepancy indicates that changes in permeability and absorption may not be parallel during PCS. It is possible that these changes also may be affected by nutritional factors, drug therapy, as well as toxic substances. (Hepatology 1995; 21:1167-1173.)
Genetic hemochromatosis and Wilson'S disease: Role for oxidant stress?
Authors: Britton Robert S. , Brown Kyle E.
Published: March 2007

Hereditary haemochromatosis is characterised by iron overload that may lead to tissue damage. Free iron is a potent promoter of hydroxyl radical formation that can cause increased lipid peroxidation and depletion of chain-breaking antioxidants. We have therefore assessed lipid peroxidation and antioxidant status in 15 subjects with hereditary haemochromatosis and age/sex matched controls. Subjects with haemochromatosis had increased serum iron (24.8 (19.1-30.5) vs. 17.8 (16.1-19.5) μmol/L, P = 0.021) and % saturation (51.8 (42.0-61.6) vs. 38.1 (32.8-44.0), P = 0.025). Thiobarbituric acid reactive substances (TBARS), a marker of lipid peroxidation, were increased in haemochromatosis (0.59 (0.48-0.70) vs. 0.46 (0.21-0.71) μmol/l, P = 0.045), and there were decreased levels of the chain-breaking antioxidants alpha-tocopherol (5.91 (5.17-6.60) vs. 7.24 (6.49-7.80) μmol/mmol cholesterol, P = 0.001), ascorbate (51.3 (33.7-69.0) vs. 89.1 (65.3-112.9), P = 0.013), and retinol (1.78 (1.46-2.10) vs. 2.46 (2.22-2.70) μmol/l, P - 0.001). Patients with hereditary haemochromatosis have reduced levels of antioxidant vitamins, and nutritional antioxidant supplementation may represent a novel approach to preventing tissue damage. However, the use of vitamin C may be deleterious in this setting as ascorbate can have prooxidant effects in the presence of iron overload.

The RRR-alpha-tocopherol (vitamin E) content in plasma from 46 patients with liver diseases and 23 healthy controls was determined by high performance liquid chromatography and electrochemical detection. Patients were divided into three groups: alcoholic liver diseases (n = 17; group A), hemochromatosis (n 17;group B) and Wilson's disease (n 12;group C). Lipidstandardized alpha-tocopherol levels were determined to neutralize differences due to hyperlipemia. The ratio of serum vitamin E to serum lipids (cholesterol, triglycerides, phospholipids) was highest in healthy controls and in patients in group A with cirrhosis and normal transaminases and bilirubin. Patients in group A with acute or chronic ethanol intoxication and high bilirubin levels had a 37% lower lipid-standardized vitamin E level than controls. Patients in group B with hemochromatosis, showing high serum iron (>180 μg/dl), a low free iron binding capacity (<8 μmol/l) and high ferritin-levels (>450 μg/l), had a 34% lower vitamin E/lipid ratio than healthy controls. No significant lowering of the vitamin E/lipid ratio was observed in the other patients in group B. A significant decrease (37%) in the vitamin E/lipid ratio was only detectable in patients with Wilson's disease (group C) showing high free serum copper (>10 μg/dl). The data support a role for free radicals in the pathogenesis of active liver diseases. © Journal of Hepatology.
Parenchymal FE and collagen gene expression: An iron-clad association?
Authors: Friedman Scott L.
Published: March 2007

Background/Aims:

Liver fibrosis and cirrhosis represent common pathological findings in humans with iron overload. This study was undertaken to assess whether in vivo targeting of iron to liver parenchymal or nonparenchymal cells would differently affect collagen gene activity.

Methods:

Rats were treated with an iron diet or intramuscular injections of iron dextran, and in situhybridization analyses on liver samples were performed.

Results:

These iron treatments determined parenchymal or reticuloendothelial cell iron overload, respectively. The typical distribution of iron into different liver cells was documented by histochemistry and confirmed by in situ hybridization analysis with a ferritin L complementary RNA probe. In iron-fed rats, in situ hybridization analysis identified a signal for collagen type I messenger RNA into nonparenchymal cells in zones I and II. In rats with nonparenchymal cell iron overload, no activation of collagen gene expression was detected into or near iron-laden nonparenchymal cells. These findings were also confirmed by quantitative Northern blot analysis.

Conclusions:

The results of this study indicate that, regardless of the total hepatic iron burden, selective localization of iron into liver cells (i.e., parenchymal cells) is required for the activation of collagen gene during longterm iron overload in rodents.
Permissiveness of kupffer cells for simian immunodeficiency virus (SIV) and morphological changes in the liver of rhesus monkeys at different periods of SIV infection
Authors: Persidsky Yury , Steffan Anne-Marie , Gendrault Jean-Louis , Hurtrel Bruno , Berger Stefan , Royer Cathy , Stutte Hans-Jochen , Muchmore Elizabeth , Aubertin Anne-Marie , Kirn André
Published: March 2007

The pathogenesis of liver injury, which remains unclear in the course of human immunodeficiency virus infection, can be investigated in simian immunodeficiency virus—infected macaques, which develop an immunodeficiency disease resembling human acquired immune deficiency syndrome (AIDS). We studied the livers of 21 monkeys infected with simian immunodeficiency virus (SIVmac251) for 4 days to 39 months and detected viral antigens in Kupffer cells, macrophages, and lymphocytes in 65% of the livers tested. Virus-containing cells were present in 5 out of 9 livers tested as early as 4 days postinoculation. The number of positive cells as well as their content in viral proteins substantially increased in sinusoidal cells with the progression of the disease. Morphological features and double immuno-labeling indicated that Kupffer cells constituted the predominant cell type containing viral antigens. The presence of multinucleated giant cells displaying the ultrastructural features of resident liver macrophages was another sign of the productive infection of Kupffer cells in vivo, which was attested by the observation of budding, immature, and mature SIV particles. Kupffer cell hyperplasia and hypertrophy were evident and appeared to be related to the development of SIV infection, because a close correlation was found between antigenemia and the surface area occupied by these cells. The Kupffer cells contained apoptotic lymphocytes, indicating that resident liver macrophages could play a role in the uptake of such cells from the blood. The production of tumor necrosis factor α (TNFα) and, possibly, interferon-α by Kupffer cells, the expression of vascular adhesion molecule-1, (VCAM-1), intralobular and periportal inflammation, and the proliferation and expansion of bile duct cells were other signs of liver involvement in SIV infection.
Hepatic and portal vein thrombosis in cirrhosis: Possible role in development of parenchymal extinction and portal hypertension
Authors: Wanless Ian R. , Wong Florence , Blendis Lawrence M. , Greig Paul , Heathcote Jenny E. , Levy Gary
Published: March 2007

Obliterative lesions in portal veins (PVs) and hepatic veins (HVs) of all sizes are known to occur in cirrhotic livers. PV lesions have generally been attributed to thrombosis, but the pathogenesis of the HV (veno-occlusive) lesions is unknown. We have studied 61 cirrhotic livers removed at transplantation to clarify the prevalence, distribution, and pathogenesis of venous lesions, as well as the association of these lesions with other morphological features and clinical morbidity. Intimal fibrosis that is highly suggestive of healed HV or PV thrombosis was found in at least 70% and 36% of livers, respectively. The distribution of HV lesions was patchy and largely confined to veins between 0.1 and 3 mm in diameter, suggesting multifocal origin in small veins. PV lesions were more uniform throughout the liver, suggesting origin in large veins with propagation to the small veins. HV lesions were associated with regions of confluent fibrosis (focal parenchymal extinction), and PV lesions were associated with regional variation in the size of cirrhotic nodules and a history of bleeding varices. These observations suggest that thrombosis of medium and large PVs and HVs is a frequent occurrence in cirrhosis, and that these events are important in causing progression of cirrhosis.
Effects of hepatocyte growth factor on the growth and metabolism of human hepatocytes in primary culture
Authors: Gómez-Lechón María José , Castelli José , Guillén Isabel , o'Connor Enrique , Nakamura Toshikazu , Fabra Ricardo , Trullenque Ramón
Published: March 2007

The effect of recombinant human hepatocyte growth factor (h-rHGF), a potent mitogen for hepatocytes, was investigated in primary cultures of human hepatocytes. Here, we describe a series of experiments to investigate the kinetics of its mitogenic action, as well as its metabolic effects on cultured human hepatocytes. The h-rHGF is a potent signal for initiating DNA synthesis in human hepatocytes, with maximal stimulatory effects at 10 ng/mL (0.1 pmol/L). The kinetics of DNA synthesis showed a lag of about 48 to 72 hours, followed by a maximum at 96 hours. At least 48 hours of continuous exposure to h-rHGF are required to initiate DNA synthesis in quiescent human hepatocytes. Cell cycle analysis by flow cytometry showed that most of quiescent 2c cells have left G0/G1 and entered the cell cycle (S and G2/M phases) by 96 hours of continuous exposure to h-rHGF. When compared with other growth factors, h-rHGF was a much more potent mitogen. The effects of 10 ng/mL (0.1 pmol/L) h-rHGF on DNA synthesis were only achieved by 1.5 pmol/L epidermal growth factor (EGF), 0.1 μmol/L insulin, or 1 μmol/L glucagon. It is noteworthy that the effect of h-rHGF was potentiated by glucagon but not by insulin or EGF. The stimulatory effect of HGF on DNA synthesis was gradually inhibited by h-rHGF transforming growth factor beta (TGF-β) in the range 1 to 10 ng/ml. The HGF also influenced the expression of other hepatic genes. This mitogenic factor stimulated the synthesis of the negative acute-phase plasma protein, albumin, and inhibited the synthesis of the positive one, α1-antichymotrypsin, whereas it moderately influenced the synthesis of fibrinogen. Although glucagon per se had no effects on cell growth, the synergism between glucagon and h-rHGF could be of relevance facilitating the mobilization of glycogen and the stimulation of human hepatocyte growth.
Serum cholestanol, cholesterol precursors, and plant sterols during placebo-controlled treatment of primary biliary cirrhosis with ursodeoxycholic acid or colchicine
Authors: Miettinend Tatu A. , Färkkilä Martti , Vuoristo Matti , Karvonen Anna-Liisa , Leino Rauli , Lehtola Juhani , Friman Claes , Seppälä Kari , Tuominen Juhani
Published: March 2007

A randomized placebo-controlled 2-year study was performed in 69 patients with primary biliary cirrhosis (PBC) on serum lipids during ursodeoxycholic acid (URSO) and colchicine treatments. In addition to serum bilirubin and alkaline phosphatase (AFOS), two variables considered to reflect liver function, serum lipoproteins, cholesterol precursors (squalene, Δ8-cholestenol, lathosterol and desmosterol), markers of cholesterol synthesis, cholestanol and plant sterols (campesterol and sitosterol), markers of liver function and cholesterol absorption, were studied before and during the treatments. Serum bilirubin was inconsistently improved by URSO, whereas improvement of AFOS values was better by URSO than colchicine, especially in patients with initially more advanced PBC. Serum total cholesterol was reduced by both drugs, very low—density lipoprotein (VLDL) and high-density lipoprotein (HDL) cholesterol by URSO. Cholesterol precursor sterols were increased by both URSO and colchicine mainly in patients with initially less severe PBC. On the other hand, the cholestanol values were markedly increased initially, and the values were related to bilirubin during the 2-year period, were further increased in the placebo group, and reduced in the URSO and colchicine groups, so that the improvement was highest in the URSO-treated patients with the severe form of PBC. The increase of the serum plant sterols, particularly that of sitosterol, was retarded by the two drugs so that the campesterol/sitosterol ratio, which was related to serum bilirubin, was increased especially in the cases with initially more advanced PBC. The present findings show that the serum cholestanol level and the campesterol/sitosterol ratio, two variables characterizing end-stage PBC, are modestly but significantly improved by long-term URSO or colchicine treatment, but virtually none of the excessively increased cholestanol values are normalized.
Cholesterol metabolism in liver and gallbladder mucosa of patients with cholesterolosis
Authors: Sahlin Staffan , Ståhlberg Dagny , Einarsson Kurt
Published: March 2007

The objective of this study was to investigate possible pathogenetic factors for cholesterolosis. Liver tissue, gallbladder mucosa, and gallbladder bile were collected in patients with cholesterol gallstones (GS) (14 patients with and 14 patients without cholesterolosis) and gallstone-free (GSF) subjects (11 with and 21 without cholesterolosis) undergoing cholecystectomy. In cholesterolosis, the gallbladder mucosa was characterized by a fivefold increase in esterified cholesterol and normal content of free cholesterol. The hepatic levels of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity, governing cholesterol synthesis, and acyl coenzyme A: acyltransferase activity, catalyzing the esterification of cholesterol, were similar in patients with and without cholesterolosis. Also in the gallbladder mucosa, the 3-hydroxy-3-methylglutaryl coenzyme A reductase activity was similar in patients with and without cholesterolosis. The acyl coenzyme A: acyltransferase activity of the gallbladder mucosa was increased in the GSF subjects with cholesterolosis. The nucleation time of gallbladder bile was shorter in the GSF subjects with cholesterolosis compared with the time of those without cholesterolosis. Occurrence of cholesterol crystals, lipid composition, and cholesterol saturation of gallbladder bile were not significantly influenced by the absence or presence of cholesterolosis. The study has confirmed that cholesterolosis is associated with a several-fold increased level of esterified cholesterol. The data suggest that patients with cholesterolosis have normal hepatic cholesterol formation and esterification. The local synthesis of cholesterol in the gallbladder mucosa seems to be normal. A positive correlation was obtained between the cholesterol saturation of bile and the content of esterified cholesterol in the gallbladder mucosa in the whole series of patients. However, because cholesterolosis may occur in both GSF subjects and patients with cholesterol GS with or without cholesterol-saturated bile, the uptake of cholesterol from the gallbladder bile may not be the only mechanism for the development of cholesterolosis.
Expression of CD44 in human hepatocellular carcinoma cell lines
Authors: Haramaki Makoto , Yano Hirohisa , Fukuda Kazunori , Momosaki Seiya , Ogasawara Sachiko , Kojiro Masamichi
Published: March 2007

CD44 is a glycosylated cell surface adhesion molecule expressed on a diverse range of cells and has several variant forms, some of which are involved in metastasis of cancer cells. Because little is known about CD44 in human hepatocellular carcinoma (HCC), we investigated its expression in tissue specimens from primary lesions (12 cases), in smear specimens from peritoneal effusions (2 cases), and in cell lines (HCC cell lines, KIM-1, KYN-1, KYN-2, KYN-3, HAK-1A, and HAK-1B; combined hepatocholangiocarcinoma cell lines, KMCH-1 and KMCH-2; and bile duct carcinoma cell lines, KMC-1 and KMBC). Immunohistochemical studies using monoclonal antibody recognizing epitope Group 1 of human CD44 molecule showed that HCC cells in all tissue specimens, including the original tumors of one smear specimen and HAK-1A, were negative for CD44; whereas, HCC cells in two-smear specimens, KIM-1, KYN-2, KYN-3, HAK-1A, HAK-1B, KMCH-1, KMC-1, and KMBC, showed positive reactions on the cell membrane. Immunostain-positive cell lines showed a positive cell rate of 51.9% to 99.8% by flow cytometric analysis. Western blotting detected CD44 protein of hemopoietic type in KIM-1, KYN-3, HAK-1A, and HAK-B and epithelial type in KMC-1 and KMBC. Southern blotting of complementary DNA amplified after reverse transcriptase—polymerase chain reaction (RT-PCR) detected hemopoietic type and some variant forms with longer insertion in all cell lines but KMCH-2, whereas hemopoietic type and variants with minor insertion were only detectable in tissue specimens. These findings suggest that HCC cells in ascites and in culture often express CD44, but those in tissue do not at protein level.
Thrombomodulin inhibits intrahepatic spread in human hepatocellular carcinoma
Authors: Suehiro Taketoshi , Shimada Mitsuo , Matsumata Takashi , Taketomi Akinobu , Yamamoto Kazuharu , Sugimachi Keizo
Published: March 2007

Thrombomodulin (TM) converts thrombin from procoagulant into anticoagulant protein to activate protein C. Thrombin also plays an important role in the metastatic process of cancer cells. We performed an immunohistochemical and clinicopathological study of TM in 141 cases with resected hepatocellular carcinoma (HCC) measuring less than 6 cm in diameter. Twenty-five specimens (17.73%) stained positive for TM. TM was found in the cytoplasm and surface of cancer cells. The clinicopathological findings according to the positive of TM are examined in HCC. The preoperative plasma TM level of the patients with tissue that stained positive for TM was significantly higher than that of the patients with negative results; for the postoperative TM level, there were no differences between them. In addition, the frequencies of intrahepatic metastasis, tumor thrombus in the portal vein, and capsular infiltration were significantly lower in patients whose tissue stained positive for TM than in patients whose tissue stained negative for TM. The recurrence freedom rate was significantly higher in patients whose tissue stained positive for TM than patients whose tissue stained negative for TM. Thus, TM-producing HCC shows a slow intrahepatic spread. Therefore, these findings suggest that TM may inhibit the adhesion of tumor cells to the portal vein because of anticoagulant activity and thus prevent the spread of intrahepatic metastasis.
Increase of deoxycholate in supersaturated bile of patients with cholesterol gallstone disease and its correlation with de novo syntheses of cholesterol and bile acids in liver, gallbladder emptying, and small intestinal transit
Authors: Shoda Junichi , He Bing-Fang , Tanaka Naomi , Matsuzaki Yasushi , Osuga Toshiaki , Yamamori Shunji , Miyazaki Hiroshi , Sjövall Jan
Published: March 2007

A total of 100 nonobese and normolipidemic subjects (29 control subjects, 49 patients with cholesterol stones [CSs], and 22 patients with brown pigment stones) were studied to elucidate the pathogenetic contributions of deoxycholate (DC) to supersaturated bile formation with special reference to de novo syntheses of cholesterol and bile acids in the liver. A higher proportion of DC was observed in gallbladder bile from patients with CSs (CSs; 21.7 ± 1.4%, mean ± SEM, vs. control subjects; 10.2 ± 0.9%). Cholesterol saturation in bile was elevated parallel to the increase of DC (r = 0.48; P = 0.0002), irrespective of the existence of stones. In a comparison between the 52 subjects with increased DC in bile ( > 10% of biliary bile acids) and the 20 subjects without the increase ( < 10%), the molar percentage of cholesterol in bile was significantly higher in the former (9.4 ± 0.5%) than in the latter (6.7 ± 0.4%) (P < 0.001). Consistent with the decrease in steady-state level of low-density lipoprotein (LDL) receptor—messenger RNA (mRNA), the catalytic activity and mRNA level of microsomal hepatic 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase, the rate-limiting enzyme for de novo cholesterol synthesis, were significantly lower in the former (2.9 ± 0.3 pmol/min/mg protein) than in the latter (5.1 ± 0.6) (P < 0.0001). Biliary molar percentage of bile acids was significantly lower in the former (69.8 ± 1.1%) than in the latter (75.2 ± 1.5%) (P < 0.01). However, contrary to expectations, the catalytic activity and mRNA level of cholesterol 7α-hydroxylase, the rate-limiting enzyme for bile acid synthesis, were significantly higher in the former (5.8 ± 0.4 pmol/min/mg protein) than in the latter (3.7 ± 0.6) (P < 0.01). The magnitude of the impaired gallbladder emptying (control subjects; 78.4 ± 4% vs. CSs; 58 ± 3%; P < 0.0005) together with the prolonged small intestinal transit (control subjects; 126 ± 9 minutes vs. CSs; 198 ± 9 minutes; P < 0.01) correlated significantly with the increased percentage of DC in bile. It is concluded that in cholesterol gallstone disease an increase of DC in bile, linked to an impaired gallbladder emptying together with a prolonged small intestinal transit, may play a significant role in downregulating de novo cholesterol synthesis but not bile acid synthesis in the liver. It is further suggested that the formation of supersaturated bile may be attributed to a hypersecretion of cholesterol (derived from an exogeneous source) driven by the flow of DC through the liver, combined with a normosecretion or hyposecretion of bile acids, the pool size of which is reduced because of changes in distribution, e.g., a sequestration into the hypomotile gallbladder and intestine.
Diagnostic value of fine-needle puncture of the gallbladder: Side effects, safety, and prognostic value
Authors: Tudyka Jürgen , Kratzer Wolfgang , Kuhn Klaus , Janowitz Paul , Wechsler Johannes Georg , Adler Guido
Published: March 2007

Bile sampling without the risk of contamination by pancreatic and duodenal secretions and avoiding unpredictable influences of general anesthesia during biliary surgery on biliary analytics are feasible with percutaneous puncture of the gallbladder. In 207 patients with gallstones, gallbladder puncture was performed under local anesthesia with a 22-gauge spinal needle under continuous real-time ultrasound guidance. Bile samples were investigated for biliary lipids and nucleation time. Complete aspiration of gallbladder bile could be achieved in all patients without complications such as bleeding, bile leak, or inflammation. Of these patients, 11.6% reported mild abdominal problems, 3.4% required analgetics, and in 1.0% biliary colics were observed. Elective cholecystectomy was performed in 1 patient. Of the bile samples, 10.1% were contaminated with bactobilia. Biliary lipids, cholesterol saturation index (CSI), total lipid concentration (TLC), and bacteriological contamination were independent of gallstone number, whereas patients with solitary gallbladder stones exhibited a significantly longer nucleation time (NT) in comparison with those with multiple stones. In patients with gallstones, fine-needle puncture of the gallbladder represents an important diagnostic procedure and can be performed within minutes without major side effects if performed by an experienced sonographer.
Kupffer cell iron overload induces intercellular adhesion molecule-1 expression on hepatocytes in genetic hemochromatosis
Authors: Stå Per , Broomé Ulrika , Scheynius Annika , Befrits Ragnar , Hultcrantz Rolf
Published: March 2007

The mechanisms underlying iron-induced liver fibrogenesis in patients with genetic hemochromatosis are poorly understood. We studied signs of Kupffer cell activation and inflammatory responses in liver biopsy specimens obtained from 15 patients with untreated and six patients with treated hemochromatosis. Immunohisto-chemistry was performed on 11 of the untreated and all treated patients. Three of the untreated patients (20%) had cirrhosis and eight (53%) had fibrosis. None had chronic active hepatitis (CAH). Immunohistochemistry indicated that 55% of the untreated patients had sparse intercellular adhesion molecule-1 (ICAM-1) expression by hepatocytes, and all of these had Kupffer cell iron overload. No ICAM-1 expression was seen by hepatocytes in treated patients or healthy controls. ICAM-1 was strongly expressed by hepatocytes from control patients with inflammatory liver disease. HLA-DR reactivity was seen on sinusoidal cells in all groups, but not on hepatocytes except for two of the control patients with CAH. Twenty-seven percent of the untreated hemochromatosis patients displayed moderate infiltration by CD3-positive lymphocytes. Electron microscopy of samples from untreated hemochromatosis patients showed hypertrophic Kupffer cells containing iron-rich remnants of phagocytosed hepatocytes. Fat-storing cells close to iron-laden hepatocytes contained multiple lipid droplets and adjacent collagen fibril bundles. Thus, in patients with untreated genetic hemochromatosis and Kupffer cell iron overload, hepatocytes occasionally express ICAM-1. In regions with heavy iron overload, Kupffer cell hypertrophy and transition of fat-storing cells are seen. Our findings indicate that release of factors from iron-loaded, activated Kupffer cells is of importance for the transformation of fat-storing cells and increased collagen deposition seen in genetic hemochromatosis.
Calculation of child and adult standard liver volume for liver transplantation
Authors: Urata Koichi , Kawasaki Seiji , Matsunami Hidetoshi , Hashikura Yasuhiko , Ikegami Toshihiko , Ishizone Shinpachi , Momose Yoshitaka , Komiyama Atsushi , Makuuchi Masatoshi
Published: March 2007

Despite refinements in surgical techniques for liver transplantation, liver size disparity remains one of the most common problems in pediatric patients. Optimal liver graft size remains unknown and the volume of diseased liver in the recipient is not indicative of the volume (standard liver volume [LV]) optimal for the recipient's metabolic demands. To establish a formula for calculating the standard LV in the pediatric and adult populations for liver transplantation, whole LVs were measured using computed tomography (CT) in 96 patients (65 pediatric and 31 adolescent or adult subjects) with normal liver whose disease conditions did not seem to affect body weight (BW) or LV. In the 96 subjects, the ratio of estimated LV to BW decreased gradually as age increased until approximately 16 years, when it started to level off. On the other hand, there seemed to be a directly proportional relationship between the estimated LV in vivo and body surface area (BSA) (r = 0.981; r
2 = 0.962; P < 0.0001) in the subjects as a whole, and the formula, LV (mL) = 706.2 × BSA (m2) + 2.4, was established from the measured data by simple regression analysis. Another predicting equation, LV (mL) = 2.223 × BW (kg)0.426 × body height (BH) (cm)0.682, was produced by multiple regression analysis (r
2 = 0.969; P < 0.0001). Considering its simplicity of use, we adopted the first formula for predicting standard LV in an individual patient.