bluuebunny/crossref_metadata_2025 huggingface dataset:
A murine model of type 2 autoimmune hepatitis: Xenoimmunization with human antigens
Authors: Lapierre Pascal , Djilali-Saiah Idriss , Vitozzi Susana , Alvarez Fernando
Published: March 2004

Autoimmune hepatitis (AIH) is characterized by an immune-mediated injury of the hepatic parenchyma of unknown pathogenesis. Type 2 AIH is identified by the presence of anti-liver-kidney microsomes type 1 (anti-LKM1) and anti-liver cytosol type 1 (anti-LC1) autoantibodies. The current study shows that a murine model of AIH can be generated by DNA immunization against type 2 AIH self-antigens (P450 2D6 and formiminotransferase-cyclodeaminase). A pCMV plasmid containing the N-terminal region of mouse CTLA-4 and the antigenic region of human CYP2D6 (672-1,377 bp) and human formiminotransferase cyclodeaminase (FTCD; 1,232-1,668 bp) was used for DNA immunization of C57BL/6 female mice. Immunized mice showed elevated levels of alanine aminotransferase (ALT), with peaks at 4 and 7 months postinjection. Periportal, portal, and intralobular liver inflammatory infiltrates were observed at histology. Mainly CD4+ lymphocytes, but also CD8+ and B lymphocytes, were found in the liver. Cytotoxic-specific T cells were found in both the liver and spleen of these animals. Mice developed anti-LKM1 and anti-LC1 antibodies of immunoglobulin G2 (IgG2) subclass, against specific mouse autoantigens. The ALT levels correlated with both the presence of anti-LKM1/anti-LC1 antibodies and the presence of liver necroinflammation. In conclusion, in mice, DNA immunization against human autoantigens breaks tolerance and induces an autoimmune liver disease. Molecular mimicry between foreign and self-antigens explains the liver injury. This model of AIH resembles human type 2 AIH and will be helpful for the study of its pathogenesis. (Hepatology 2004;39:1066-1074.)
Analyses of hepatocellular proliferation in a mouse model of α-1-antitrypsin deficiency
Authors: Rudnick David A. , Liao Yunjun , An Jae-Koo , Muglia Louis J. , Perlmutter David H. , Teckman Jeffrey H.
Published: March 2004

α-1-Antitrypsin (α1-AT) deficiency is the most common cause of metabolic pediatric liver disease. Hepatocellular injury is caused by toxicity of the mutant α-1-antitrypsin Z (α1-ATZ) molecule retained within hepatocytes. In these studies, we used the PiZ transgenic mouse model of α1-AT deficiency to examine hepatocellular proliferation in response to chronic liver injury resulting from this metabolic disease. The results showed increased hepatocellular proliferation and caspase 9 activation in male PiZ mice compared with female PiZ and wild-type mice. Hepatic α1-AT mRNA and protein expression also were increased in male PiZ mice, suggesting that greater hepatocellular proliferation and caspase activation in males results from increased hepatotoxicity associated with greater intracellular α1-ATZ accumulation. Testosterone treatment of female PiZ mice increased α1-ATZ expression and hepatocellular proliferation to a level comparable with that in males. In PiZ mice, hepatocytes devoid of intracellular α1-AT globules had a proliferative advantage compared with globule-containing hepatocytes. However, this advantage is relative because both globule-containing and globule-devoid hepatocytes exhibited comparable proliferation after partial hepatectomy. In conclusion, these data indicate that intracellular retention of mutant α1-ATZ is associated with a regenerative stimulus leading to increased hepatocellular proliferation, that gender-specific signals influence the degree of α1-AT expression and associated hepatic injury, and that hepatocytes devoid of α1-ATZ have a proliferative advantage over cells that accumulate the mutant protein. This selective proliferation suggests that hepatocellular transplantation may be applicable for treatment of this and other slowly progressive metabolic liver diseases (Hepatology 2004;39:1048-1055.)
Variation of hepatic glucuronidation: Novel functional polymorphisms of the UDP-glucuronosyltransferase UGT1A4
Authors: Ehmer Ursula , Vogel Arndt , Schütte Jan Karl , Krone Britta , Manns Michael P. , Strassburg Christian P.
Published: March 2004

UDP-glucuronosyltransferases are a family of drug metabolizing enzymes contributing to hepatic drug metabolism and protection against environmental toxins. The aim of this study was to identify polymorphisms at the human UGT1A gene locus and to characterize their function and potential association with hepatocellular carcinoma (HCC). Genomic DNA from the blood of 363 subjects (128 patients with HCC, 235 blood donors) was analyzed for polymorphisms of the UGT1A3, UGT1A4, UGT1A8, UGT1A9, UGT1A10 genes using polymerase chain reaction, sequencing analysis. Recombinant variant UGT protein was analyzed by activity assays. In the UGT1A8 gene an A173G variant and a conserved G to A exchange at position 765 were detected in 25% and 15%. UGT1A9 exhibited two variants C3Y and M33T in 1% and 3%. UGT1A10 exhibited conserved nucleotide exchanges (128 G→A and 696 C→T) in 2% and 13%. In the UGT1A3 gene a W11R, a V47A variant, and a conserved G to A exchange at position 81 with an incidence of 65%, 58%, and 65%, respectively, were identified. UGT1A4 exhibited a P24T and an L48V variant in 8% and 9%. UGT1A SNPs were not associated with HCC. UGT1A4 P24T and L48V exhibited reduced glucuronidation activities: β-naphthylamine 30% and 50%, and dihydrotestosterone 50% and 0%, respectively. In conclusion, the high prevalence of SNPs throughout the human UGT1A gene locus illustrates a genetic basis of interindividual variations of hepatic metabolism. Two polymorphisms of the hepatic UGT1A4 protein show a differential metabolic activity toward mutagenic amines and endogenous steroids, altering hepatic metabolism and detoxification. (Hepatology 2004;39:970-977.)
Hepatitis C virus-infected patients report communication problems with physicians
Authors: Zickmund Susan , Hillis Stephen L. , Barnett Mitchell J. , Ippolito Laura , LaBrecque Douglas R.
Published: March 2004

We examined the prevalence and nature of perceived problems in the interaction between physicians and patients diagnosed with hepatitis C virus (HCV) infection. This cross-sectional study included 322 outpatients diagnosed with chronic HCV infection and treated at a tertiary referral hospital's hepatology clinic. Patients were asked to provide demographic information and to complete a semistructured interview, the Sickness Impact Profile (SIP) and Hospital Anxiety Depression (HAD) scale. A team of two blinded coders analyzed the interviews. A total of 131 (41%) study patients reported communication difficulties with physicians involved in their care. The main difficulties were the poor communication skills of physicians (91 [28%]), physician incompetence regarding the diagnosis and treatment of HCV infection (74 [23%]), feelings of being misdiagnosed, misled, or abandoned (51 [16%]), or being stigmatized by their physician (29 [9%]). Patients were twice as likely to report difficulties with subspecialists as compared with generalists. Nonresponse with antiviral therapy correlated with perceived physician conflict even after adjusting for treatment in relation to the time of interview, whereas previous or ongoing substance abuse and mode of acquisition did not. In a multivariate model, patients' psychosocial problems were the best predictors of communication difficulties. In conclusion, a substantial number of patients with HCV infection report difficulties when interacting with physicians, which may be due to coexisting emotional or social problems. However, perceived stigmatization by physicians and a sense of abandonment reflect the need for further educational efforts. These should target both specialists and primary care providers to inform them about the psychosocial challenges facing these patients. (Hepatology 2004;39:999-1007.)
Host factors and failure of interferon-α treatment in hepatitis C virus
Authors: Gao Bin , Hong Feng , Radaeva Svetlana
Published: March 2004

Failure of interferon-α (IFN-α) treatment in patients with chronic hepatitis C virus (HCV) infection is a challenging obstacle for clinical and experimental hepatology. Both viral and host factors have been implicated in reducing responsiveness to IFN-α therapy. The role of viral factors has been studied extensively and has been summarized in several review articles; however, much less attention has been paid to host factors. In this paper, we review evidence of host factor involvement in IFN-α treatment failure. We discuss possible underlying mechanisms responsible for these effects. Potential therapeutic strategies to enhance the effectiveness of IFN-α therapy for HCV are also proposed. (Hepatology 2004;39:880ndash;890.)
Abnormal hepatocystin caused by truncating PRKCSH mutations leads to autosomal dominant polycystic liver disease
Authors: Drenth Joost P. H. , Tahvanainen Esa , te Morsche Rene H. M. , Tahvanainen Pia , Kääriäinen Helena , Höckerstedt Krister , van de Kamp Jiddeke M. , Breuning Martijn H. , Jansen Jan B. M. J.
Published: March 2004

Mutations in protein kinase C substrate 80K-H (PRKCSH), encoding for the protein hepatocystin, cause autosomal dominant polycystic liver disease (PCLD), which is clinically characterized by the presence of multiple liver cysts. PCLD has been documented in families from Europe (Netherlands, Belgium, Finland) as well as from the United States. In this article, we report results from extensive mutational analysis of the PRKCSH gene in a group of 14 PCLD families and 65 singleton cases of Dutch and Finnish descent with multiple simple liver cysts. We identified PRKCSH mutations in 12 families and in 3 sporadic cases. In 8 of 10 Finnish families we detected the 1437+2delTG splice-site mutation. In Dutch families, we found 2 other mutations that affect correct splicing of PRKCSH: 292+1 G>C (2 families) and 1338-2 A>G (1 family). In another Dutch family, we detected a novel deletion (374-375delAG) in exon 6, predicting an abnormal shortened protein. Investigation of the carrier haplotypes identified a common founder chromosome in unrelated individuals in each of the 3 identified splice-site mutations. In 2 Finnish families with dominantly inherited PCLD, and in 62 of 65 sporadic cases with multiple simple liver cysts, we failed to demonstrate any PRKCSH mutation. This corroborates the notion that autosomal dominant PCLD is genetically heterogeneous. In conclusion, we propose that, on the basis of our results, genetic screening for PRKCSH gene mutations should be limited to patients either with a positive family history for PCLD or who have severe polycystic liver disease. (Hepatology 2004;39:924-931.)
HLA and cytokine gene polymorphisms are independently associated with responses to hepatitis B vaccination
Authors: Wang Chengbin , Tang Jianming , Song Wei , Lobashevsky Elena , Wilson Craig M. , Kaslow Richard A.
Published: March 2004

Variable immune responses to hepatitis B virus (HBV) infection and recombinant HBV vaccines have been associated with polymorphisms in several genes within the human leukocyte antigen (HLA) complex. Analyses of polymerase chain reaction (PCR)-based genotyping data from 164 North American adolescents vaccinated with recombinant HBV products confirmed that HLA-DRB1*07 (relative odds [RO] = 5.18, P < .0001) and human immunodeficiency virus type 1 (HIV-1) infection (RO = 3.91, P < .001) were both associated with nonresponse to full-dose vaccination. Further associations were observed with single nucleotide polymorphisms (SNPs) at the IL2 and IL4 loci along with insertion/deletion variants at the IL12B locus (P = .003-.01). Host genetic associations were independent of one another as well as other HLA (A, B, C, and DQB1) and cytokine gene (IL4R, IL6, IL10, and TNF) variants. Statistical adjustments for nongenetic factors (gender, ethnicity, age, HIV-1 infection, and vaccination protocols) did not substantially alter the strengths of the genetic relationships. The overall distribution pattern of genetic variations was similar between the analyzed vaccinees and additional adolescents (n = 292) from the same cohort. In conclusion, DRB1*07 (or a closely linked allele) and immunoregulatory cytokine gene polymorphisms correlate with variable immune response to recombinant HBV vaccines. (Hepatology 2004;39:978-988.)
Increased vascular heme oxygenase-1 expression contributes to arterial vasodilation in experimental cirrhosis in rats
Authors: Chen Yung-Chang , Ginès Pere , Yang Jianhui , Summer Sandra N. , Falk Sandor , Russell Nash S. , Schrier Robert W.
Published: March 2004

Vascular heme oxygenase (HO) regulates vascular tone in normal conditions and in some pathologic circumstances (e.g., sepsis). However, its possible role in the pathogenesis of arterial vasodilation in cirrhosis is unknown. To address this question, the expression and activity of HO in arterial vessels was studied in rats at 1, 2, and 4 weeks after bile duct ligation (BDL) or sham operation. A progressively increased expression of HO-1 was found in aorta and mesenteric arteries of BDL rats in a close chronologic relationship with the progression from acute cholestatic liver injury (1 week) to the fully developed cirrhosis with intense systemic arterial vasodilation (4 weeks). No changes were found in the expression of the constitutive isoform HO-2. HO-1 was mainly located in vascular smooth muscle cells of the arterial wall. Aortic HO activity increased in parallel with the expression of HO-1 (up to 600% in rats with cirrhosis compared with sham rats) and correlated with hemodynamic parameters. Increased expression of HO-1 and HO activity were also found in other organs, such as liver and spleen, though to a lesser extent compared with vascular tissue. The acute administration of an inhibitor of HO to cirrhotic rats, at a dose that normalized aortic HO activity, was associated with significantly greater effects on arterial pressure, total peripheral vascular resistance, and cardiac index, compared with effects in sham rats. In conclusion, these findings are consistent with a role for HO in the pathogenesis of arterial vasodilation in cirrhosis. (Hepatology 2004;39:1075-1087.)
Prognostic value of altered oral glutamine challenge in patients with minimal hepatic encephalopathy
Authors: Romero-Gómez Manuel , Grande Lourdes , Camacho Inés
Published: March 2004

Oral glutamine challenge (OGC) has been found to be safe, and an altered response predicts elevated risk of overt hepatic encephalopathy (HE) in patients with minimal hepatic encephalopathy (MHE). We assessed the survival prognosis of patients with cirrhosis, but without current overt HE, who have an altered OGC and MHE. MHE was inferred using 3 neuropsychological tests. Venous ammonia concentrations were measured pre- and post-60 minutes of a 10 g oral glutamine load. The median follow-up was 25.2 months, by which time 22 patients had had bouts of overt HE and 18 had died from liver-related causes. The results in 126 patients with cirrhosis, indicated 25 with MHE and abnormal OGC response. Survival among patients who developed overt HE was 59% at 1 year and 38% at 3 years. In patients without HE, survival was 96% and 86% at 1 and 3 years, respectively (log-rank 50.9, P < .0001). The presence of MHE was not related to survival (log-rank 2.21, P = .23). Patients with MHE and abnormal OGC test had elevated mortality risk (log-rank 13.1, P = .0003). Multivariate analyses indicated Child-Pugh score (hazard ratio [HR] 1.46; 95% CI, 1.46-2.08), and MHE plus altered OGC response (HR 5.5; 95% CI, 1.81-16.6) were predictors of mortality, whether from liver-related or non-liver-related causes. In conclusion, a pathological OGC response in patients with MHE appears to be associated with lower survival rate and may prove useful in the selection of candidates for liver transplantation. (Hepatology 2004;39:939-943.)
Kupffer cells required for high affinity peptide-induced deletion, not retention, of activated CD8+ T cells by mouse liver
Authors: Kuniyasu Yuhshi , Marfani Suhail Mohammed , Inayat Irteza Bin , Sheikh Shehzad Zafar , Mehal Wajahat Zafar
Published: March 2004

The immune response to foreign antigens in the liver is often suboptimal and this is clinically relevant in chronic persistence of hepatotropic viruses. In chronic infection with the hepatitis C virus, activated CD8+ T cells specific for viral epitopes are present in the peripheral blood and the liver, yet viral clearance is unusual. To define the fate of activated CD8+ entering the liver, we developed a mouse model of portal vein injection of activated CD8+ T cells in vivo. Activated CD8+ T cells are retained very efficiently by the liver and undergo an approximately 8-fold expansion in the first 48 hours. This expansion is followed by apoptosis and a decline in numbers of the retained cells over the next 4 days. The presence of high affinity (HA) antigen does not affect the initial retention by the liver but greatly limits the expansion in the first 48 hours by increasing apoptosis of the retained cells. In the absence of Kupffer cells, the initial retention and expansion are unchanged, but HA antigen does not limit the expansion of the liver CD8+ T cell pool. In conclusion, these data identify a previously unknown phase of CD8+ T cell expansion after entering the liver, demonstrate that HA antigen limits the hepatic CD8+ T cell pool by inducing apoptosis, and that this effect requires Kupffer cells. Interfering with antigen presentation by Kupffer cells may be a strategy to limit HA antigen-induced deletion of activated CD8+ T cells entering the liver. (Hepatology 2004;39;1017-1027.)
5′-methylthioadenosine modulates the inflammatory response to endotoxin in mice and in rat hepatocytes†
Authors: Hevia Henar , Varela-Rey Marta , Corrales Fernando J. , Berasain Carmen , Martínez-Chantar María L. , Latasa Ujue M. , Lu Shelly C. , Mato José M. , García-Trevijano Elena R. , Avila Matías A.
Published: March 2004

5′-methylthioadenosine (MTA) is a nucleoside generated from S-adenosylmethionine (AdoMet) during polyamine synthesis. Recent evidence indicates that AdoMet modulates in vivo the production of inflammatory mediators. We have evaluated the anti-inflammatory properties of MTA in bacterial lipopolysaccharide (LPS) challenged mice, murine macrophage RAW 264.7 cells, and isolated rat hepatocytes treated with pro-inflammatory cytokines. MTA administration completely prevented LPS-induced lethality. The life-sparing effect of MTA was accompanied by the suppression of circulating tumor necrosis factor-α (TNF-α), inducible NO synthase (iNOS) expression, and by the stimulation of IL-10 synthesis. These responses to MTA were also observed in LPS-treated RAW 264.7 cells. MTA prevented the transcriptional activation of iNOS by pro-inflammatory cytokines in isolated hepatocytes, and the induction of cyclooxygenase 2 (COX2) in RAW 264.7 cells. MTA inhibited the activation of p38 mitogen-activated protein kinase (MAPK), c-jun phosphorylation, inhibitor kappa B alpha (IκBα) degradation, and nuclear factor κB (NFκB) activation, all of which are signaling pathways related to the generation of inflammatory mediators. These effects were independent of the metabolic conversion of MTA into AdoMet and the potential interaction of MTA with the cAMP signaling pathway, central to the anti-inflammatory actions of its structural analog adenosine. In conclusion, these observations demonstrate novel immunomodulatory properties for MTA that may be of value in the management of inflammatory diseases. (Hepatology 2004;39:1088-1098.)
Progression of HCC in mice is associated with a downregulation in the expression of hepatocyte nuclear factors
Authors: Lazarevich Natalia L. , Cheremnova Oksana A. , Varga Ekaterina V. , Ovchinnikov Dmitry A. , Kudrjavtseva Elena I. , Morozova Olga V. , Fleishman Daria I. , Engelhardt Natalia V. , Duncan Stephen A.
Published: March 2004

Hepatocyte nuclear factors (HNF) play a critical role in development of the liver. Their roles during liver tumorigenesis and progression of hepatocellular carcinomas (HCC) are, however, poorly understood. To address the role of HNFs in tumor progression, we generated a new experimental model in which a highly differentiated slow-growing transplantable mouse HCC (sgHCC) rapidly gives rise in vivo to a highly invasive fast-growing dedifferentiated variant (fgHCC). This in vivo model has allowed us to investigate the fundamental mechanisms underlying HCC progression. A complete loss of cell polarity, a decrease in cell-cell and cell-extracellular matrix (ECM) adhesion, elevation of telomerase activity, and extinction of liver-specific gene expression accompanies tumor progression. Moreover, cells isolated from fgHCCs acquired the ability to proliferate rapidly in culture. These alterations were coupled with a reduced expression of several liver transcription factors including HNF4, a factor essential for hepatocyte differentiation. Forced re-expression of HNF4α1 in cultured fgHCC cells reversed the progressive phenotype and induced fgHCC cells to re-establish an epithelium and reform cell-ECM contacts. Moreover, fgHCC cells that expressed HNF4α1 also re-established expression of the profile of liver transcription factors and hepatic genes that are associated with a differentiated hepatocyte phenotype. Importantly, re-expression of HNF4α1 in fgHCC reduced the proliferation rate in vitro and diminished tumor formation in congenic recipient mice. In conclusion, loss of HNF4 expression is an important determinant of HCC progression. Forced expression of this factor can promote reversion of tumors toward a less invasive highly differentiated slow-growing phenotype. (Hepatology 2004;39;1038-1047.)
Sinusoidal endothelial cell and hepatocyte death following cold ischemia-warm reperfusion of the rat liver
Authors: Huet Pierre-Michel , Nagaoka Marcia R. , Desbiens Geneviève , Tarrab Esther , Brault Antoine , Bralet Marie-Pierre , Bilodeau Marc
Published: March 2004

Cold ischemia-warm reperfusion (CI-WR) injury of the liver is characterized by marked alterations of sinusoidal endothelial cells (SECs), whereas hepatocytes appear to be relatively unscathed. However, the time course and mechanism of cell death remain controversial: early versus late phenomenon, necrosis versus apoptosis? We describe the occurrence and nature of cell death after different periods of CI with University of Wisconsin (UW) solution and after different periods of WR in the isolated perfused rat liver model. After 24- and 42-hour CI (viable and nonviable livers, respectively), similar patterns of liver cell death were seen: SEC necrosis appeared early after WR (10 minutes) and remained stable for up to 120 minutes. After 30 minutes of WR, apoptosis increased progressively with WR length. Based on morphological criteria, apoptotic cells were mainly hepatocytes within liver plates or extruded in the sinusoidal lumen. In addition, only after 42-hour CI were large clusters of necrotic hepatocytes found in areas of congested sinusoids. In these same livers, the hepatic microcirculation, evaluated by means of the multiple-indicator dilution technique, revealed extracellular matrix disappearance with no-flow areas. In conclusion, different time courses and mechanisms of cell death occur in rat livers after CI-WR, with early SEC necrosis followed by delayed hepatocyte apoptosis. These processes do not appear to be of major importance in the mechanism of graft failure because they are similar under both nonlethal and lethal conditions; this is not the case for the loss of the extracellular matrix found only under lethal conditions and associated with hepatocyte necrosis. (Hepatology 2004;39:1110-1119.)
Human resource management problems over the life cycle of small to medium‐sized firms
Authors: Rutherford Matthew W. , Buller Paul F. , McMullen Patrick R.
Published: March 2004

This study uses a sample of 2,903 small to medium‐sized firms to examine the manner in which HR problems
vary over the organizational life cycle. We found that a four‐stage model was appropriate. Interestingly,
firm age did not emerge as a significant indicator of stage—the firms' HR problems varied across stages
defined by growth. Training problems were highest in high‐growth firms and lowest in low‐growth
firms; compensation problems were highest in moderate‐growth firms and lowest in high‐growth
firms; and recruiting problems were highest in no‐growth firms and lowest in low‐growth firms.
© 2004 Wiley Periodicals, Inc.
The adoption of high‐involvement practices and processes in emergent and developing firms: A descriptive and prescriptive approach
Authors: Ciavarella Mark A.
Published: March 2004

One of the most pressing challenges entrepreneurs face is how to do more with less. More often than not, the
solution lies in the human resource component of the venture. As such, it is important that entrepreneurs
understand how practices and processes designed to tap the knowledge, skills, and abilities of their employees can
lead to greater productivity and performance. This article develops prescriptive arguments to implicate that
adoption of involvement practices and processes by entrepreneurs early in the organization's life may extend
desired stages such as growth, or may delay or negate unwanted stages such as maturity and decline.
© 2004 Wiley Periodicals, Inc.
Strategic human capital management in SMEs: An empirical study of entrepreneurial performance
Authors: Hayton James C.
Published: March 2004

How can human resource management (HRM) practices promote entrepreneurial performance in small and
medium‐sized enterprises (SMEs)? This article discusses the association between human
capital management (HCM) and other contemporary HRM practices and the ability of SMEs to be
entrepreneurial. In a study of 99 SMEs, HRM practices that promote employee discretionary behavior, knowledge
sharing, and organizational learning are found to be positively associated with entrepreneurial performance.
Two contingencies are also identified for this relationship. First, the use of strategic HCM practices enhances
the observed positive association. Second, these relationships are strongest for SMEs operating in
high‐technology industries. © 2004 Wiley Periodicals, Inc.
Dietary carotenoids and risk of colon cancer: Case‐control study
Authors: Nkondjock André , Ghadirian Parviz
Published: March 2004

Some epidemiological studies suggest that consumption of fruits and vegetables with a high carotenoid content may protect against colon cancer (CC). The evidence, however, is not completely consistent. Given the inconsistencies in findings in previous studies and continued interest in identifying modifiable risk factors for CC, a case‐control study of French‐Canadian in Montreal, Canada, was undertaken to examine the possible association between dietary carotenoids and CC risk and to investigate whether this association varies in relation to lifestyle factors such as smoking or diet, and particularly the high consumption of long‐chain polyunsaturated fatty acids (LCPUFA). A total of 402 colorectal cases (200 males and 202 females) and 688 population‐based controls matched for age, gender and place of residence were interviewed. Dietary intake was assessed through a validated food frequency questionnaire that collected information on over 200 food items and recipes. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated in unconditional logistic regression models. After adjustment for important variables such as total energy intake, no association was found between dietary intake of carotenoids and CC risk. For women with high intakes of LCPUFA, an inverse association was found between lutein + zeaxanthin and CC risk. ORs were 0.41; 95%CI (0.19–0.91), p=0.03 for eicosapentaenoic acid, and OR=0.36, 95%CI (0.19–0.78), p=0.01 for docosahexaenoic acid, when the upper quartiles of intake were compared to the lower. Among never‐smokers, a significantly reduced risk of CC was associated with intake of β‐carotene [OR=0.44, 95%CI (0.21–0.92) and p=0.02], whereas an inverse association was found between lycopene intake and CC risk [OR=0.63, 95%CI (0.40–0.98) and p=0.05] among smokers. The results of our study suggest that a diet rich in both lutein + zeaxanthin and LCPUFAs may help prevent CC in French‐Canadian females. © 2004 Wiley‐Liss, Inc.
Study of puupehenone and related compounds as inhibitors of angiogenesis
Authors: Castro M. Estrella , González‐Iriarte Mauricio , Barrero Alejandro F. , Salvador‐Tormo Nélida , Muñoz‐Chápuli Ramón , Medina Miguel Ángel , Quesada Ana R.
Published: March 2004

Puupehenone, a sesquiterpene produced by certain sponges, was selected in the course of a blind screening for new potential inhibitors of angiogenesis. In our study, we compare the potential anti‐angiogenic activities of puupehenone and another 11 related compounds that were either natural products from marine origin or their synthetic derivatives. The effects of these compounds were determined with cell growth and differentiation assays on bovine aorta endothelial cells. Our results show that these compounds are weak inhibitors to cell growth and are not selective for endothelial cells. However, contrary to cell growth, the differentiation of endothelial cells into tubular structures was completely inhibited by 7 of these compounds at concentrations equal or lower than 3 μM. Three of these compounds, isozonarol, 8‐epipuupehedione and 8 epi‐9,11‐dihydropuupehedione, completely inhibited the in vivo angiogenesis in the CAM assay at doses equal or lower than 30 nmol/egg. Further characterisation showed that these 3 terpenes also inhibited endothelial cell production of urokinase and invasion. One compound (8‐epipuupehedione) inhibited endothelial cell migration in a dose‐dependent manner. The anti‐angiogenic properties of the selected compounds, the simplicity of their structures and the feasibility of their synthesis make them attractive drugs for further evaluation in the treatment of angiogenesis‐related pathologies. © 2004 Wiley‐Liss, Inc.
Impact of reproductive factors and lactation on breast carcinoma in situ risk
Authors: Meeske Kathleen , Press Michael , Patel Alpa , Bernstein Leslie
Published: March 2004

Incidence rates for breast carcinoma in situ (CIS) have increased markedly over the past 20 years. Breast CIS, detected primarily on mammography, now represents 30–45% of all screened detected breast cancers. We conducted a large population‐based case‐control study to evaluate the impact of reproductive factors and lactation on breast CIS risk. Case subjects were newly diagnosed with breast CIS at ages 35–64 years between March 1, 1995 and May 31, 1998 (n = 567), resided in Los Angeles County and were born in the United States. Control subjects (n = 614), identified through random digit dialing, fulfilled the same eligibility criteria and were required to have had at least one screening mammogram in the 2‐year period before their interview. Women with a positive family history of breast cancer had a 2‐fold increase in breast CIS risk. Parous women were at reduced risk relative to nulligravid women (odds ratio [OR] = 0.67, 95% confidence interval [CI] = 0.46–1.00). Among nulliparous women, pregnancy was unrelated to breast CIS risk. Among parous women, risk declined with each additional term pregnancy (p‐trend = 0.003). No associations were found with age at first term pregnancy, induced abortion or miscarriage. Long duration of breast‐feeding (≥24 months) was associated with increased risk (OR = 2.00, 95% CI = 1.11–3.60). The observed effects of family history and pregnancy on breast CIS risk are consistent with those observed for invasive breast cancer. The results for breast‐feeding are contrary to what has been observed in studies of invasive breast cancer. © 2004 Wiley‐Liss, Inc.
POBASCAM, a population‐based randomized controlled trial for implementation of high‐risk HPV testing in cervical screening: Design, methods and baseline data of 44,102 women
Authors: Bulkmans Nicole W.J. , Rozendaal Lawrence , Snijders Peter J.F. , Voorhorst Feja J. , Boeke A. Joan P. , Zandwijken Gladys R.J. , van Kemenade Folkert J. , Verheijen René H.M. , v Groningen Krijn , Boon Mathilde E. , Keuning Hans J.F. , van Ballegooijen Marjolein , van den Brule Adriaan J.C. , Meijer Chris J.L.M.
Published: March 2004

Cytological cervical screening is rather inefficient because of relatively high proportions of false negative and false positive smears. To evaluate the efficiency of high‐risk human papillomavirus (hrHPV) testing, by GP5+/6+ PCR‐enzyme immunoassay (EIA), in conjunction with cytology (Intervention Group) to that of the classical cytology (Control Group), we initiated the Population Based Screening Study Amsterdam (POBASCAM). POBASCAM is a population‐based randomized controlled trial for implementation of hrHPV testing in cervical screening. The outcome measure is the proportion of histologically confirmed ≥CIN3 lesions in each study arm up to and including the next screening round after 5 years. We present the design, methods and baseline data of POBASCAM. When, in the next 5 years, the follow‐up will be completed, the data obtained will be used in model studies, including a cost‐effectiveness study, to advise the Dutch Ministry of Public Health in deciding whether cervical screening should be based on combined hrHPV and cytology testing instead of cytology alone. Between January 1999 and September 2002, 44,102 women (mean age = 42.8 years; range = 29–61) that participated in the regular Dutch screening program were included in our study. In the Intervention Group the distribution of cytology and hrHPV by cytology class was as follows: normal cytology 96.6% (3.6% hrHPV positive); borderline and mild dyskaryosis (BMD) 2.5% (34.6% hrHPV positive); and moderate dyskaryosis or worse (>BMD) 0.8% (88.3% hrHPV positive), i.e., 0.4% moderate dyskaryosis (82.9% hrHPV positive), 0.3% severe dyskaryosis (92.5% hrHPV positive), 0.1% carcinoma in situ (95.2% hrHPV positive), <0.1% suspected for invasive cancer (hrHPV positive 100.0%). In the Control Group 96.5% of the women had normal cytology, 2.4% BMD and 0.8% >BMD, i.e., 0.4% moderate dyskaryosis, 0.3% severe dyskaryosis, 0.1% carcinoma in situ, <0.1% suspected for invasive cancer. The presence of hrHPV was age‐dependent, decreasing from 12.0% at 29–33 years to 2.4% at 59–61 years. Among women with a positive hrHPV test, the prevalence of BMD was age‐dependent ranging from 20.2% at 29–33 years to 7.8% at 54–58 years. In contrast, the risk of >BMD of 13.7% among women with a positive hrHPV test was not age‐dependent. Our study indicates that large‐scale hrHPV testing by GP5+/6+ PCR‐EIA in the setting of population‐based cervical screening is practically feasible, is accepted by both participating women and general practitioners and yields highly reproducible results. © 2004 Wiley‐Liss, Inc.
Antisense oligodeoxynucleotides for urokinase‐plasminogen activator receptor have anti‐invasive and anti‐proliferative effects in vitro and inhibit spontaneous metastases of human melanoma in mice
Authors: D'Alessio Silvia , Margheri Francesca , Pucci Marco , Del Rosso Angela , Monia Brett P. , Bologna Mauro , Leonetti Carlo , Scarsella Marco , Zupi Gabriella , Fibbi Gabriella , Del Rosso Mario
Published: March 2004

We have targeted the urokinase‐type plasminogen activator receptor (uPAR) with phosphorothioate antisense oligonucleotides (aODN) in vitro to evaluate the anti‐invasive and anti‐proliferative effects of uPAR down‐regulation, as well as in vivo to evaluate anti‐tumor and anti‐metastatic activity. aODN‐dependent uPAR downregulation in vitro was induced in cells of human melanoma, mammary carcinoma, ovarian carcinoma and SV‐40‐transformed embryonic lung fibroblasts. uPAR was determined by an antibody‐based assay and by semiquantitative polymerase chain reaction. Cell invasion was evaluated by Matrigel invasion assay and cell proliferation by direct cell counting. aODN reduced uPAR, invasion and proliferation in all the treated cell lines. Following aODN treatment, human melanoma cells exhibited a strong decrease of uPAR‐dependent ERK1/2 activation and were used in vivo to control metastasis in CD‐1 male nude (nu/nu) mice by uPAR aODN injection. 60 mice were injected in the hind leg muscles with a suspension of 106 melanoma cells. After 4 days, when a tumor mass of about 350 mg was evident in all the mice injected, 20 mice were treated i.v. with aODN and 20 with dODN at 0.5 mg/day for 5 consecutive days. Twenty control mice were not treated. A second and third cycle of treatment was administered at 2‐day intervals. Treatment with aODN resulted into a 78% reduction of lung metastases and 45% reduction of the primary tumor mass with no loss of body weight. Our results suggest to evaluate the utility of uPAR aODN in controlling the metastatic spreading of human melanoma. © 2004 Wiley‐Liss, Inc.
Interferon resistance promotes oncolysis by influenza virus NS1‐deletion mutants
Authors: Muster Thomas , Rajtarova Julia , Sachet Monika , Unger Hermann , Fleischhacker Reinhard , Romirer Ingrid , Grassauer Andreas , Url Angelika , García‐Sastre Adolfo , Wolff Klaus , Pehamberger Hubert , Bergmann Michael
Published: March 2004

NS1 protein of influenza virus is a virulence factor that counteracts Type I interferon (IFN)‐mediated antiviral response by the host. A recombinant influenza A virus that lacks the NS1 protein only replicates efficiently in systems that contain defective IFN pathways. We demonstrate that the conditional replication properties of NS1‐modified influenza A virus mutants can be exploited for the virus‐mediated oncolysis of IFN‐resistant tumor cells. IFN resistance in analyzed tumor cell lines correlated with a reduced expression of STAT1. Addition of exogenous IFNα or supernatant of virus‐infected endothelial cells inhibited viral oncolysis in IFN‐sensitive but not in IFN‐resistant cell lines. The oncolytic potential of NS1‐modified influenza A virus mutants could be exploited in vivo in a SCID mouse model of a subcutaneously‐implanted human IFN‐resistant melanoma. The data indicate that IFN‐resistant tumors are a suitable target for oncolysis induced by NS1‐modified influenza virus mutants. STAT1 might serve as a marker to identify these IFN‐resistant tumors. © 2004 Wiley‐Liss, Inc.
Neovascular targeting chemotherapy: Encapsulation of paclitaxel in cationic liposomes impairs functional tumor microvasculature
Authors: Strieth Sebastian , Eichhorn Martin E. , Sauer Birgitta , Schulze Brita , Teifel Michael , Michaelis Uwe , Dellian Marc
Published: March 2004

Cationic liposomes have been shown to be internalized selectively by angiogenic tumor endothelial cells after intravenous injection. Therefore, encapsulation of cytotoxic substances in cationic liposomes is a new approach to target tumor vasculature. It was the aim of our study to quantify the effects of paclitaxel encapsulated in cationic liposomes (MBT‐0206) on tumor microvasculature and growth in vivo. Experiments were performed in the dorsal skinfold chamber preparation of Syrian Golden hamsters bearing syngeneic A‐Mel‐3 melanomas. Tumors were treated with intravenous infusion of MBT‐0206 (20 mM) resulting in an effective paclitaxel dose of 5 mg/kg body weight (b.w.). Control animals received conventional paclitaxel in Cremophor EL (Taxol®; 5 mg/kg b.w.), unloaded cationic liposomes (20 mM) or the solvent 5% glucose, respectively. Using intravital microscopy, tumor growth and effects on intratumoral microvasculature were analyzed. Tumor growth was significantly retarded after treatment with MBT‐0206 compared to the treatment with paclitaxel. Analysis of intratumoral microcirculation revealed a reduced functional vessel density in tumors after application of liposomal paclitaxel. At the end of the observation time, vessel diameters were significantly smaller in animals treated with paclitaxel encapsulated in cationic liposomes while red blood cell velocity was less affected. This resulted in a significantly reduced blood flow in vessel segments and a reduced microcirculatory perfusion index in these animals. Histochemical TUNEL stain was vessel‐associated after treatment with liposomal paclitaxel in contrast to few apoptotic tumor cells in the control groups. Our data demonstrate that encapsulation of paclitaxel in cationic liposomes significantly increased the antitumoral efficacy of the drug. Remarkable microcirculatory changes indicate that encapsulation of paclitaxel in cationic liposomes resulted in a mechanistic switch from tumor cell toxicity to an antivascular therapy. © 2004 Wiley‐Liss, Inc.
B7‐2 expression above a threshold elicits anti‐tumor immunity as effective as interleukin‐12 and prolongs survival in murine B‐cell lymphoma
Authors: Pizzoferrato Eva
Published: March 2004

The costimulatory molecule, B7‐2, is expressed by various lymphomas, but this level of expression is not sufficient to generate effective anti‐tumor immunity in vivo. To determine whether up‐regulated expression of the costimulatory molecule, B7‐2, leads to more effective anti‐tumor immunity in vivo, the A20 murine model of B‐cell lymphoma was used. A20 tumor cells express major histocompatibility complex (MHC) I and II molecules and moderate constitutive levels of B7‐2. While B7‐1 and B7‐2 have been introduced into tumor cells lacking these molecules, studies have not been conducted to determine whether tumors that constitutively express B7‐1 or B7‐2 can be made more immunogenic by increasing the expression of these molecules. In this report, A20/B7‐2 transfectants expressing greater levels of B7‐2 were rejected in syngeneic mice, and systemic immunity against the A20 parental cells was generated. Treatment with the A20/B7‐2 variant cells significantly improved the survival of tumor‐bearing mice. Coinjection with IL‐12 secreting variants did not further augment the anti‐tumor immunity observed for B7‐2 therapy alone. Both CD8+ T cells and natural killer (NK) cells mediated the anti‐tumor immune response observed in A20/B7‐2 immunized mice. In mice that developed tumors after immunization with the A20/B7‐2 variant cells, resected tumor cells were shown to express lower levels of B7‐2 than the transfected variants. These results suggest that the level of costimulation is important for the generation of anti‐tumor immunity and for host survival. In addition, tumors appear to be able to evade the immune response by downregulating the expression of B7‐2 below a threshold level. © 2004 Wiley‐Liss, Inc.
Olf/EBF proteins are expressed in neuroblastoma cells: Potential regulators of the Chromogranin A and SCG10 promoters
Authors: Persson Paula , Manetopoulos Christina , Lagergren Anna , Nygren Jens , Gisler Ramiro , Axelson Håkan , Sigvardsson Mikael
Published: March 2004

The childhood malignancy neuroblastoma is derived from developmentally arrested sympathetic nervous system precursor cells. To obtain further insight into the molecular processes involved in the formation of these tumors, we decided to investigate the functional role of Olf/EBF (O/E) transcription factors in human neuroblastoma cells. We here report that O/E‐1 and O/E‐2 are expressed at variable levels in neuroblastoma cell lines and that O/E proteins could be identified by electrophoretic mobility shift assays. To identify potential neuronal target genes for O/E proteins in neuroblastoma cells we investigated the ability of a set of neuronal promoters to interact with O/E‐1 in electrophoretic mobility shift assays. This analysis suggested that the Chromogranin A (CgA) and SCG10 promoters both contained binding sites for O/E‐1. O/E‐1 was able to activate the CgA promoter in vivo and mutation of the O/E‐1 binding site in the CgA promoter reduced the functional activity of the element to about 60% of the wild‐type in neuroblastoma cells, supporting the idea that O/E proteins may be involved in the control of the CgA promoter. Furthermore, overexpression of O/E‐1 in hippocampal progenitor cells led to neurite outgrowth, indicative of a role for O/E proteins in neuronal differentiation. © 2004 Wiley‐Liss, Inc.
Explant pathology study of decellularized carotid artery vascular grafts
Authors: Hilbert Stephen L. , Boerboom Lawrence E. , Livesey Stephen A. , Ferrans Victor J.
Published: March 2004

The purpose of this study was to evaluate the morphologic findings in small‐diameter freeze‐dried decellularized carotid artery grafts implanted in goats as carotid artery interposition grafts for 6–7 months. Unimplanted decellularized carotid artery grafts did not contain intact cells; however, remnants of smooth muscle cells were present in the media. The extracellular matrix was well preserved. All decellularized grafts were patent at explant, without significant dimensional changes or aneurysm formation. Their luminal surfaces were lined by a thin neointima, consisting of myofibroblasts, collagen, and a discontinuous layer of endothelial cells. Histologic evidence of calcification within the explants was not observed; however, electron microscopy showed calcification of minute remnants of cell membranes. Inflammatory cells were not present in the graft wall. Host cell migration was greatest in the adventitia along the length of the graft. Migration of host cells into the media was more apparent close to the anastomoses, forming cellular nests rich in extracellular proteoglycans, whereas cell migration into areas subjacent to the lumen was minimal. Ingrowth of host blood vessels was not observed. These results demonstrate satisfactory structural and morphologic features of a decellularized carotid artery small‐diameter graft implanted for up to 7 months. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 69A: 197–204, 2004
Preformed grafts of porcine small intestine submucosa (SIS) for bridging segmental bone defects
Authors: Moore Douglas C. , Pedrozo Hugo A. , Crisco Joseph J. , Ehrlich Michael G.
Published: March 2004

Previous studies suggest that fresh, morselized porcine small intestine submucosa (SIS) may have promise in the treatment of large bone defects. This study evaluated the bone regenerative potential of preformed tubular SIS grafts, designed to provide a scaffold for regeneration of diaphyseal bone. Critical length segmental defects in the femurs of male rats were either left unfilled (n = 11) or filled with morselized cancellous bone (n = 12), or spanned with intramedullary tubes (n = 12) or periosteal sleeves (n = 12) fabricated from SIS. All of the animals were euthanized 12 weeks postoperatively. Healing was assessed with biweekly radiographs, routine histology, and mechanical testing. Copious new bone formed in the defects of all of the animals treated with cancellous bone; 10 of the 12 animals in that group had healed their defects. In contrast, no new bone was formed in the defects left unfilled or treated with SIS; only fibrous tissue was found. In both of the SIS‐treated groups, the SIS persisted at twelve weeks. The cellular response to the SIS involved a mild mononuclear infiltrate in the loose or delaminated superficial layers of the tubes and sleeves, with few cells in the deeper layers. The results of this study cast doubt on the ability of SIS to support or stimulate growth of bone across a critical length segmental bone defect. Additional work will be required to determine whether our results reflect the protocols used to prepare and fabricate the SIS grafts used in the study or the inherent inability of SIS to support new bone growth. © 2004 Wiley Periodicals, Inc. J Biomed Mater Res 69A: 259–266, 2004