bluuebunny/crossref_metadata_2025 huggingface dataset:
A Morphometric and Immunohistochemical Study on Angiogenesis of Human Metastatic Carcinomas of the Liver
Authors: Terayama N , Terada T , Nakanuma Y
Published: November 2004

Background/Aims:

To clarify the angiogenetic process and the origin of tumor vessels in human metastatic liver carcinomas.

Methods:

One hundred autopsy livers with metastatic carcinomas were studied by immunohistochemistry for von Willebrand factor, by lectin histochemistry for

Ulex europaeus

agglutinin I (UEA–I), and by morphometry of the density of tumor vessels in the liver metastases. In addition, tumor vessels were observed three–dimensionally in silicone rubber–injected livers with metastases.

Results:

Tumor vessels in metastatic liver carcinomas were positive for von Willebrand factor and UAE–I agglutinin I receptor. Tumor vessels first appeared in liver metastases of 200 μm in diameter. The density of the tumor vessels in the metastases increased up to 3mm in diameter, and it remained stable over 3mm. Sinusoidal endothelial cells around the liver metastases were positive for these endothelial markers. Sinusoidal endothelial cells were frequently in continuity with vessels in the metastases. Silicone rubber–injected specimens showed that blood vessels arising from surrounding sinusoids entered into the metastases.

Conclusions:

The angiogenesis of liver metastases may progress stepwise as the metastases enlarge, and capillarization of sinusoidal endothelium around the liver metastases may occur. Tumor vessels of liver metastases may in part originate from sinusoidal endothelial cells neighboring the tumors.
Role of Sinusoidal Endothelial Cells of the Liver in Concanavalin A–Induced Hepatic Injury in Mice
Authors: Knolle P A , Gerken G , Löser E , Dienes H P , Gantner F , Tiegs G , zum Büschenfelde Meyer K , Lohse A W
Published: November 2004

CD4+ T lymphocytes have been identified as being responsible for organ damage in the murine model of experimental liver injury induced by intravenous injection of concanavalin A (Con A). Liver sinusoidal endothelial cells (SEC) and Kupffer's cells (KC) are among the first cells that come into contact with lymphocytes in the liver sinusoid. We aimed to investigate the respective role of these cell populations in the initial steps of T–cell–mediated liver injury in Con A–induced hepatitis. By electron microscopy, we could show that intravenously applied Con A bound predominantly to SEC but not to KC. KC depletion by gadolinium chloride treatment of mice did not result in protection from liver injury, indicating that KCs are not primarily involved in the generation of liver injury. We could show that a CD4+ T–cell line (LNC.2) displayed selective cytotoxicity toward SEC (>50%) but not KC (12%) or fibroblasts (5%) in the presence of Con A

in vitro

. Microscopic observation revealed that the SEC monolayer was rapidly destroyed by LnC2 in the presence of Con A. Specificity of the Con A–induced cytotoxicity was shown by the ability of a competitive ligand, methyl–α–D–mannopyranoside, to reduce T–cell–mediated cytotoxicity to SEC by more than 50%. Tumor necrosis factor α (TNF–α) was produced by LnC2 in high amounts after Con A stimulation (>6 ng/mL), but antiserum to TNF–α did not reduce LnC2–mediated cytotoxicity toward SEC. In conclusion, we could show for the first time that liver SECs have accessory function and are selectively destroyed by CD4+ T lymphocytes in the presence of Con A. We speculate that SEC damage is an early event in T–cell–mediated liver injury recruiting T lymphocytes from the sinusoidal circulation. Loss of the SEC barrier function then exposes underlying hepatocytes to further attack by activated T lymphocytes. These results offer a model of initiating events in T–cell– mediated liver diseases, such as viral or autoimmune hepatitis, and suggest an important role for sinusoidal endothelial cells.
Alterations in Enzymatic Functions in Hepatocytes and Hepatocellular Carcinomas From Ras –Transduced Livers Resemble the Effects of Insulin
Authors: Pearline R V , Lin Y , Shen K J , Brunt E M , Bowling W M , Hafenrichter D G , Kennedy S , Flye M W , Ponder K P
Published: November 2004

An understanding of how oncogenes affect differentiated liver functions might lead to improved treatments for liver cancer or other disorders where liver–specific functions are compromised. A retroviral vector that coexpressed β–galactosidase (β–gal) and activated

Ras

genes (

Ras–gal

) was transduced into a small fraction of adult rat hepatocytes

in vivo

. Hepatocytes from

Ras–gal

–transduced diethylnitrosamine– untreated livers and hepatocellular carcinomas (HCC) from

Ras–gal

– transduced diethylnitrosamine–treated rats were analyzed for liver functions by performing histochemical assays on liver sections.

Ras–gal

– transduced hepatocytes failed to express gluconeogenic, ketogenic, and urea pathway enzymes. In contrast, several enzymes involved in fat synthesis were strongly activated, and microvesicular fat accumulated. These metabolic changes are induced in normal livers by insulin, a hormone that activates

p21–ras

. The deregulation of

p21–ras

may inhibit these liver–specific functions and may induce fat synthesis in both malignant and nonmalignant liver diseases. Furthermore, treatment with drugs that inhibit the attachment of

p21–ras

to the plasma membrane might reverse these changes. The alterations in enzymatic functions in the HCCs were similar to those observed in the hepatocytes, although each of the two cancers had a region that abruptly lost its expression of liver–specific enzymes and acquired the expression of genes that are more characteristic of oval or bile ductule cells. This suggests that a single genetic event in a malignant cell may dramatically alter its apparent phenotype. The identification of this putative gene might lead to insights into the regulation of the phenotype of normal cells in the liver.
Hepatic Sinusoidal Fibrosis Induced by Cholesterol and Stilbestrol in the Rabbit: 1. Morphology and Inhibition of Fibrogenesis by Dipyridamole
Authors: Wanless I R , Belgiorno J , Huet P
Published: November 2004

This study documents the hepatic morphology and the ultrastructure of a model of hepatic fibrosis in rabbits. Rabbits were given a cholesterol– supplemented diet (1%), a stilbestrol diet (10 mg subcutaneously twice a week), or both treatments simultaneously for 7 weeks. Rabbits given the combined treatment developed sinusoidal and portal fibrosis with only a mild disturbance of acinar vascular relationships. Ultrastructurally, there was marked widening of the spaces of Disse by collagen fibers, basement membrane material adjacent to endothelial cells and hepatocytes, blunted hepatocellular microvilli, activated stellate cells, lipid droplets in endothelial cells and hepatocytes, and degranulated platelets in sinusoids. The hepatic hydroxyproline content was markedly increased (12.0 ± 5.2 vs. 4.8 ± 1.5 mmol/g of liver dry weight;

P

< .001). Plasma bile acids were markedly increased (222 ± 180 vs. 12 ± 5 in controls;

P

< .001). Dipyridamole (25 mg every 12 hours) that was given in addition to cholesterol and stilbestrol decreased the hepatic collagen content (–49% and –48%, in two experiments;

P

< .05 in both) and splenomegaly. This model provides a reliable method for the production of extensive sinusoidal fibrosis with capillarization of sinusoids. Hepatocellular degeneration is only mild to moderate, and fibrosis occurs slowly without the sudden pathological changes that occur with other models of hepatic fibrosis, such as with the administration of CCl4 or galactosamine. The mechanism of injury may involve the accumulation of bile salts or the generation of free radicals from cholesterol oxidation products. The possibility that the sinusoidal release of platelet–derived factors may have a role in the activation of stellate cells (lipocytes) is supported by the suppression of fibrogenesis by dipyridamole.
Hepatic Sinusoidal Fibrosis Induced by Cholesterol and Stilbestrol in the Rabbit: 2. Hemodynamic and Drug Disposition Studies
Authors: Mastai R , Laganiere S , Wanless I R , Giroux L , Rocheleau B , Huet P
Published: November 2004

We assessed hepatic functions and systemic and splanchnic hemodynamics in a new model of hepatic sinusoidal fibrosis. Fibrosis was induced by the simultaneous administration for 8 weeks of diethyl–stilbestrol (DES) (10 mg twice weekly, subcutaneously) and cholesterol–supplemented diet (1%) in rabbits. A marked and progressive impairment of hepatic function was observed during the 8 weeks of treatment with a significant decrease in indocyanine green (ICG) systemic clearance (– 89%;

P

< .001) and aminopyrine elimination (–69%;

P

< .001). In fibrotic animals, hyperdynamic circulation was found with an increased cardiac output (+73%,

P

< 0.01) and a decreased peripheral vascular resistance (–50%;

P

< .005), as evaluated by the microsphere technique in animals that were awake. The total portal venous inflow was not significantly modified in fibrotic rabbits. However, since there was a marked increase in the liver weight, the portal venous inflow was significantly decreased when expressed per gram of liver weight (–30%;

P

< .05). In contrast, the hepatic artery blood flow was markedly increased, even when expressed per gram of liver weight (+95%;

P

< .01). Portal pressure was significantly increased in treated rabbits (from 7.4 ± .4 to 14.4 ± .6 mm Hg,

P

< .01). This new experimental model could prove useful to evaluate the influence of extensive perisinusoidal fibrosis on exchanges between plasma and hepatocytes, particularly of protein–bound substances.
Frequency and Significance of Antibodies to Actin in Type 1 Autoimmune Hepatitis
Authors: Czaja A J , Cassani F , Cataleta M , Valentini P , Bianchi F B
Published: November 2004

Antibodies to actin have been proposed as diagnostic markers for type 1 autoimmune hepatitis. Our aims were to determine (1) if testing for antibodies to actin is superior to testing for smooth muscle antibodies (SMA); (2) if these antibodies identify patients with distinctive clinical features; and (3) if the production of antibodies to actin is associated with genetic risk factors for autoimmune hepatitis. Sera from 99 patients with type 1 autoimmune hepatitis were tested. The frequencies of HLA B8, DR3, DR4, and A1–B8–DR3 in patient subsets were compared with those in 80 normal subjects. Seventy–three patients (74%) had antibodies to actin. Antibodies to actin were found more commonly in patients with SMA than in patients without them (86% vs. 7%,

P

< .0001). Screening only for antibodies to actin and antinuclear antibodies (ANA) failed to establish the diagnosis of autoimmune hepatitis in 5 patients. Patients with antibodies to actin were younger than seronegative patients. They were also more commonly DR3–positive than normal subjects and more frequently B8–positive than patients with non–actin–associated SMA (49% vs. 0%,

P

= .004). Only patients with antibodies to actin died of liver failure (6% vs. 0%), and 10 of 11 patients requiring liver transplantation were seropositive for these antibodies. Indeed, death and liver transplantation occurred more frequently in these patients than in actin–negative patients with ANA (19% vs. 0%, P = .03). We conclude that routine screening for antibodies to actin may miss patients with type 1 autoimmune hepatitis. Antibodies to actin are associated with HLA B8 and DR3, and they identify patients with a poor prognosis.
Immunohistochemical Evidence of Disease Recurrence After Liver Transplantation for Primary Biliary Cirrhosis
Authors: Van de Water J , Gerson L B , Ferrell L D , Lake J R , Coppel R L , Batts K P , Wiesner R H , Gershwin M E
Published: November 2004

Whether primary biliary cirrhosis (PBC) recurs after liver transplantation has remained an interesting and controversial issue; rejection, viral hepatitis, and drug effects all may mimic recurrent PBC histologically and biochemically. Furthermore, reliable clinical criteria for PBC recurrence are lacking. In this study, the issue of disease recurrence using a well–characterized monoclonal antibody (MAb), C355.1, that reacts with the immunodominant mitochondrial autoantigen of PBC (pyruvate dehydrogenase complex [PDC–E2]) was addressed. When used in an immunohistochemical assay, C355.1 produces intense apical staining of bile duct epithelium specifically in liver sections of patients with PBC and may be the earliest known marker of PBC. Immunohistochemical and histological analysis of serial liver biopsy specimens of 67 patients pre– and post–orthotopic liver transplantation (OLT), including 38 patients with PBC and 29 non–PBC liver disease controls, was performed. Sections were stained with MAb C355.1 or the control MAb C315 and analyzed to determine whether there was a recurrence of apical reactivity in the bile ducts of the posttransplantation biopsy specimens. The immunohistochemical staining was correlated with the histological findings and serum biochemistries at the time of the biopsy. Our data indicate that a significant number of patients who underwent transplantation for PBC (28 of 38) but not controls (0 of 29) develop a staining pattern of liver bile duct epithelium with MAb C355.1 that is indistinguishable from the pretransplantation pattern. Of the 28 patients with this apical staining pattern, 8 were characterized histologically as possible recurrent PBC, 2 as chronic rejection, 2 as acute rejection, 9 as nonspecific changes, 4 as normal or near normal, and 3 had other histological changes. Only 50% of the patients with apical C355.1 staining had liver enzyme levels suggestive of cholestasis. Thus, there appears to be immunohistochemical evidence that supports the concept of recurrence of PBC after OLT. The appearance of biliary epithelial abnormalities before the clinical appearance of disease is important not only for liver transplantation but also for understanding the natural history of PBC.
Intracaval Endovascular Ultrasonography for Preoperative Assessment of Retrohepatic Inferior Vena Cava Infiltration by Malignant Hepatic Tumors
Authors: Kaneko T , Nakao A , Nomoto S , Endo T , Itoh S , Takagi H
Published: November 2004

The accurate diagnosis of inferior vena cava infiltration is important not only for preoperative staging but also for performing surgery for hepatic tumor involving the inferior vena cava (IVC). We investigated the diagnostic value of intracaval endovascular ultrasonography (ICEUS) in the diagnosis of invasion of the IVC by a prospective study of 30 consecutive patients with hepatic tumors with possible invasion of the IVC. The procedure was performed using an 8F, 20–MHz intravascular ultrasonography (US) from the right femoral vein. Twenty–nine of the 30 patients underwent surgery, and the tumor was resected in 25 cases, including 3 cases of combined resection of the IVC. The diagnostic criterion for IVC infiltration with ICEUS was obliteration of a single echogenic layer of the IVC wall or intracaval tumor mass. The results of ICEUS were compared with those of computed tomography (CT), cavography, conventional US, and intraoperative US. Vascular invasion was identified in 7 of 30 cases and was subsequently confirmed by pathological examination of four resected specimens, including one autopsy and two operative findings. The sensitivity, specificity, and overall accuracy of ICEUS for the diagnosis of the IVC invasion was 100%, 95.8%, and 96.7%, respectively, against 85.7%, 60.8%, and 66.7% for CT, and 71.4%, 73.9%, and 73.3%, respectively, for cavography. The sensitivity, specificity, and overall accuracy of conventional US was 87.5%, 59%, and 66.7%, respectively; those of intraoperative US were 100%, 81.8%, and 86.2%, respectively. ICEUS is a useful technique that can precisely evaluate the IVC for possible hepatic tumor invasion.
Exogenous Spermine Induces Maturation of the Liver in Suckling Rats
Authors: Wéry I , Kaouass M , Deloyer P , Buts J , Barbason H , Dandrifosse G
Published: November 2004

In the present study, we investigated the effects of spermine on postnatal liver maturation in suckling rats. The animals were given spermine either

per os

(8 μmol) or by intraperitoneal injection (1 μmol), once daily for three or five days. The percentage of liver cells in different cell cycle phases and of diploid cells in the parenchyma was estimated. The protein content, ornithine aminotransferase (OAT) activity, and content of DNA polyamines and receptors for polymeric immunoglobulins (RPI) were also measured in liver extracts. The ingestion of spermine had the following effects: the percentage of the cells in S and G2M phases of the cell cycle diminished the percentage of diploid cells increased the content of polymeric immunoglobulin receptors increased; the OAT activity increased; the contents of putrescine and spermidine decreased and almost reached adult values; and the spermidine/spermine ratio became similar to that observed in the liver of adult rats. These phenomena were detected 40 hours after the beginning of oral spermine treatment. The intraperitoneal injection of spermine had no effect on the OAT activity, but it decreased the spermidine content and enhanced the spermine content. Our data demonstrated for the first time that dietary polyamines play a role in the initiation of liver postnatal maturation in suckling rats.
Apoptosis of Neutrophils and Their Elimination by Kupffer Cells in Rat Liver
Authors: Shi J , Fujieda H , Kokubo Y , Wake K
Published: November 2004

The fate of neutrophils in the peripheral circulation is poorly understood. In this study, the role of Kupffer cells in eliminating aged and apoptotic neutrophils was investigated. Liver, spleen, lung, and blood samples from Wistar rats were examined by light and electron microscopy, the terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate–biotin nick end labeling (TUNEL) method, and immunohistochemistry after the intravenous injection of OK–432, a streptococcal preparation. Neutrophils were trapped predominantly in the periportal and midzonal regions of hepatic lobules and were in contact with endothelial cells and Kupffer cells, or were surrounded by Kupffer cells. The trapping of neutrophils peaked after 6 hours. Apoptotic neutrophils, with or without buds, were found in the lumen of hepatic sinusoids as early as 6 hours, reached maximal levels after 12 hours, and represented greater than 60% of the total number of neutrophils in the liver. The presence of apoptotic neutrophils was correlated with the degree of neutrophil phagocytosis. Double–staining showed that TUNEL–positive neutrophils were phagocytosed or encircled by ED1– or ED2–positive Kupffer cells. In contrast, apoptosis and phagocytosis of neutrophils were rare in the spleen, lung, and peripheral blood. These results suggested that the appearance of apoptotic neutrophils in the hepatic sinusoids and their rapid clearance by Kupffer cells occurs after the invasion of bacteria (i.e., bacteremia or bacteriotoxemia) or the release of inflammatory mediators into the blood stream. These findings have important implications for the regulation of neutrophil homeostasis, the limitation of inflammation and tissue injury, and provide insight into the physiological removal of circulating, senescent neutrophils.
Hepatocyte Growth Factor Levels in Liver and Serum Increase During Chemical Hepatocarcinogenesis
Authors: Burr A W , Hillan K J , McLaughlin K E , Ferrier R , Chapman C , Mathew J , Burt A D
Published: November 2004

Hepatocyte growth factor (HGF) is mitogenic for hepatocytes and some tumor cell lines. Elevations in plasma HGF levels have been detected in patients with hepatocellular carcinoma (HCC), and it is possible that HGF is involved in the promotion and/or progression of tumor growth. We measured serum and liver tissue HGF levels during chemically induced hepatocarcinogenesis. Wistar rats were given diethylnitrosamine (DEN) in drinking water for 10 weeks with controls receiving drinking water only. Animals were killed at 10, 16, and 19 weeks. Liver HGF levels were determined from immunoblotted protein by scanning densitometry, and serum HGF levels were measured by sandwich enzyme–linked immunosorbent assay (ELISA). HGF was also immunolocalized in fixed liver tissue sections. In DEN–treated animals, at 10 weeks, there was necroinflammation but no dysplasia. Serum HGF was elevated compared with controls (

P

< .001) but there was no increase in liver HGF. At 16 weeks, there was liver cell dysplasia with minimal necroinflammation; serum and tissue HGF levels were both significantly elevated above controls. At 19 weeks, hepatocellular carcinomas (HCC) were present in five of six DEN–treated animals; liver HGF (

P

< .05) and serum HGF (

P

< .001) were both elevated compared with controls. HGF was localized in basement membranes around bile ducts and vessels and some perisinusoidal cells. Increased HGF immunolabeling was observed at 16 and 19 weeks, but dysplastic hepatocytes and tumor cells were HGF–negative. HGF may serve as a growth promoter at early stages during liver tumor development acting through possibly endocrine and paracrine pathways. Recent observations have described HGF as being mitoinhibitory for HCC cell lines; it is possible therefore that the continued up–regulation of HGF in the latter stages of our DEN model may inhibit tumor cell growth, and thus represent a form of antitumor host response.
Hypervariable Region Sequence in Cryoglobulin–Associated Hepatitis C Virus in Sera of Patients With Chronic Hepatitis C: Relationship to Antibody Response Against Hypervariable Region Genom
Authors: Aiyama T , Yoshioka K , Okumura A , Takayanagi M , Iwata K , Ishikawa T , Kakumu S
Published: November 2004

Essential mixed cryoglobulinemia is frequently associated with hepatitis C virus (HCV) infection, with the formation of HCV antigen/antibody complexes. The hypervariable region (HVR) of the HCV E2/NS1 region is thought to include epitopes for neutralizing antibodies, but it remains uncertain whether cryoglobulins (CGs) contain such antibody–bound HCV. Thus, we studied HVR clones isolated from cryoprecipitate and supernatant in the sera of four chronic hepatitis C patients with cryoglobulinemia, and expressed as fusion proteins with glutathione S–transferase (GST). Patients' sera were tested for antibody binding to the proteins. The rate of anti–HVR antibody–positive clones was significantly higher in cryoprecipitate (89% ± 13%,

P

< .05) than in supernatant (41% ± 25%). Both HCV RNA and anti–HVR antibody were more concentrated in cryoprecipitates compared with those of serum and supernatant in two patients tested. Anti–HVR antibody–positive clones in cryoprecipitate showed common amino acid (aa) sequences in each of the four patients. Similarly, all the antibody–positive clones in supernatant showed the same aa sequences for three of the four patients. When aa sequences were compared with those of reported isolates with genotype 1b, the mean percentage of aa difference was greater in the clones from supernatant and in anti–HVR antibody–negative clones than in the clones from cryoprecipitate and in the antibody–positive clones, respectively. These findings indicate that serum CG contains anti–HVR antibody–bound HCV in patients with chronic hepatitis C. Anti–HVR antibody–free individual clones, which were more frequently noted in supernatant, showed closely related sequences, but which were of a heterogeneous quasispecies nature.
The Valine Catabolic Pathway in Human Liver: Effect of Cirrhosis on Enzyme Activities
Authors: Taniguchi K , Nonami T , Nakao A , Harada A , Kurokawa T , Sugiyama S , Fujitsuka N , Shimomura Y , Hutson S M , Harris R A , Takagi H
Published: November 2004

The activities of key enzymes in the valine catabolic pathway-branched–chain aminotransferase, branched–chain alpha–keto acid dehydrogenase complex, methacrylyl (MC)–coenzyme A (CoA) hydratase (crotonase), and 3– hydroxyisobutyryl–CoA (HIB–CoA) hydrolase-were measured in normal and cirrhotic human livers. Unlike rat liver, which does not contain branched–chain aminotransferase, the aminotransferase activity in the normal liver was measurable and is increased somewhat in cirrhosis of the human liver. The total activity of branched–chain alpha–keto acid dehydrogenase complex in the normal human liver was approximately 1% of that in rat liver, and 20% to 30% of the complex was in the active form in both normal and cirrhotic livers. Only the actual activity of the enzyme was significantly decreased by cirrhosis. These results suggest that human liver is less active than rat liver in the catabolism of branched–chain amino and alpha–keto acids. Activities of MC–CoA hydratase and HIB–CoA hydrolase in human liver were very high compared with that of branched–chain alpha–keto acid dehydrogenase complex, suggesting an important role for these enzymes in catabolism of a potentially toxic compound, MC–CoA, formed as an intermediate in the catabolism of valine and isobutyrate. Cirrhosis resulted in a significant decrease in HIB–CoA hydrolase activity but had no effect on the citrate synthase activity, suggesting that the decrease in HIB–CoA hydrolase activity does not reflect a general decrease in mitochondria but that it may contribute to cellular damage that culminates in liver failure.
Differential Recognition of the 52–Kd Ro(Ss–A) Antigen by Sera From Patients With Primary Biliary Cirrhosis and Primary Sjogren's Syndrome
Authors: Dorner T , Feist E , Held C , Conrad K , Burmester G R , Hiepe F
Published: November 2004

Antibodies against the 52–kd Ro(SS–A) protein are significantly associated with the primary Sjogren's syndrome (pSS). A small proportion of patients suffering from primary biliary cirrhosis (PBC) with secondary Sjogren's syndrome (PBC/SS) who are serologically characterized by antimitochondrial type 2 antibodies also express anti–52–kd Ro(SS–A) antibodies. The primary B–cell–derived antigenic responses by autoimmune sera were analyzed in both entities using truncated recombinant proteins to examine whether different epitopes are associated with these diseases. Sera were collected from 25 patients with pSS and 9 anti–52–kd Ro(SS–A)–positive patients suffering from PBC/SS. B–cell epitope mapping was performed using different 52–kd Ro(SS–A) fusion proteins in enzyme–linked immunosorbent assay (ELISA) and immunoblotting. Sera from patients with pSS showed the broadest reactivity against antigenic epitopes at AA 153–245 compared to the significantly limited reactivity against AA 228–245 of sera from patients with PBC. B–cell epitopes within AA 190–245 represent immunodominant epitopes recognized by pSS sera in a significantly higher degree than by PBC sera, which react predominantly with AA 228– 245 (

P

< .0001). Anti–La(SS–B) antibodies were significantly associated with anti–52–kd Ro(SS–A) in sera from patients with pSS compared with PBC patients (

P

< .025). Thus, the antibody response to 52–kd Ro(SS–A) in PBC appears to be induced differently than in pSS. Although a limited immune response to 52–kd Ro(SS–A) occurs in PBC/SS patients, a more extended epitope spreading is evident in patients with pSS.
High Frequency of The Mage–1 Gene Expression in Hepatocellular Carcinoma
Authors: Yamashita N , Ishibashi H , Hayashida K , Kudo J , Takenaka K , Itoh K , Niho Y
Published: November 2004

The

MAGE–1

gene, originally isolated from a human melanoma cell line, directs the expression of a potential tumor–rejection antigen, MZ2–E. This antigen is recognized by autologous cytotoxic T lymphocytes in association with a major histocompatibility complex class I molecule (HLA–A1), and has provided a basis for specific immunotherapy for melanoma patients. Here we show a high frequency of expression of the

MAGE–1

gene in hepatocellular carcinoma (HCC). We examined the expression of the

MAGE–1

gene in cell lines originated from hepatoma cells and in tumor and nontumor tissues of livers with HCC by reverse–transcription polymerase chain reaction (RT–PCR) and subsequent Southern blotting.

MAGE–1

messenger RNA (mRNA) was expressed in three of four HCC cell lines (75%) and in 16 of 20 (80%) resected HCCs though none was detected in nontumor tissues. The high frequency expression of

MAGE–1

gene in HCCs suggests the possibility as a target for tumor– specific immunotherapy for HCC patients.
Fulminant Hepatic Failure in Rats: Survival and Effect on Blood Chemistry and Liver Regeneration
Authors: Eguchi S , Kamlot A , Ljubimova J , Hewitt W R , Lebow L T , Demetriou A A , Rozga J
Published: November 2004

A reproducible experimental animal model of fulminant hepatic failure (FHF) resembling the clinical condition is needed. We have developed such a model in the rat by combining resection of the two anterior liver lobes (68% liver mass) with ligation of the right lobes pedicle (24% liver mass), resulting in liver necrosis; the remaining two omental lobes (8% liver mass) are left intact. Adult Sprague–Dawley rats (250–300 g) were used. Survival time was determined in 60 rats. Because maintenance of body temperature at 37 degrees C shortened survival time by half, FHF rats were not warmed during the postinduction period and were allowed to gradually enter a state of mild to moderate hypothermia (29–32 degrees C). Additionally, 42 FHF rats were killed in batches of six rats each 2, 6, 12, 18, 24, 30, and 36 hours postoperatively to evaluate changes in blood chemistry (glucose, lactate, liver function tests, prothrombin time) and to assess liver regenerative response in the residual omental liver lobes (weight, protein content, incorporation of bromodeoxyuridine [BrdU], expression of proliferation cell nuclear antigen [PCNA], mitotic activity), plasma levels of hepatocyte growth factor (HGF) and transforming growth factor beta (TGF–beta1), and tissue expression of the HGF and it's receptor c–met. Rats undergoing partial hepatectomy of 68% (PH; n = 42) and a sham operation (SO; n = 42) served as controls. All SO and PH controls survived. PH rats showed only transient decreases in body temperature, signs of modest early hepatic dysfunction (hyperlactemia, hyperammonemia, prolonged PT time), and normal restitution of liver mass. All FHF rats became comatose by 24 hours postoperatively. Most animals (90%) died within 24–48 hours postoperatively (mean, 39 ± 11 hours). Changes in blood chemistry reflected rapid development of liver failure. Plasma HGF levels were markedly elevated and at all time points were higher than in PH controls (

P

< .05). At the same time, expression of HGF and c–met messenger RNA in the remnant liver was delayed. Plasma TGF–beta1 levels increased early (18 hours) and remained twofold to threefold higher than that of PH and SO controls (

P

< .05). There was only a 20% increase in the weight of the remnant liver lobes due to swelling. No hepatocytes stained positively for BrdU and PCNA, and none showed mitotic figures. In contrast, all PH controls showed vigorous liver regeneration. In conclusion, we have developed and characterized a novel model of FHF in rats that has a number of physiological and biochemical features seen clinically in FHF, including severely impaired ability of the residual liver tissue to regenerate.
Stimulation of Bile Acid Independent Bile Flow With Bromo–Cyclic Guanosine Monophosphate
Authors: St-Pierre M V , Dufour J , Arias I M
Published: November 2004

The second messenger, cyclic guanosine monophosphate (cGMP), mediates the actions of nitric oxide, natriuretic peptides, and microbial toxins on cellular contractility and electrolyte movement. Because both hepatocellular contractility and electrolyte secretion participate in bile formation, we investigated the actions of cGMP on this process in intact liver. In rat liver perfused with 8–bromo–cyclic GMP (bcGMP) at 0.5 and 3 micromol/min, bile flow increased by 5% and 31%, respectively. The biliary excretion of the bile acid, taurocholate ([3H]–labeled; 1 micromol/min) and of the organic anion, bromosulfophthalein ([35S]–labeled; tracer dose), was unchanged. The paracellular and transcytotic pathways of biliary excretion, assessed by horseradish peroxidase (HRP), were unaffected. BcGMP was concentratively secreted into bile and the accompanying 30% increase in the biliary clearance of erythritol suggested that the choleresis was primarily osmotic in nature. Unlike cyclic adenosine monophosphate (cAMP), which stimulates bile acid dependent bile flow and transcytosis, bcGMP increased bile acid independent bile flow mainly as a result of its concentrative biliary secretion.
The United Nations Convention on the Assignment of Receivables in International Trade: insolvency aspects
Authors: Bazinas Spiros V.
Published: November 2004

Breaking new ground, the UN Convention on the Assignment of Receivables in International Trade refers all priority conflicts with respect to receivables to the law of single and easily determinable jurisdiction, and one that is most likely going to be the insolvency jurisdiction, namely to the law of the assignor's place of business or, in the case of places of business in more than one State, the assignor's central administration. In the case of an insolvency proceeding in another jurisdiction, the mandatory rules of that juridiction displace any priority rule of the law of the assignor's location only if that priority rule is manifestly contrary to the public policy of that jurisdiction. In such a case, the balance of the priority rules of the law of the assignor's location prevails over the priority rules of the insolvency jurisdiction with the exception of rules relating to preferential rights. In any case, the Convention ensures that priority rules do not interfere with basic insolvency rights, such as those relating to stays, avoidance actions and to the performance of contracts or maintenance of the estate. Copyright © 2004 John Wiley & Sons, Ltd.
Web information tracking using ontologies
Authors: Maedche Alexander
Published: November 2004

Bringing knowledge management to practice one typically has to focus on concrete problems that exist in the daily work of the knowledge worker. We consider the task of tracking relevant information on the Web as important and time consuming, and thus as a concrete problem. In this paper we introduce an integrated approach for Web information tracking using ontologies. The overall approach has been implemented within a case study carried out at DaimlerChrysler AG. Copyright © 2004 John Wiley & Sons, Ltd.
Natural language interface for multi‐agent contracting system (MACS)
Authors: Yoon Victoria , Montano Bonnie Rubenstein , Wilson Teresa , Lowry Stuart , Liebowitz Jay
Published: November 2004

Facing serious imbalances in the skills and experience of our highly talented and specialized civilian workforce, the Naval Postgraduate School has initiated the External Acquisition Research Program (EARP) to promote advances in basic and applied research as related to US defense acquisition. One of the projects funded under EARP over a 2 year period is the Multi‐Agent Contracting System (MACS) project (Liebowitz et al., 2000. Journal of Knowledge‐Based Systems 13: 241–250). MACS was designed to provide advice in the pre‐award phase of a defense contract. The original version of MACS allowed users to query the system using a list of predetermined keywords from pull‐down menus. An exact string of keywords had to be used to obtain an answer from MACS, forcing the users to memorize a list of keywords. This inﬂexible user interface could have caused problems for those users who did not know the right keywords and, in turn, reduced the usability of MACS. The objective of this study is to extend the capability of MACS by allowing users to interface with the system using natural language. The natural language user interface will provide the users with more ﬂexibility in formulating queries on the pre‐award contracts, increasing the user friendliness of its interface. The paper presents the methods used to develop a natural language user interface for MACS. Copyright © 2004 John Wiley & Sons, Ltd.
Adhesion of human U937 macrophages to phosphorylcholine‐coated surfaces
Authors: Gong Yong‐Kuan , Luo Li , Petit Alain , Zukor David J. , Huk Olga L. , Antoniou John , Winnik Françoise M. , Mwale Fackson
Published: November 2004

A new type of amphiphilic phosphorylcholine (PC) polymer was used in this work to develop a cell culture surface that allows the attachment of U937 macrophages. The PC polymer was a random copolymer of N‐isopropylacrylamide (45%), N‐(phosphorylcholine)‐N′‐(ethylenedioxy‐bis(ethyl)) acrylamide (41%), and the hydrophobic monomer N‐(n‐octadecyl) acrylamide (14%). Polypropylene (PP) films (1 cm2) were coated with the polymer solution by immersion. U937 macrophage suspensions were applied on PC polymer–coated surfaces and incubated for up to 72 h at 37°C. While U937 cells did not adhere to PP, ammonia, nitrogen, or oxygen plasma‐treated surfaces, they attached rapidly on PC‐coated surfaces (< 1 h), proliferated, and stayed attached to the modified surface for at least 72 h, suggesting that unique features of the PC polymer, and the U937 macrophages, are responsible for the attachment of these cells. We compared the effect of Co2+ and Cr3+ ions on the expression of bone‐resorbing cytokines (TNF‐α, IL‐6, IL‐1β) in U937 macrophages cultured on PC‐coated surfaces to the response of U937 macrophages in suspension. Cytokine gene expression was analyzed by reverse transcription polymerase chain reaction (RT‐PCR). Addition of Co2+ and Cr3+ ions led to a significant increased expression of TNF‐α in both cultured and suspension cells. On the other hand, Co2+ and Cr3+ ions had a weak stimulatory effect or no effect on IL‐1β and IL‐6, respectively, in both cultured and suspension cells. In conclusion, the use of PC polymer–modified surfaces might offer promising new opportunities for the culture of human U937 cells and may also point to the mechanism by which macrophages interact with lipid bilayers of biological membranes. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 72A: 1–9, 2005
The influence of microtextured basal lamina analog topography on keratinocyte function and epidermal organization
Authors: Downing Brett R. , Cornwell Kevin , Toner Mehmet , Pins George D.
Published: November 2004

The rational design of future bioengineered skin substitutes requires an understanding of the mechanisms by which the three‐dimensional microarchitecture of tissue scaffolds modulates keratinocyte function. Microtextured basal lamina analogs were developed to investigate the relationship between the characteristic topography at the dermal–epidermal interface of native skin and keratinocyte function. Microfabrication techniques were used to create master patterns, negative replicates, and collagen membranes with ridges and channels of length scales (e.g., grooves of 50–200 μm in depth and width) similar to the invaginations found in basal lamina at the dermal–epidermal junction of native skin. Keratinocytes were seeded on the surfaces of basal lamina analogs, and histological analyses were performed after 7 days of tissue culture at the air–liquid interface. The keratinocytes formed a differentiated and stratified epidermis that conformed to the features of the microtextured membranes. Morphometric analyses of immunostained skin equivalents suggest that keratinocyte stratification and differentiation increases as channel depth increases and channel width decreases. This trend was most pronounced in channels with the highest depth‐to‐width ratios (i.e., 200 μm deep, 50 μm wide). It is anticipated that the findings from these studies will elucidate design parameters to enhance the performance of future bioengineered skin substitutes. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 72A: 47–56, 2005
Differentiation and drug resistance relationships in leukemia cells
Authors: Camarasa Maria V. , Castro‐Galache Maria D. , Carrasco‐García Estefanía , Garcia‐Morales Pilar , Saceda Miguel , Ferragut Jose A.
Published: November 2004

It is well established that the effectiveness of anticancer drugs may result from combined cytotoxic and differentiation activities on tumor cells. Also, differentiating agents are able to alter the susceptibility of cancer cells to antineoplastic drug therapy. However, the acquisition and/or development of drug resistance that frequently appears in anticancer treatment can impair these interactions between differentiation agents and cytotoxic drugs. In the present study, we report that the acquisition of resistance to anthracyclines in two humans, promyeolocytic leukemia HL‐60 and eythroleukemia K562 cell lines, results in a restricted maturation process induced by differentiating agents with respect to that exhibited by their corresponding drug‐sensitive counterparts. Interestingly, differentiating agents are able to decrease the overexpression of drug‐efflux pumps as it is the case of MRP1 in the resistant HL‐60 cells, thus increasing the sensitivity of cells to drug treatment. In addition, susceptibility of the drug‐sensitive cells to certain apoptotic stimuli is significantly reduced after differentiation. The results here reported indicate complex interactions between cytotoxic (drug therapy) and non‐cytotoxic (differentiation) cancer treatments, which should be taken into account to improve therapeutic efficiency. © 2004 Wiley‐Liss, Inc.
PTEN expression elicited by EGR‐1 transcription factor in calyculin A‐induced apoptotic cells
Authors: Okamura Hirohiko , Yoshida Kaya , Morimoto Hiroyuki , Haneji Tatsuji
Published: November 2004

PTEN is a tumor suppressor gene encoding a phosphatase that negatively regulates cell survival mediated by the PI3‐kinase‐Akt pathway. The gene for transcription factor EGR‐1 is an early response gene essential for cellular growth, proliferation, and differentiation. Protein phosphatase inhibitors including calyculin A and okadaic acid are potent inducers of apoptosis in several cell lines; however, the molecular mechanisms underlying their action are unknown. The purpose of this study was to examine the expression of PTEN and EGR‐1 and the phosphorylation status of EGR‐1 and Akt in calyculin A‐treated human squamous carcinoma cells (SCCTF). Phosphorylation of EGR‐1 and upregulation of PTEN expression were observed to occur in SCCTF cells treated with calyculin A in time‐ and dose‐dependent fashions. The level of phosphorylated Akt decreased as the expression of PTEN protein increased in the calyculin A‐treated SCCTF cells. Calyculin A‐stimulated expression of EGR‐1 and PTEN might be p53 independent, because the expression of them was also detected in p53‐null Saos‐2 cells. RNA interference using double‐stranded RNA specific for the EGR‐1 gene inhibited not only EGR‐1 expression but also PTEN expression in SCCTF cells treated or not with calyculin A. Calyculin A induced nuclear fragmentation and chromatin condensation in SCCTF cells. The present results suggest that the level of PTEN expression and the phosphorylation status of Akt were associated with apoptosis induced by calyculin A. These observations also support the view that EGR‐1 regulates PTEN expression in the initial steps of the apoptotic pathway. © 2004 Wiley‐Liss, Inc.
Bone marrow stromal cells express two distinct splice variants of ER‐α that are regulated by estrogen
Authors: Sanyal A. , Riggs B.L. , Spelsberg T.C. , Khosla S.
Published: November 2004

Estrogen plays a critical role in bone metabolism in both sexes. While the major action of estrogen is to inhibit bone resorption, it is now clear that early osteoblastic (or stromal) cells are a target for estrogen action, mediating the effects of estrogen on bone formation as well as resorption. However, little is known about the expression or regulation of the estrogen receptor (ER)‐α in these cells. The expression of ER‐α is regulated by a complex set of promoters and ER‐α splice variants are present in different tissues. Thus, we sought to define the ER‐α splice variants and their regulation by estrogen in the mouse bone marrow stromal cell line, ST‐2, which can be induced to differentiate into mature osteoblasts. ST‐2 cells expressed the mRNAs and proteins for both the 66 and 46 kDa forms of ER‐α; the latter lacks the AF‐1 domain and can transduce estrogen signaling in some tissues, while serving as a dominant negative receptor in others. Using primers specific for each of the five 5′‐untranslated exons of ER‐α, we found that ST‐2 cells utilized only the promoters upstream of exons F and C (in contrast to most reproductive tissues, which utilize promoters upstream of virtually all the five exons). Moreover, 17β‐estradiol (10−8 M) treatment of ST‐2 cells markedly diminished levels of the 66 kDa as well as the 46 kDa ER‐α proteins, largely through suppression of the transcript arising from the F1 promoter. These data thus indicate that: (1) bone marrow stromal cells express at least two variants of ER‐α and (2) estrogen down regulates the ER‐α mRNA and protein in these cells. © 2004 Wiley‐Liss, Inc.
High affinity leptin receptors are present in human mesenchymal stem cells (MSCs) derived from control and osteoporotic donors
Authors: Hess Rodrigo , Pino Ana María , Ríos Susana , Fernández Mireya , Rodríguez J. Pablo
Published: November 2004

There are disparate observations on central and peripheral effects of leptin, but several studies consistently support its role as a link between fat and bone. Bone marrow stroma contains mesenchymal stem cells (MSCs), which differentiate into osteoblasts and adipocytes, among others. In this study we assessed the expression of leptin receptors protein in MSCs from control and osteoporotic postmenopausal donors and their change during osteogenic and adipogenic differentiation. Also, we assessed the effects of leptin on osteogenic and adipogenic differentiation of these cells. We demonstrated high affinity leptin binding (KD = 0.36 ± 0.02 nM) in both types of cells. Binding was very low under basal, but increased significantly (2–3 times) through osteogenic and adipogenic differentiation. Osteoporotic MSCs showed lower leptin binding capacity than control cells at an early osteogenic and adipogenic differentiation time, which could restrict cell sensitivity to the protective action of leptin. In this regard, we observed that leptin significantly inhibited adipocyte differentiation in control but not in osteoporotic MSCs, while it exerted a low stimulatory effect on calcium deposition (10%–20%) in both types of MSCs cells. In summary, we report the presence of high affinity leptin receptors on control and osteoporotic MSCs, which were modified distinctly by osteogenic and adipogenic stimulation and a direct and distinct effect of leptin on both type of cells. © 2004 Wiley‐Liss, Inc.
Sensitive noninvasive marker for the diagnosis of probable bacterial or viral infection
Authors: Jortani Saeed A. , Pugia Michael J. , Elin Ronald J. , Thomas Meera , Womack Edward P. , Cast Todd , Valdes Roland
Published: November 2004

Urinary trypsin inhibitor (uTi) is a product of elastase‐mediated degradation of interleukin‐α‐inhibitor (I‐α‐I). Its activity increases in the urine of patients with a malignancy, inflammation, or infection, or in late pregnancy. The objective of this study was to compare the sensitivity of uTi in urine with that of serum quantitative C‐reactive protein (CRP) for diagnosing infection, as indicated by white cell response and clinical assessment. Ninety controls and 171 patients with various systemic infections were enrolled. We measured uTi enzymatically on a Cobas Fara (Roche Diagnostics). Patients were separated into bacterial, probable bacterial, viral, or probable viral groups based on the results of a complete blood count with differential (CBC), urinalysis (UA), and clinical assessment. In the bacterial (n=70) and control (n=90) groups, the uTi values (mean±SE) were 25.3±3.1 mg/L and 2.8±0.8 mg/L, respectively. uTi (at 2.7 mg/L) had a diagnostic sensitivity of 91% and specificity of 82% (AUC=0.889), whereas CRP (at a cutoff of 10 mg/L) had a sensitivity and specificity of 82% and 96%, respectively (AUC=0.921). As a marker of infection (positive in both bacterial and viral groups), uTi had a sensitivity of 91% (AUC=0.884) vs. 89% (AUC=0.828) for CRP. Our data indicate that uTi has sufficient clinical sensitivity for screening systemic infections, and may have diagnostic value as a noninvasive test. J. Clin. Lab. Anal. 18:289–295, 2004. © 2004 Wiley‐Liss, Inc.