bluuebunny/crossref_metadata_2025 huggingface dataset:
Long‐term Results of Total Adrenalectomy for Cushing's Disease
Authors: Nagesser Suresh K. , van Seters Arnoud P. , Kievit Job , Hermans Jo , Krans H. Michiel J. , van de Velde Cornelis J.H.
Published: August 2002

The objective of this study was to present the long‐term results of total adrenalectomy for Cushing's disease. Forty‐four patients undergoing total adrenalectomy for Cushing's disease between 1953 and 1989 at Leiden University Medical Center, The Netherlands, were studied retrospectively. Remission was achieved in 42 patients (95%), with a mean duration of 19 years. Adrenal remnants were observed in 12 patients (27%), and were without clinical consequence in the majority of patients, but caused early recurrent disease in 2 patients. Nine patients (20%) experienced Addisonian crises up to 30 years following treatment. Nelson's syndrome developed in 10 patients (23%) 7–24 years following total adrenalectomy. Prior pituitary irradiation was a protective factor against Nelson's syndrome as it delayed its onset (p= 0.025). On the other hand, subnormal dose or noncontinuous glucocorticoid replacement therapy was associated with increased risk of development of Nelson's syndrome (p= 0.047). The incidence of Nelson's syndrome increased with prolonged follow‐up, and female patients seemed to be at increased risk. Quality‐of‐life assessment showed less favorable scores on mental health and health perception scales, for which no explanation can be found except the long‐lasting metabolic effects of Cushing's disease, even when successfully treated. In conclusion, total adrenalectomy remains the final treatment for Cushing's disease. The presence of adrenal remnants which can cause recurrent disease and the development of Nelson's syndrome during prolonged follow‐up enhance the need for continued regular follow‐up. Pituitary irradiation prior to total adrenalectomy delays the onset of Nelson's syndrome.
Changes in Intestinal Transit Time after Proctocolectomy Assessed by the Lactulose Breath Test
Authors: Bruewer Matthias , Stern Josef , Herrmann Silke , Senninger Norbert , Herfarth Christian
Published: August 2002

After proctocolectomy with ileal pouch‐anal anastomosis (IPAA) patients have increased stool frequency and intermittently use antidiarrheal medication. In addition to other factors, gastrointestinal transit time (MTT) could influence stool frequency. The aim of this study was to investigate how MTT changes after IPAA and to study whether MTT has an influence on daily stool frequency. In a prospective trial MTT was investigated with the lactulose breath test in 12 patients undergoing surgery for chronic ulcerative colitis (CUC) or familial adenomatous polyposis coli (FAPC) at different stages: before proctocolectomy, after IPAA with loop ileostomy, and 3 months and 1 year after ileostomy closure. MTT was also measured in 12 patients with IPAA, 12 patients with subtotal colectomy and ileorectal anastomosis (IRA), and 8 patients with conventional proctocolectomy and Brooke ileostomy (CPC) several years after surgery. Twelve healthy volunteers served as controls. Before IPAA, MTT was prolonged in CUC versus FAPC and controls. After restoration of gut continuity MTT was markedly accelerated. After 1 year MTT was slowed again, though values before proctocolectomy and those in controls were not reached. Several years after surgery MTT was significantly prolonged in IPAA and IRA versus controls. In CPC, MTT could not be determined by lactulose breath test. Stool frequency showed an inverse correlation to MTT in IPAA. In conclusion, this study shows that orocecal and oropouch transit are accelerated in the early postoperative period after (procto)colectomy but prolonged in the long‐term course. Adaptation of the small bowel takes longer than 1 year. Impairment of stool frequency may be partly due to this adaptation.
Ultrasound of the Breast
Authors: Kossoff Marjorie Britt
Published: August 2002

Breast ultrasound (US) has developed into an essential imaging modality for evaluation and management of the patient with breast symptoms or mammographic abnormalities. US is usually an adjunct to mammography and the clinical examination but in the young patient presenting with symptoms sonography may be the initial or sole imaging procedure. A screening role has not been established, but there may be a subset of high‐risk patients for whom US will prove useful as a screening adjunct. Proper selection of equipment, careful attention to the technical aspects of scanning, and awareness of artifacts are necessary to avoid misinterpretation and to demonstrate subtle changes in breast architecture which indicate a malignancy. Thorough knowledge of the range of normal appearances of the breast and the alterations produced by breast disease are essential requisites for accurate evaluation and lesion characterization. Two of the most important clinical advances in breast US have been the development of criteria that allow improved benign/malignant differentiation of solid breast lesions and the use of US to guide interventional procedures. Other developments such as Doppler and contrast agents remain investigational.
Ultrasound of the Spleen
Authors: Andrews Matthew W.
Published: August 2002

Ultrasound (US) is a very useful means of noninvasively examining the spleen. Imaging is generally achieved via an intercostal approach using gray scale US supplemented by color flow to assess vasculature. Normal spleen appears uniform with vessels radiating and converging at the hilum. US readily identifies accessory spleens, a common congenital variation. Splenomegaly can be identified by US, however, it is not useful in differentiating the many causes of an enlarged spleen. Focal splenic masses are identified as cystic or solid by US, and features such as calcification, wall thickening, internal debris, and gas may be demonstrated in cystic type masses. Granulomatous calcification may be seen with US and cavernous hemangiomata typically have a characteristic US appearance. US can demonstrate typical features of splenic infarction. Splenic injury can be detected with US, however, whether US is the most appropriate first line investigation for suspected splenic injury is yet to be determined. It is a very useful follow‐up modality for monitoring splenic injuries detected by either US or computed tomography which are treated conservatively.
Review of Venous Vascular Ultrasound
Authors: Phillips Gareth W.L.
Published: August 2002

Duplex ultrasound (US) has become the new gold standard in the assessment of acute deep vein thrombosis. In view of the large number of cases with persistent changes, all cases should be reassessed at 6 months to document the extent of residual disease. The role of duplex US in chronic venous disease is less well established but it is evolving as the single most useful examination and is likely to supplant other noninvasive methods in the near future.
Metabolic Management of Severe Acute Pancreatitis
Authors: Windsor John A. , Hammodat Hisham
Published: August 2002

The metabolic management of severe acute pancreatitis involves early identification of patients with severe pancreatitis, aggressive fluid resuscitation, organ support, and careful monitoring in an intensive care environment. Recent evidence has helped to define the roles of enteral feeding, prophylactic antibiotics, endoscopic retrograde cholangiopancreatography, computed tomography, and fine‐needle aspiration for bacteriology. The most difficult decision in the management of these patients is whether surgery is required and which of the complementary approaches to necrosectomy and drainage is appropriate. Key metabolic events in the acinar cell, pancreas, and intestines are now being unraveled, as is the basis for the systemic manifestations and organ dysfunction associated with pancreatitis. This gives hope for the development of more specific metabolic interventions, which will likely target the maintenance of intestinal integrity and function, preservation of pancreatic microcirculation, and balanced modulation of the inflammatory response.
Interferon and Ribavirin for Patients With Chronic Hepatitis C Who Did Not Respond to Previous Interferon Therapy: A Meta–Analysis of Controlled and Uncontrolled Trials
Authors: Cheng Steve J. , Bonis Peter A.L. , Lau Joseph , Pham Nhu Q. , Wong John B.
Published: August 2002

The efficacy of interferon (IFN) combined with ribavirin for the treatment of patients with hepatitis C who failed to respond to initial IFN therapy is not well established. The primary goal of this study was to perform a systematic review of the literature evaluating the efficacy of combination therapy in nonresponders. Studies were retrieved from MEDLINE, abstracts of scientific meetings, and review of the bibliographies of retrieved studies. Controlled trials were included in the primary analysis whereas uncontrolled trials and trials reported as abstracts were included for sensitivity analysis. The primary endpoints were biochemical and virologic response. A combined estimate of the odds ratio (OR) for each endpoint was obtained by using the random effects model. The number needed to treat (NNT) was calculated by taking the inverse of the pooled risk difference. Nine controlled trials (789 patients) were identified. Six months after treatment, the overall sustained biochemical and virologic responses to 24 weeks of combination therapy were 15.2% and 13.2% with a common OR of 3.8 (95% confidence interval [CI] 2.2–6.7) and 4.9 (95% CI 2.1–11.2) compared with patients treated with IFN monotherapy. The pooled risk difference for the sustained virologic response (SVR) to combination therapy was 7% (95% CI 2–13). The NNT was 14 (95% CI 8–50), suggesting that approximately 14 patients would need to be treated with 6 months of combination therapy for 1 patient to have a SVR. A number of variables were associated with a high response rate in individual studies. Sensitivity analysis of preliminary trials suggest a higher response rate with longer duration of therapy and non–type 1 genotypes.
Human and Rat Hepatic Stellate Cells Express Neurotrophins and Neurotrophin Receptors
Authors: Cassiman David , Denef Carl , Desmet Valeer J. , Roskams Tania
Published: August 2002

The expression of neurotrophins and neurotrophin receptors in non–neural tissue is related to tissue remodeling, differentiation, proliferation and migration of target cells. The literature yields contradictory results on neurotrophin and neurotrophin receptor expression in the liver. We show immunoreactivity to antibodies to nerve growth factor (NGF), brain–derived neurotrophin (BDNF), neurotrophin 3 (NT–3), neurotrophin 4/5 (NT–4/5), the low–affinity nerve growth factor receptor p75 and the high–affinity tyrosine kinase receptors (Trk) B and C in hepatic stellate cells and weak reactivity for BDNF, NT–3, and NT–4/5 in hepatocytes, in cryosections of human and rat liver, in normal and varying pathologic conditions. Immunoreactivity is unequivocally localized to hepatic stellate cells by double staining with α–smooth muscle actin (α–SMA) and desmin, studied by confocal laser scanning microscopy. Finally, the presence of mRNA transcripts for the different neurotrophins and neurotrophin receptors, with the exception of Trk–B, is shown by reverse transcription polymerase chain reaction (RT–PCR) on RNA extracted from freshly isolated rat hepatic stellate cells, compared with hepatocyte RNA. Hepatocyte RNA was found to contain BDNF, NT–3, NT–4/5 mRNA (which is compatible with the immunohistochemical findings) and Trk–A mRNA. In conclusion, hepatic stellate cells are a source of several neurotrophins in the liver and they express neurotrophin receptors. These findings correspond with the known involvement of hepatic stellate cells in tissue remodeling, their production of extracellular matrix components and their proliferation in acute necrotizing liver pathology. In analogy with findings in other organs and systems, neurotrophins are hypothesized to play a role in the pathophysiology of liver disease.
Hepatitis C Virus Infection in A Community in the Nile Delta: Risk Factors for Seropositivity
Authors: Habib Mostafa , Mohamed Mostafa K. , Abdel-Aziz Fatma , Magder Laurence S. , Abdel-Hamid Mohamed , Gamil Foda , Madkour Salah , Mikhail Nabiel N. , Anwar Wagida , Strickland Thomas G. , Fix Alan D. , Sallam Ismail
Published: August 2002

The purpose of this study was to identify risk factors for hepatitis C virus (HCV) infection in a rural village in the Nile Delta with a high prevalence of antibodies to HCV (anti–HCV). One half of the village households were systematically selected, tested for anti–HCV, and interviewed: 973 of 3,999 (24.3%) subjects were anti–HCV–positive (reflecting prior HCV infection but not necessarily current liver disease), with nearly equal prevalence among males and females. Anti–HCV prevalence increased sharply with age among both males and females, from 9.3% in those 20 years of age and younger to >50% in those older than 35, suggesting a cohort effect with reduced transmission in recent years. Multivariate regression was used to estimate independent effects of risk factors on seropositivity. Among those over 20 years of age, the following risk factors were significantly associated with seropositivity: age (P < .001); male gender (odds ratio [OR] = 2.5, 95% CI = 1.3–4.7); marriage (OR = 4.1, 2.4–6.9); anti–schistosomiasis injection treatment (OR = 2.0, 1.3–2.9); blood transfusion (OR = 1.8, 1.1–2.9), invasive medical procedure (surgery, catheterization, endoscopy, and/or dialysis) (OR = 1.5, 1.1–1.9); receipt of injections from “informal” health care provider (OR = 1.3, 1.0–1.6); and cesarean section or abortion (OR = 1.4, 1.0–1.9). Exposures not significantly related to anti–HCV positivity in adults included: history of, or active infection with, Schistosoma mansoni, sutures or abscess drainage, goza smoking in a group, and shaving by community barbers. Among those 20 years old or younger, no risk factors were clearly associated with anti–HCV positivity; however, circumcision for boys by informal health care providers was marginally associated with anti–HCV (OR = 1.7, 1.0–3.0). Prevention programs focused primarily on culturally influenced risks in rural Egyptian communities are being implemented and evaluated.
Differential Regulation of Hepatic Bile Salt and Organic Anion Transporters in Pregnant and Postpartum Rats and the Role of Prolactin
Authors: Cao Jingsong , Huang Liyue , Liu Yong , Hoffman Tim , Stieger Bruno , Meier Peter J. , Vore Mary
Published: August 2002

We characterized expression and activity of the bile salt transporters Na+/taurocholate (TC) cotransporting polypeptide (Ntcp), and bile salt export pump (Bsep), and the expression of organic anion transporting polypeptides 1 and 2 (Oatp1 and 2) and multidrug resistance associated protein–2 (Mrp2) in pregnancy and throughout lactation in rats. The V
max for Na+/TC cotransport in basolateral liver plasma membrane was increased 1.7–fold in 2 days postpartum relative to control and pregnant rats. This correlated well with an increase in Ntcp messenger RNA (mRNA) and a 2–fold increase in Ntcp protein. Ntcp mRNA remained significantly elevated until 14 days postpartum but had begun to decline by 21 days postpartum. The maximal secretory rate (nmol/min/g liver) for TC in the single pass isolated perfused liver was also increased by 10%, 31%, and 24% at 2, 14, and 21 days postpartum and correlated with increased expression of Ntcp and Bsep mRNA and protein. Infusion of ovine prolactin (oPRL) to ovariectomized rats increased expression of both Ntcp and Bsep mRNA and protein. These data indicate a coordinate increased expression of bile salt transporters postpartum and by PRL. Mrp2 mRNA was stable in pregnancy and postpartum, whereas Mrp2 protein expression decreased significantly in pregnancy, but returned to control levels postpartum. Organic anion transporting polypeptide 2 (Oatp2) mRNA was decreased in pregnancy and increased postpartum, but changes in Oatp2 protein were not significant. Oatp1 mRNA and protein were unchanged in pregnancy and postpartum.
Hepatitis B Virus-Specific T–Cell Proliferation and Cytokine Secretion in Chronic Hepatitis B E Antibody-Positive Patients Treated With Ribavirin and Interferon Alfa
Authors: Rico Miguel A. , Quiroga Juan Antonio , Subirá Dolores , Castañón Susana , Esteban José Miguel , Pardo Margarita , Carreño Vicente
Published: August 2002

Immune elimination of hepatitis B virus (HBV) during antiviral therapy depends on the activation of T–cell responses, which are generally impaired in chronic hepatitis B. HBV–specific T helper (Th)–cell reactivity has been assessed post–treatment in liver and peripheral blood of 18 anti–HBe–positive patients with chronic hepatitis B administered combined ribavirin/interferon alfa (IFN–α) therapy. The results showed that patients with undetectable HBV DNA by quantitative polymerase chain reaction under combination therapy were able to mount an HBV–specific CD4+ Th–cell proliferative response and such T–cell reactivity is detectable 1 year after HBV DNA clearance. Hepatitis B virus core (HBcAg) and e (HBeAg) antigen–specific Th–cell proliferation was found more frequently in the liver and peripheral blood in those patients who sustained the alanine aminotransferase (ALT) normalization together with HBV DNA loss. However, HBV–specific IFN–γ production in vitro in peripheral blood mononuclear cells augmented in 4 of 5 sustained responders and all 13 nonresponders, interleukin 10 (IL–10) production decreased in all 5 sustained responders but increased in 7 of 13 nonresponders. Furthermore, intrahepatic HBcAg plus HBeAg–specific Th–cell proliferation only occurred in sustained responders (2 of 3, 67%, vs. 0 of 9; P = .045) whose cells showed in vitro significantly increased productions in HBcAg/HBeAg–specific IFN–γ and IL–12 compared with nonresponders in whom IFN–γ and IL–12 productions decreased together with increased IL–10 secretion. In conclusion this study indicates that combined therapy with ribavirin and IFN–α for chronic hepatitis B not only significantly reduces viremia levels but also induces lasting CD4+ T–cell proliferation and Th1 cytokine release at the site of infection, which may lead to sustained eradication of the HBV.
Prevalence and Significance of Hepatitis B Virus (Hbv) Pre–S Mutants in Serum and Liver At Different Replicative Stages of Chronic Hbv Infection
Authors: Fan Yu-Fen , Lu Cheng-Chan , Chen Wen-Chi , Yao Wei-Jen , Wang Hui-Ching , Chang Ting-Tsung , Lei Huan-Yao , Shiau Ai-Li , Su Ih-Jen
Published: August 2002

Several types of naturally occurring pre–S mutants in sera or liver tissues in patients with chronic hepatitis B virus (HBV) infection have been identified. To clarify the prevalence and significance of emergence of pre–S mutants, 140 sera and 18 resected livers from patients with HBV were studied. Replicative status was designated as high, intermediate, and low based on the HBV–DNA levels in serum or the expression of HBV antigens in liver. In vitro transfection and Western blot analysis were performed to characterize expression and secretion of HBsAg by the mutant constructs. Five major types (I to V) of pre–S deletion mutants in serum and liver and 2 types (VI and VII) in liver were identified. Pre–S mutant was 6.4% at high replicative phase, 13% at intermediate, and 37.5% at low or nonreplicative phases in serum. In livers, the same tendency existed: pre–S2 deletion mutants emerged and prevailed at a low replicative phase in hepatocytes that expressed a novel marginal pattern of HBsAg and usually clustered in groups. The deletion sequence of pre–S2 region coincides with human leukocyte antigen–restricted T– and B–cell epitopes. In vitro HBsAg was retained in the hepatocytes and synthesis and secretion of major surface antigen decreased for most of the pre–S mutants. Pre–S mutants prevailed with evolution of chronic HBV, probably under immune pressure. Emergence of pre–S mutants may account for the life–long persistence and discrepancy of HBsAg in serum and liver in HBV and may confer growth advantage in view of the clustering proliferation of hepatocytes harboring pre–S2 mutant.
Dna Prime/Canarypox Boost—Based Immunotherapy of Chronic Hepatitis B Virus Infection in A Chimpanzee
Authors: Pancholi Preeti , Lee Dong-Hun , Liu Qingyan , Tackney Charles , Taylor Patricia , Perkus Marion , Andrus Linda , Brotman Betsy , Prince Alfred M.
Published: August 2002

There are about 200 million chronic hepatitis B virus (HBV) carriers at high risk of development of cirrhosis and hepatocellular carcinoma. Termination of the carrier state may avert these risks. We have investigated immunotherapy for chronic HBV infection in a chimpanzee HBV carrier using recombinant DNA–based immunization followed by a recombinant canarypox booster. One week after the booster, HBV DNA declined greater than 400–fold and remained undetectable by the quantitative polymerase chain reaction (PCR) assay for 186 weeks. Plasma levels of hepatitis B surface antigen (HBsAg) declined for only a short time. The decline in HBV DNA correlated with a boost in gamma interferon production without a corresponding boost in cytotoxic T lymphocyte levels, and decline in the transcriptional template or covalently closed circular DNA level. Confirmation of these findings requires further studies in chimpanzees and/or in humans.
Irregular Regeneration of Hepatocytes Is An Important Factor in the Hepatocarcinogenesis of Liver Disease
Authors: Ueno Yukiko , Moriyama Mitsuhiko , Uchida Toshikazu , Arakawa Yasuyuki
Published: August 2002

The goal of this study was to assess the relationship between the degree of irregular regeneration of hepatocytes (IR) and the development of hepatocellular carcinoma (HCC). We examined the liver biopsy specimens from patients with chronic liver disease (CLD), and the noncancerous liver tissue in the surgically resected specimens from patients with HCC. Liver tissues were obtained from 18 patients with B–viral CLD, 38 with C–viral CLD, and 5 with nonB–nonC (NBNC)–CLD who underwent liver biopsy, and 24 patients with B–viral HCC, 67 with C–viral HCC, and 17 with NBNC–HCC who underwent surgical resection. The IR score of the noncancerous region of the liver of the HCC patients and that of the liver biopsy specimens of the CLD patients, were compared. Of the 108 samples of noncancerous liver tissue from the HCC patients, 70% showed an IR score of moderate or higher (IR = 3 or 4), and only 1.9% showed an IR score of none or minimal (IR = 0 or 1). The degree of IR of hepatocytes in the HCC patients was significantly more severe than that in the CLD patients in each etiological type of CLD. High IR may reflect the histologic expression of genetic instability, namely a carcinogenic state, although prospective studies are needed to determine its relationship to the risk for developing HCC.
Expression of Cyclooxygenase–2 Promotes the Release of Matrix Metalloproteinase–2 and –9 in Fetal Rat Hepatocytes
Authors: Callejas Nuria A. , Casado Marta , Díaz-Guerra María J.M. , Boscá Lisardo , Martín-Sanz Paloma
Published: August 2002

Treatment of primary cultures of fetal hepatocytes with proinflammatory cytokines, lipopolysaccharide (LPS), and hepatocyte growth factor promoted the expression of cyclooxygenase–2 (COX–2) and the synthesis of high amounts of prostaglandins (PGs). Under these conditions, the active forms of the matrix metalloproteinases–2 and –9 (MMPs) were released to the extracellular medium. This process was inhibited when the synthesis of PGs was suppressed pharmacologically with COX–2 inhibitors. Addition to the cell cultures of PGE2 promoted the release of MMPs through a mechanism that involved the expression of COX–2 and the synthesis of additional PGs. Kinetic analysis of the secretion of MMPs in response to LPS and PGE2 showed a similar time course, with a lag period of 6 hours, which suggests that PGE2 does not act directly on the mechanism of MMP processing and release. Inhibitors of protein kinase A, p38 MAP kinase, phosphatidylinositol–3–kinase, and nuclear factor κB (NF–κB) activation impaired the release of MMPs in response to PGE2 challenge, indicating the involvement of multiple steps in the process. The ability of fetal hepatocytes to release MMPs in response to growth factors and inflammatory stimuli constitutes a model for the study of the extracellular matrix remodeling that accompanies most liver diseases.
A Meta–Analysis of Endoscopic Variceal Ligation for Primary Prophylaxis of Esophageal Variceal Bleeding
Authors: Imperiale Thomas F. , Chalasani Naga
Published: August 2002

Despite publication of several randomized trials of prophylactic variceal ligation, the effect on bleeding–related outcomes is unclear. We performed a meta–analysis of the trials, as identified by electronic database searching and cross–referencing. Both investigators independently applied inclusion and exclusion criteria, and abstracted data from each trial. Standard meta–analytic techniques were used to compute relative risks and the number needed to treat (NNT) for first variceal bleed, bleed–related mortality, and all–cause mortality. Among 601 patients in 5 homogeneous trials comparing prophylactic ligation with untreated controls, relative risks of first variceal bleed, bleed–related mortality, and all–cause mortality were 0.36 (0.26–0.50), 0.20 (0.11–0.39), and 0.55 (0.43–0.71), with respective NNTs of 4.1, 6.7, and 5.3. Among 283 subjects from 4 trials comparing ligation with β–blocker therapy, the relative risk of first variceal bleed was 0.48 (0.24–0.96), with NNT of 13; however, there was no effect on either bleed–related mortality (relative risk [RR], 0.61; confidence interval [CI], 0.20–1.88) or all–cause mortality (RR, 0.95; CI, 0.56–1.62). In conclusion, compared with untreated controls, prophylactic ligation reduces the risks of variceal bleeding and mortality. Compared with β–blockers, ligation reduces the risk for first variceal bleed but has no effect on mortality. Prophylactic ligation should be considered for patients with large esophageal varices who cannot tolerate β–blockers. Subsequent research should further compare ligation and β–blockers to determine the effect on mortality, and measure ligation's cost–effectiveness.
Resistance to Fulminant Hepatitis and Carcinogenesis Conferred by Overexpression of Retinoblastoma Protein in Mouse Liver
Authors: Ichihara Toshiaki , Komagata Yoshinori , Yang Xiao-Li , Uezato Tadayoshi , Enomoto Katsuhiko , Koyama Kenji , Miyazaki Jun-ichi , Sugiyama Toshihiro , Miura Naoyuki
Published: August 2002

Previously, retinoblastoma (Rb) transgenic mice were produced under the control of the Rb gene promoter and showed dwarf characteristics. Here, we created transgenic mice, in which the human Rb gene was controlled by the hepatocyte nuclear factor–1 gene promoter/enhancer and was expressed primarily in the liver. The liver of these novel transgenic mice was normally developed. Intriguingly, these mice showed resistance to fulminant hepatitis induced by anti–Fas antibody as well as resistance to chemical carcinogenesis in the liver. These results show that the Rb protein acts as an anti–apoptotic and anti–oncogenic agent in vivo. Our novel construct may be useful as a gene cassette in gene therapy for prevention of fulminant hepatitis and hepatoma.
Tumor Hepatocytes and Basement Membrane-Producing Cells Specifically Express Two Different Forms of the Endostatin Precursor, Collagen Xviii, in Human Liver Cancers
Authors: Musso Orlando , Theret Nathalie , Heljasvaara Ritva , Rehn Marko , Turlin Bruno , Campion Jean-Pierre , Pihlajaniemi Taina , Clement Bruno
Published: August 2002

Endostatin is an endogenous inhibitor of angiogenesis and tumor growth in mice, which may be generated by proteolytic cleavage of collagen XVIII. In normal tissues, 2 variants of the endostatin precursor, namely the SHORT and LONG forms, regulate tissue specificity. We analyzed 53 human liver biopsies (18 hepatocellular carcinomas, 16 metastases of colorectal cancer, 3 cholangiocarcinomas, and 16 controls) by RNA dot blots, double–labeling immunohistochemistry, and in situ hybridization, using common and variant–specific probes. Tumor hepatocytes expressed the LONG form, whereas cholangiocarcinoma cells expressed the SHORT form, which was deposited in tumor basement membranes. Metastatic colorectal carcinoma cells did not express collagen XVIII. In the stromal compartment of primary and metastatic cancers, myofibroblasts and vascular endothelial cells expressed the SHORT form. Both basement membrane components, collagen IV and the SHORT collagen XVIII form, were codistributed and their mRNA levels strongly correlated (R = .75, P < .001). In addition, freshly isolated human hepatocytes expressed the LONG form and culture–activated stellate cells the SHORT form. Moreover, the full–length LONG form is a plasma protein. Thus, the LONG form is a hepatocyte–specific variant, and the SHORT form is a major component of the tumor extracellular matrix in primary and metastatic liver cancers. In the clinical context, the global expression of the endogenous endostatin precursor, collagen XVIII, in liver cancer results from the combined expression profiles of tumor cells, stromal cells, and nontumor hepatocytes at the advancing edge of the tumor, particular to each type of cancer.
Normal Liver Regeneration in P50/Nuclear Factor κB1 Knockout Mice
Authors: DeAngelis Robert A. , Kovalovich Kellen , Cressman Drew E. , Taub Rebecca
Published: August 2002

Nuclear factor κB (NF–κB) is rapidly activated during liver regeneration following partial hepatectomy or carbon tetrachloride (CCl4)–mediated liver injury and is felt to be important in the antiapoptotic and regenerative responses. After partial hepatectomy, livers of mice deficient in the p50 subunit of NF–κB (p50-/-) showed a loss of NF–κB and decreased STAT3 transcription factor DNA binding activities. However, nuclear levels of the NF–κB p65 subunit were increased and peaked earlier in p50-/- livers. Both messenger RNA and cytoplasmic protein levels of the NF–κB inhibitor IκBα were lower in p50-/- livers, potentially accounting for the increase in p65 protein. Small effects on gene expression posthepatectomy were observed in p50-/- livers, but no effects were seen on hepatocyte DNA synthetic or mitotic responses, serum enzyme levels, or overall liver mass restoration. After CCl4 treatment, hepatocyte DNA synthesis and mitosis and serum enzyme levels were similar in p50-/- and p50+/+ mice, and histologic analysis indicated a slight decrease in overall damage in p50-/- livers. After injection of Fas antibody, p50-/- livers showed an earlier onset of nuclear changes consistent with apoptosis. These data indicate that absence of p50 affects certain protein and gene activation pathways following partial hepatectomy, CCl4, and Fas treatment but does not impair overall liver regeneration. Interleukin 6 (IL–6) levels were reduced but still adequate to support regeneration. We hypothesize that increased levels of the NF–κB p65 subunit in p50-/- livers may provide compensation for the absence of p50, thereby allowing normal liver regeneration and repair following liver injury.
Effect of Nitric Oxide on Shock–Induced Hepatic Heme Oxygenase–1 Expression in the Rat
Authors: Hoetzel Alexander , Vagts Dierk A. , Loop Torsten , Humar Matjaz , Bauer Michael , Pahl Heike L. , Geiger Klaus K. , Pannen Benedikt H.J.
Published: August 2002

Recent evidence suggests that the hepatic expression of heme oxygenase–1 (HO–1) may preserve hepatocellular integrity after hemorrhagic shock and resuscitation (HR). Because nitric oxide (NO) has been shown to modulate HO–1 expression in cultured cells in vitro, we determined its potential role in the regulation of HO–1 expression after HR in the rat liver in vivo. HO–1 mRNA and protein were highly induced and HO enzyme activity was higher after HR when compared with time–matched sham controls. Administration of the NO donor, molsidomine (MOL) (3 mg · kg-1), during resuscitation attenuated the accumulation of HO–1 mRNA and protein and the rise in HO activity. In addition, MOL prevented the shock–induced increase in DNA binding activity of the transcription factor, activator protein–1 (AP–1), but did not alter the activity of nuclear factor–erythroid 2 related factor (Nrf–2), nuclear transcription factor–κB (NF–κB), and hypoxia–inducible factor–1 (HIF–1). The suppressing action of MOL was not confined to HO–1, because the hepatic expression of the 70–kd major heat shock protein (HSP) in response to HR was also diminished. Moreover, MOL prevented the HR–induced increase in the serum activity of alanine transaminase (ALT) and α–glutathione–S–transferase (α–GST) that could otherwise be observed after HR. In contrast, the NO synthase inhibitor, N
ω–nitro–l–arginine methyl ester (l–NAME) (1 mg · kg-1), had either no or only minor effects on the primary experimental endpoints. These findings would be consistent with a reduction of shock–induced liver damage by exogenous NO, which in turn prevents the subsequent activation of injury–sensitive transcription factors, thus attenuating the expression of stress–inducible proteins such as HO–1.
In Vitro And In Vivo Activation of Rat Hepatic Stellate Cells Results In De Novo Expression of L–Type Voltage–Operated Calcium Channels
Authors: Bataller Ramón , Gasull Xavier , Ginès Pere , Hellemans Karine , Görbig Nieves M. , Nicolás Josep M. , Sancho-Bru Pau , De Las Heras Dara , Gual Arcadi , Geerts Albert , Arroyo Vicente , Rodés Juan
Published: August 2002

Following chronic liver injury, hepatic stellate cells (HSCs) transdifferentiate into myofibroblast–like cells, which develop contractile properties and contribute to increased resistance to blood flow. We investigated whether this phenotypic activation includes changes in the expression of L–type voltage–operated Ca2+ channels (VOCC), which mediate Ca2+ influx and regulate cell contraction in vascular cell types. Rat HSCs were studied in the quiescent phenotype and after their activation in vitro (cultured on plastic for 14 days) and in vivo (isolated from rats with CCl4–induced cirrhosis). Patch–clamp studies showed Ca2+ currents through L–type VOCC in HSCs activated both in vitro and in vivo, whereas no currents were detected in quiescent HSCs. Moreover, binding studies with 3H–isradipine, a specific L–type VOCC antagonist, showed a large number of binding sites in activated HSCs, while no specific binding was found in quiescent HSCs. Finally, messenger RNA (mRNA) encoding L–type VOCC was not detected in quiescent HSCs as assessed by reverse transcription–polymerase chain reaction (RT–PCR) and Northern blot analysis, whereas it was present in activated HSCs. Stimulation of L–type VOCC with KCl resulted in a marked increase in [Ca2+]i followed by cell contraction in HSCs activated both in vitro and in vivo, whereas no effects were observed in quiescent HSCs. We conclude that the activation of HSCs is associated with up–regulation of L–type VOCC that mediate Ca2+ influx and cell contraction. These results may be relevant to the pathogenesis of portal hypertension.
Frequent Loss of Heterozygosity At The Hcr1 ( Hepatocarcinogenesis Resistance ) Locus on Chromosome 10 in Primary Hepatocellular Carcinomas From Lff1 Rat Strain
Authors: De Miglio Maria R. , Muroni Maria R. , Simile Maria M. , Virdis Patrizia , Asara Giuseppina , Frau Maddalena , Calvisi Diego F. , Seddaiu Maria A. , Pascale Rosa M. , Feo Francesco
Published: August 2002

Hepatocarcinogenesis sensitivity (Hcs1, 2) and resistance (Hcr1–3) loci have been identified by linkage analysis on rat chromosomes 7 and 1, and 10, 4, and 8, respectively. Cytogenetic studies documented deletions on chromosomes 3 and 6 of neoplastic rat hepatocytes. Hepatocellular carcinomas (HCCs) were produced in F1 hybrid rats between Long–Evans (LE) and Fisher 344 (F344) rats. Scanning of the above chromosomes for loss of heterozygosity (LOH) showed allelic imbalance (AI) at multiple regions on chromosomes 6, 7, and 10q. Detailed deletion mapping of chromosome 10 localized a putative suppressor Hcr1 gene to within a 3.2–cM interval flanked by D10Rat51 and D10Rat121. Two other distinct regions with frequent AIs were found inside the Hcr1 locus, at marker loci including DNaseI and Mrp genes, and in a segment including 4 consecutive markers (D10Rat64, D10Rat182, D10Rat113, D10Rat216). In 40% of HCCs, AI was seen at the p53 locus. AI on chromosome 7 occurred at the Hcs1 locus, where is located c–myc, which is amplified in HCCs, suggesting allelic gain. Most AIs occurred in poorly/moderately differentiated carcinomas, and a few events were seen in well–differentiated tumors on chromosomes 7 and 10. These data suggest that alteration of a cluster of oncosuppressor genes on 10q is important for HCC progression. The existence of AI on segments of rat chromosomes 6, 7, and 10, syntenic to chromosomal segments of human HCCs where chromosomal gains or deletions occur, suggests a commonalty of some molecular events in the pathogenesis of HCCs in rats and humans. Our map provides information toward cloning putative oncosuppressor genes associated with this carcinoma. (Hepatology 2001;33:1110–1117.)
Long–Term Follow–Up Study of Sustained Biochemical Responders With Interferon Therapy
Authors: Shindo Michiko , Hamada Kazushige , Oda Yoko , Okuno Tadao
Published: August 2002

A proportion of chronic hepatitis C patients who were treated with interferon have a sustained normalization of transaminase levels after interferon therapy without hepatitis C virus (HCV)–RNA clearance. We determined their clinical characteristics and long–term outcome in relation to progression to liver cirrhosis (LC) and the development of hepatocellular carcinoma (HCC). A total of 250 patients with chronic hepatitis C who were treated with interferon were studied for 8 to 11 years' posttherapy. Sixty–seven patients (27%) were complete responders with clearance of HCV RNA. Twenty–six (10%) were biochemical responders who had sustained normal alanine transaminase (ALT) levels without viral clearance. The remaining patients were short–term responders (n = 70) and nonresponders (n = 87). Biochemical responders were older, had higher levels of pretreatment HCV RNA in serum than complete responders, and had less advanced liver histology than nonresponders. Histologic grading scores decreased significantly at the end of therapy, while the staging scores did not change significantly. The annual incidence of cirrhosis was 0% in biochemical and complete responders, which was significantly lower than nonresponders and the controls (P = .0001). The annual incidence of HCC was 0.37% in complete responders and 0.50% in biochemical responders, which was significantly lower than nonresponders (P = .0001 for both). Our findings suggest that biochemical responders had high pretreatment viral levels with less advanced liver histology, and their long–term outcome appeared to be good irrespective of the persistence of the virus. (Hepatology 2001;33:1299–1302.)
Increased Risk of Tumor Seeding After Percutaneous Radiofrequency Ablation for Single Hepatocellular Carcinoma
Authors: Llovet Josep M. , Vilana Ramon , Brú Concepció , Bianchi Lluís , Salmeron Joan Manuel , Boix Loreto , Ganau Sergi , Sala Margarita , Pagès Mario , Ayuso Carmen , Solé Manel , Rodés Joan , Bruix Jordi
Published: August 2002

Radiofrequency (RF) ablation is an alternative to percutaneous ethanol injection (PEI) for single nonsurgical hepatocellular carcinoma (HCC) and is currently used as adjuvant therapy before liver transplantation. This phase II study assesses the treatment–related complications and response rate of RF for the treatment of single HCC ≤5 cm. Percutaneous RF was performed under conscious sedation and ultrasound (US) guidance with an electrical generator connected to a single cooled–tip electrode. Neoplastic cells in peripheral blood (reverse transcription–polymerase chain reaction for alpha fetoprotein [AFP] messenger RNA) were analyzed before and after RF. Treatment response was assessed by spiral computed tomography (CT) at 1 month and every 3 months by US or spiral CT thereafter. Thirty–two patients (20 men; age 67 ± 4 years; 78% hepatitis C virus; 24 Child–Pugh A) with a mean tumor size of 2.8 cm (25 patients ≤3 cm) were treated by RF (1.25 sessions; mean time, 22.1 ± 2 minutes). Adjuvant PEI was performed in 9 cases. Complete response was achieved in 21 patients (65%), being significantly higher for HCC ≤3 cm (76% vs. 29%, P = .03). After a median follow–up of 10 months, 8 patients showed treatment–related morbidity. Four of them (12.5%) showed biopsy–proven needle–track seeding detected between 4 to 18 months. Neoplastic seeding was related to subcapsular location (P = .009), poor differentiation degree (P = .02), and baseline AFP levels (P = .02). Thus, RF ablation with cooled–tip needle for HCC is associated with a high risk of neoplastic seeding. Iatrogenic dissemination was related to subcapsular location or an invasive tumoral pattern, and has to be considered when selecting curative treatments for HCC or adjuvant therapies before liver transplantation.
Steatosis Accelerates the Progression of Liver Damage of Chronic Hepatitis C Patients and Correlates With Specific Hcv Genotype and Visceral Obesity
Authors: Adinolfi Luigi E. , Gambardella Michele , Andreana Augusto , Tripodi Marie-françoise , Utili Riccardo , Ruggiero Giuseppe
Published: August 2002

The role of steatosis in the progression of liver damage in chronic hepatitis C (CHC) was studied. Enrolled were 180 consecutive liver biopsy–proven CHC patients and 41 additional subjects with a known duration of infection. We evaluated the histological activity index (HAI), grade of fibrosis and steatosis, body mass index (BMI; kg/m2), distribution of body fat, HCV genotype, and levels of HCV RNA. Eighty six (48%) patients showed steatosis, and a higher prevalence was observed in genotype 3a infection (P < .01). A correlation between the grade of steatosis and fibrosis was observed (P < .001). Fibrosis was also associated with age (P < .001). After adjusting for age, the association between steatosis and fibrosis remained significant. The grade of steatosis also correlated with the HAI (P < .007) with a significant increase in periportal necrosis. No relation was found between steatosis and age, gender, iron storage, or levels of HCV RNA. Patients with a high grade of steatosis (>30%) showed higher serum levels of γ–GT and ALT (P < .001). Overall, steatosis was not significantly associated to BMI. Analysis by single genotype showed a significant association between the grade of steatosis and BMI in type 1 infection r = .689; P < .001) and with levels of HCV RNA in type 3a infection r = .786; P < .001). Visceral fat distribution rather than BMI proved to be associated with steatosis (P < .001). Data obtained from patients with a known date of infection confirmed that steatosis grades 3–4 were associated with a higher annual rate of fibrosis progression, and showed that alcohol and steatosis act together in increasing fibrosis (P < .05). Our data indicate that steatosis is an important cofactor in increasing liver necroinflammatory activity and in accelerating fibrosis in CHC. Visceral obesity and genotype 3a play a role in the development of steatosis.
Alterations in Tight Junctions Differ Between Primary Biliary Cirrhosis and Primary Sclerosing Cholangitis
Authors: Sakisaka Shotaro , Kawaguchi Takumi , Taniguchi Eitaro , Hanada Shinichiro , Sasatomi Kurumi , Koga Hironori , Harada Masaru , Kimura Rina , Sata Michio , Sawada Norimasa , Mori Michio , Todo Satoru , Kurohiji Toshihiko
Published: August 2002

Tight junctions (TJ) of biliary epithelial cells (BEC) and hepatocytes prevent bile regurgitation from the biliary tract. Alterations in these TJs may participate in chronic cholestatic liver diseases such as primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC). We examined the localization of 2 TJ proteins, ZO–1 and 7H6, in these diseases. Frozen sections from livers of PBC, PSC, extrahepatic cholestasis (Ex–C), and hepatitis C-associated cirrhosis (LC–C), as well as histologically normal livers, were processed for double–fluorescence immunohistochemistry. In controls and cirrhosis, 7H6 and ZO–1 colocalized surrounding the luminal space of the bile ducts and outlined the bile canalicular spaces between hepatocytes. In untreated PBC, immunostaining for ZO–1 in BEC of bile ducts 40 to 80 μm in diameter was preserved, but that for 7H6 was diminished to absent. In PBC treated with ursodeoxycholic acid (UDCA), immunostaining for 7H6 was well preserved. In PSC as well as in Ex–C, immunostaining for both 7H6 and ZO–1 was well preserved in bile ducts. In hepatocytes, ZO–1 showed preserved immunoreactivity, but immunostaining for 7H6 frequently disappeared. The percentage of bile ducts with immunostaining for 7H6 in all bile ducts with immunostaining for ZO–1 was significantly reduced in PBC compared with that in control, LC–C, Ex–C, and PSC (all P < .0001). Substantial alteration in the TJ protein occurs predominantly in bile ducts in PBC and in hepatocytes in PSC, suggesting increased paracellular permeability along different paracellular routes for bile regurgitation in these chronic cholestatic liver diseases.
Roles of Akt/Pkb and Ikk Complex in Constitutive Induction of Nf–κB in Hepatocellular Carcinomas of Transforming Growth Factor α/C– Myc Transgenic Mice
Authors: Factor Valentina , Oliver Aundrea L. , Panta Ganesh R. , Thorgeirsson Snorri S. , Sonenshein Gail E. , Arsura Marcello
Published: August 2002

NF–κB regulates liver cell death during development, regeneration, and neoplastic transformation. For example, we showed that oncogenic Ras– or Raf–mediated transformation of rat liver epithelial cells (RLEs) led to altered NF–κB regulation through IKK complex activation, which rendered these cells more resistant to TGF–β1–induced apoptosis. Thus, based on these findings, we sought to determine whether NF–κB could also be involved in tumor growth of liver cells in vivo. Hepatocellular carcinomas (HCCs) derived from bitransgenic mice harboring TGF–α and c–myc transgenes targeted specifically to the liver were compared with HCCs from c–myc single transgenic mice. Tumors from bitransgenic mice are characterized by a higher frequency of appearance, lower apoptotic index, and a higher rate of cell proliferation. Here we show that NF–κB is activated in HCCs of double TGF–α/c–myc transgenic mice, but not of c–myc single transgenic mice, suggesting that TGF–α mediates induction of NF–κB. Activation of the IKK complex was observed in the HCCs of double TGF–α/c–myc transgenic mice, implicating this pathway in NF–κB induction. Lastly, activation of the Akt/protein kinase B (PKB), which has recently been implicated in NF–κB activation by PDGF, TNF–α, and Ras, was also observed. Importantly, human HCC cell lines similarly displayed NF–κB activation. Thus, these studies elucidate an anti–apoptotic mechanism by a TGF–α–Akt/PKB–IKK pathway, which likely contributes to survival and proliferation, thereby accelerating c–myc–induced liver neoplastic development in vivo.